Notes For Biology

lOMoARcPSD|1377855
Complete lecture notes for BIOL2016
Cell Biology (University of Sydney)
Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

lOMoARcPSD|1377855
How Cells Communicate

 Protein Kinase = signaling protein that phosphorylates proteins
 Multicellular organisms:
o Cells are held together by proteins and carbs
o Chemical signaling necessary between cells (eg: hormones)
Chemical Signaling
 Extracellular signal molecules bind to receptor molecules and initiate a response

in the target cell
 Receptors located throughout the cell, ready to pick up signals
 Signal passed through various intracellular signaling proteins, from receptor to
effector protein (destination)
 Proteins and enzymes activated have effect on cell
o Metabolic enzyme  altered cell metabolism
o Gene regulatory protein  altered gene expression
o Cytoskeletal protein  altered cell shape or movement
 Signal molecules:
o Hydrophilic = protein hormone (loves water)
o Hydrophobic = steroid hormone (hates water, loves oil)
– Have carrier proteins in blood- regulates activity
Signal molecules bind to specific receptors

 Cell-surface receptors:
o Located on the plasma membrane of cell (outside
surface)

lOMoARcPSD|1377855
o When bound to by a signal molecule, the receptor becomes activated and

generates various intracellular signals that alter the behavior of the cell
 Intracellular receptors:
o Located inside the target cell (eg: inside the nucleus)
o Signal molecule must enter the cell to bind to the receptor- must ∴ be small
and hydrophobic to diffuse through the membrane
NB: hydrophobic molecules require carriers to travel
Forms of Intercellular signaling

 Contact-dependent:
o Membrane bound signal molecule on signaling cell will interact with surface
receptor on target cell
o Cells must be in direct membrane-membrane contact
 Paracrine:
o Signaling cells secrete signal molecules into extracellular fluid
o Signal molecules may then act on distant target cells or act as local mediators
 Endocrine:
o Endocrine cells (glands with no duct) secrete signal molecules into the
bloodstream and interstitial fluid, allowing them to be carried to target cells
throughout the body
o Eg: the adrenaline system- releases flight or flight hormones to the muscle
and cardio tissues

lOMoARcPSD|1377855
 Synaptic:
o Synapse = the space between 2 cells (held together by extracellular matrix)
o Signaling performed by neurons (nerve cells) that transmit signals electrically
along their axons, neurotransmitters (chemical signals) are then released at
the synapse and delivered to the target cell
 One hormone can have different effects on different tissues- signal received by
different receptors, linked to different intercellular signaling systems
 One signal can have different speed responses in different pathways
o Signals that alter protein function = FAST (acute)
o Signals that alter protein synthesis = SLOW (prolonged/chronic)
 Gap junction = tube between 2 cells that allows small molecules to pass through
 Growth factor = signals that stimulate growth and survival- stimulate stem cells to
differentiate
 Apoptosis = signal stimulating cell suicide
 Morphogen = signal that effects development or differentiation
G Proteins and Second Messengers

 NB: in contrast to the hydrophobic signal molecules that bind to intracellular
receptors, most extracellular signal molecules bind to specific receptor proteins on
the surface of the target cell but do not enter the cytosol or nucleus
 Activation = the receptor is affected by the bound molecule in such a way as to elicit
tissue response
 Agonist = a molecule that, when bound to the receptor, causes activation
 Antagonist = a molecule that binds to the receptor, without causing activation and ∴
preventing the agonist from binding
The Plasma membrane

 Composed of a phospholipid bilayer
 Outside of membrane = hydrophilic (loves water)
 Hydrophilic messengers can NOT travel through membrane
(hydrophobic center) ∴ must interact with receptor in the
membrane
 Inside of membrane = hydrophobic (fears water)
G-protein-coupled receptors (GPCRs)

 Regulate the activity of a separate protein (usually an enzyme or ion channel)
 Located within the plasma membrane
 Consist of a single polypeptide chain that weaves 7 times through the plasma
membrane (7 pass sequence)

lOMoARcPSD|1377855
 Attached to a GTP binding protein, which mediates the interaction between activated
receptor and the target protein
GTP binding proteins

 An enzyme that converts GTP to GDP
 Trimeric = 3 subunits
o α subunit mw ~ 50 kda
o Usually located in the plasma membrane
 Monomeric = 1 subunit
o Mw ~ 30 kda
o Present in the cytosol
 Cycle between an active GTP-bound form and an inactive GDP-bound form to act as
switches in intra-molecular signaling pathways
 Inactive = GDP bound
 Active = GTP bound
The GPCR Signaling complex

 When a signal molecule binds to a GPCR, the receptor undergoes a conformation
change that activates the G protein, causing the α subunit to release
its bound GDP and bind GTP in its place
 The α-GTP complex separates from the beta-gamma unit and
diffuses freely through the membrane to the enzyme adenylyl
cyclase
 Adenylyl cyclase becomes activated when bound by α-GTP,
catalyzing the production of cyclic adenosine monophosphate
(cAMP) from ATP
 cAMP is synthesized from ATP through a cyclization reaction where
two phosphate groups are removed and a cyclic bond is formed in
their place
 cAMP then activates cAMP-dependent protein kinase (PKA)
o The binding of cAMP to the regulatory subunits of PKA
induces a conformation change that causes the dissociation
and hence activation of the catalytic subunits

lOMoARcPSD|1377855
o The activated cat-subunits can then phosphorylate specific target proteins

 Activated PKA is able to assist the transcription of target genes
o When PKA is activated by cAMP, it phosphorylates CREB on a single serine
o Phosphorylated CREB then recruits a transcriptional co-activator, CREB-
binding protein,
o Activated, phosphorylated CREB can now recognize the promotor region,
cAMP response element (CRE) and hence transcription can commence
How to detect new G Proteins in the cell- mitochondria

 Isolated mitochondria are heated with SDS and mercaptoethanol to cover the protein
with negative charge
 The sample is then run through electrophoresis gel to separate into bands
 Western blot used to migrate bands to membrane
 Membrane then exposed to radioactive GTP- bands that bond with be radioactive
 Radioactivity detected with photographic film to determine which bands bound to
GTP ∴ proving the presence of G proteins in mitochondria
Phospholipase and Calcium Signaling

Phospholipase pathway:
 Phospholipids:
o Hydrophilic sugar-phosphate head
(water loves charged phosphates)
o Hydrophobic fatty acid tails
 Endoplasmic reticulum (ER)
o Used for post-transcriptional
modification
o Stores and releases Ca2+
– Ca+ can bind to negative
protein to activate
 PLC = phospholipase C (cleaves lipids)

lOMoARcPSD|1377855
 Phospholipase pathway:
o Ligand stimulates surface receptor
o Receptor stimulates associated G protein, which activates enzyme PLC
o PLC cleaves IP3 off phospholipid (PIP2)- leaving behind DAG
o IP3 is a water-soluble, intracellular mediator
– Travels through cytosol to ER
– Binds to and opens IP3-gated Ca2+-release channels (IP3 Receptors)
– Ca2+ stored in the ER is released through the open channels
o DAG can bind to and activate protein kinase C (PKC)
How does hypothalamus stimulate pathway in pituitary cell

 Hypothalamus:
o Section of small nuclei
o Located in the center of the brain (hypo = under, thalamus = top section of the
brain)
o Receives stress signals from the brain
 Under stressed circumstances, hypothalamus releases CRH and
sends to anterior pituitary cell
 Pituitary releases ACTH, which travels to the adrenal gland
 NB: VB also stimulates the release of ACTH
 REMEMBER: CRH and VP stimulate ACTH release
o VP stimulates the phospholipase pathway
o CRH stimulates the cAMP pathway
Receptor Desensitization
 Receptor Desensitization = the process whereby a prolonged exposure to a
stimulus decreases the cells’ response to that level of stimulus (ie: cell is numb to
effects of signal molecule)
o Enables cells to respond to changes in the concentration of an extracellular
signal molecule
o Prevents cell from being locked in ‘on position’ when receptor is constantly
occupied  consequence of cell locked in ‘grow and divide’ = cancer
 Desensitization can occur in various ways:
(1) Receptor Sequestration = receptor temporarily captured in endosomes
(vesicles that will transport molecules to different parts of the cell)
(2) Receptor Down-Regulation = receptor is broken down in a lysosome
(3) Receptor Inactivation = receptor inactivated on the cell surface by being
phosphorylated or methylated
(4) Inactivation of Signaling Protein = intracellular signaling proteins involved
in inducing signal are modified
(5) Production of Inhibitory Protein = inhibitory protein produced that blocks
transduction process

lOMoARcPSD|1377855
The Synthesis of PIP2

 PIP and PIP2 are produced by phosphorylation
o PI Kinase adds phosphate to phosphatidylinositol (PI)
o PIP Kinase adds phosphate to PIP  PIP2
 NB: kinases can phosphorylate sugars in phospholipids to make them ready for
cellular signaling
Phospholipase C (PLC) cleaves PIP2 into IP3 and DAG

 DAG binds to and activates PKC, which phosphorylates effector proteins
 IP3 helps release Ca2+ from the ER

lOMoARcPSD|1377855
G Protein Coupled Receptor Activates Phospholipase C (PLC)
CRH and VP
 Although CRH and VP both stimulate the release of ACTH, they use different
messenger systems
o CRH  Gs protein  adenylyl cyclase  cAMP  PKA
o VP  Gp protein  PLC  DAG  PKC
 IP3  Ca2+ released
 PKA and PKA phosphorylate effector proteins in the cell, which leads to the release
of ACTH
Effects of forskolin on β-endorphin/ACTH release

 Forskolin = an activator of cAMP
 Pituitary cells stimulated with forskolin. Conclusion:
o Forskolin stimulates the production of cAMP
o cAMP results in the release of ACTH
 ∴ If a pathway can be stimulated by forskolin then it is
a cAMP pathway

lOMoARcPSD|1377855
Ca2+ Function as a Ubiquitous Intracellular Mediator

 Many extracellular signals trigger an increase in cytosolic Ca2+ concentration
o Ca2+ conc in the cytosol is normally very low, whereas they are very high in
the extracellular fluid and ER lumen = large gradient difference
o When a signal opens a Ca2+ channel, Ca2+ rushes into the cytosol- increasing
the conc and ∴ stimulating Ca2+ responsive proteins in the cell
 Several mechanisms keep Ca2+ conc in the cytosol low in resting cells
o Ca2+ is actively pumped out of the cytosol to the cell exterior
– Ca2+-pump uses the energy of ATP hydrolysis to pump out Ca2+
– Ca2+ transport protein, couples the efflux of Ca2+ to the influx of Na+
o Ca2+ is pumped out of the cytosol into the ER and mitochondria
– Ca2+-pump uses the energy of ATP hydrolysis to pump Ca2+ into the ER
(ready for release later on)
– Sym-port transports Ca2+ into mitochondria
o Various molecules bind to Ca2+ in the cytosol (Calmodulin)
Calmodulin
 Calmodulin = the protein attracted to Ca2+
o Governs many Ca-regulated processes
o 4, high affinity Ca-binding sites
o When bound to by at least 2 Ca, it undergoes a
conformation change and becomes activated
o Can then bind to and activate other proteins
 Aids the activation of enzyme CaM-kinase II
o CaM-kinase II = large
protein complex of 12
subunits
o Ca/calmodulin binds to and
activates CaM-kinase
o CaM-kinase then auto-
phosphorylates itself to
become fully active
o Ca is released, causing
calmodulin to dissociate but
CaM-kinase remains active
due to auto-phosphorylation

lOMoARcPSD|1377855
Protein Sorting
Major Cellular Compartments
NB: all eukaryotic cells have the same basic set of membrane-enclosed organelles
 Plasma membrane = the phospholipid bilayer that surrounds a living cell
 Nucleus = contains the genome and it is the principal site of DNA and RNA synthesis
 Cytoplasm = consists of the cytosol and the cytoplasmic organelles
 Cytosol = an aqueous solution of molecules with a gel-like consistency. It is the site
of protein synthesis and degradation and performs most of the cell’s intermediary
metabolism
 Endoplasmic Reticulum (ER) = a network of membranous sacs (termed cisternae)
extending throughout the cytoplasm of a e-cell
 Golgi apparatus = stacks of 4-10 disc-shaped cisternae which, synthesize
polysaccharides (carbs), glycosylate proteins and sort molecules for storage or
secretion (NB: proteins produced in ER MUST go through Golgi if destine for the cell
exterior)
 Mitochondria = a DNA containing organelle found in animal cells, which is the site
of cellular respiration. It is surrounded by a highly permeable double membrane and
contains circular DNA molecules, RNA and small ribosomes.
 Lysosomes = a membrane-bound compartment containing hydrolytic enzymes to
breakdown and recycle many molecules
 Endosomes = a membrane-bound compartment that processes material taken up by
endocytosis (invagination) before transport to lysosomes for degradation
 Peroxisomes = a type of micro-body that contains numerous enzymes used in the
production and degradation of peroxides and the oxidation of amino acids

lOMoARcPSD|1377855
Evolution of the Nucleus

 Ancient prokaryote cells began to develop a nucleus
 Plasma membrane is invaginating (going inwards) to form a membrane around DNA
 As membrane forms, ‘fingers’ form ER
 Mitochondria (and plastids) are though to have originated when a bacterium was
engulfed by a larger pre-eukaryotic cell- could explain why mit have their own
genomes and why their lumens remain isolated from other membrane traffic
Protein Trafficking
 The synthesis of all proteins begins on ribosomes
 Their fate depends on their aa sequence:
o Many contain a sorting signal sequence, which directs their delivery from the
cytosol to outside the cell, the nucleus, the ER, mitochondria, plastids or
peroxisomes
o Nuclear localization signals are responsible for directing nuclear proteins to
the nucleus
o Others that do not have a sorting signal remain in the cytosol
 Methods of transport include:
(1) Gated transport (nuclear pores)
(2) Transmembrane transport (nuclear import receptors)
(3) Vesicular transport
 Roadmap of protein traffic:

lOMoARcPSD|1377855
Vesicular Transport- budding and fusion

 Transport vesicles bud from one compartment (donor) and fuse
with another (target) compartment
 Allows soluble components to be transferred from one lumen to
another
 NB: the membrane is also transferred such that the orientation
of proteins and lipids is preserved in its new location (ie:
membrane proteins retain the same domains facing the cytosol)
Gated Transport- nuclear pores

 Proteins move between the cytosol and the nucleus (which are
topologically equivalent) via nuclear pore complexes in the nuclear
envelope
 The Nuclear envelope = the double-membrane surrounding the nucleus
o Penetrated by pores, which are gated by nuclear pore complexes
o Envelope is continuous with the ER
 Nuclear pore complexes = selective gates that actively transport
macromolecules and macromolecular assemblies (also allow diffusion
of smaller molecules)
o 3000-4000 NPCs perforate every nuclear envelope
o Fibers guide molecules in/out
o Proteins lining the pore, restrict the size of molecule that can enter- larger
molecules require more energy to pass through
Transmembrane Transport- nuclear import receptors

 Nuclear import receptors carry proteins through the NPC- they bind to both the
cargo protein and the cargo’s nuclear localization signal (NLS)
 NLSs (encoded in primary aa sequence) must be recognized by import receptors in
order for binding between the two
 Each receptor is specific to a cargo
 Some receptors require adaptor proteins in order to bind to their cargo

lOMoARcPSD|1377855
Ran GTPase imposes directionality on transport

 The process of importing and exporting nuclear proteins through the NPC is
energized through the hydrolysis of GTP by the monomeric GTPase Ran
 Ran exists in two conformational states (depending on whether GDP or GTP is
bound), which are catalyzed by Ran-GAP and Ran-GEF
o Ran-GAP is located in the cytosol. It triggers the hydrolysis of GTP  GDP to
drive Ran-GDP through the NPC
o Ran-GEF (exchange factor) is located in the nucleus. It promotes Ran to
exchange GDP for GTP, driving Ran-GTP through the NPC
Ran’s influence on nuclear import receptors

 Some proteins shuttle back and forth between the nucleus and the cytosol and ∴
contain both NLS and nuclear export signals
 Ran has a higher affinity for receptors than cargo molecules and ∴ by binding to the
receptor in place of the cargo (GTP inside nucleus and GOP inside cytosol), Ran
promotes the receptor to release its cargo and travel back to the other side of the
NPC in order to continue transport
 The rate of import and export is regulated by shuttling proteins- if the rate of import
exceeds the rate of export, a protein will be mainly located in the nucleus (and vise
versa)

lOMoARcPSD|1377855
Protein import into the mitochondrion by TIM and TOM

 At least one signal sequence is required to direct a mitochondrial precursor protein
to its appropriate sub-compartment
 Protein translocators are required to transport molecules across the mitochondrial
membrane
o TOM = translocase outer mitochondrial membrane
o TIM = translocase inner mitochondrial membrane
 Newly synthesized mitochondrial proteins remain unfolded in the cytosol through
interactions with other proteins
 Once bound to TOM, the interacting proteins release the m-protein and it is fed into
the translocation channel
 It is then passed through TIM into the mitochondria, where it folds into its mature
form
Intracellular Vesicular Transport

Exocytosis and Endocytosis
 Exocytosis = the process by which the
biosynthetic-secretory pathway delivers
newly synthesized proteins, carbohydrates
and lipids to either the plasma membrane
or the extracellular space
o The biosynthetic-secretory pathway
leads outward from the ER towards
the Golgi and cell exterior
 Endocytosis = the process by which
components outside the plasma membrane are taken in and transported to
endosomes (where they can be recycled to the same or different parts of the plasma
membrane) or lysosomes (for degradation).
o The Endocytic pathway leads inward from the plasma membrane

lOMoARcPSD|1377855
Vesicular Transport
 Proteins are able to travel without having to cross a membrane- instead they are
passed from one compartment to another
 The lumen of each membrane enclosed compartment is topologically equivalent
 Transport vesicles continually bud off from one membrane and fuse to another,
carrying cargo
 Each vesicle that buds must be selective- only taking up appropriate molecules and
fusing with the correct target membrane
 Proteins on the donor compartment interact with proteins on the target
compartment, allowing it to dock and deliver its cargo
Road map of the cell
 Red arrows indicate inside  out (exocytosis)

 Green arrows indicate outside  in (endocytosis)
Vesicle Coat Proteins

 Transport vesicles bud off as coated vesicles- have a distinct cage of proteins
covering their cytosolic surface
 The coat is discarded before fusing to the target membrane (so that the two cytosolic
membrane surfaces can interact directly and fuse)
 Two main functions:
o Selects the appropriate molecules for transport
o Molds the forming vesicle (stabilizes the plasma membrane)

lOMoARcPSD|1377855
 Three types of protein, each used for different transport step

(1) Clathrin- transport material from the plasma membrane and between
endosome and Golgi compartments (endocyclotic)
(2) COPI- bud from Golgi compartments
(3) COPII- bud from the ER
Pulse Chase Experiments

 How to track transport of molecules through a cell
 Basic idea-
o Pulse the cell with a tracer molecule (labeled with
radioactivity or fluorescence) for a short amount of time
o Cell will absorb tracer
o Cell washed and placed in a tracer-free medium
 Example: Radioactive amino acid (leucine) added to cell culture medium to track its
transport through the cell
o Radio labeled leucine travels through the cell to the ER where it is
incorporated into protein and then released from the rough ER into the Golgi
o Radiolabelled protein is then packaged into vesicles and exocytosed into
medium
o At any point in time, different segments of the cell can be purified out (eg: the
nucleus) and the amount of radioactivity can be measured to determine the
path of leucine
o Process can be repeated at different intervals to determine the entire
transport path in cell
Clathrin Coat Structure

 Composed of three large (heavy) and three small (light) polypeptide
chains that form a three-legged structure called a triskelion
 Clathrin triskelions form a basket-like cage around the cytosolic surface
of membranes
 Adapter proteins are positioned between the Clathrin cage and the
membrane- to bind the two together and trap various transmembrane
proteins and receptors that capture cargo molecules inside the vesicle

lOMoARcPSD|1377855
Assembly and Disassembly of a Clathrin Coat

 Cargo-receptors pick up cargo molecules from one side of the membrane
 Adaptor proteins bind to both membrane-bound receptors and clathrin trikelions, ∴
mediating the formation of the cage
 The assembly of the coat introduces curvature to the membrane
 The clathrin coat is rapidly lost shorty after the vesicle forms
 LDL transported to the endosome and lysosome
Dynamin pinches off budding vesicles

 Dynamin assembles into a ring around the neck of the forming bud
 Together with other proteins, dynamin destabilizes the lipid bilayer, pinching off the
newly formed vesicle from the membrane
 Specific mutations in dynamin can either enhance or block the pinching-off process
 Type of GTP-binding protein
Formation of a COPII-Coated Vesicle

 GTP-binding protein primes membrane for COPII coating:
o Coat-recruitment GTPases = monomeric GTPases
responsible for coat assembly (eg: Sar1 protein)
o Inactive Sar1-GDP binds to Sar1-GEF in the ER
membrane, causing Sar1 to release GDP and bind
GTP
o This conformation change exposes Sar1’s
amphipathic helix (both hydrophilic and phobic
parts ∴ can slot into membrane), which inserts
into the ER plasma membrane and initiates a

lOMoARcPSD|1377855
process of membrane curvature

 Sar1 anchors two COPII proteins:
o Membrane-bound Sar1-GTP binds to a complex of two COPII
proteins, called sec23/24
o Sec 24 has several binding sites for cytoplasmic tails of cargo
receptors
 The complex attracts more COPII proteins:
o A second layer of two COPII proteins form the outer coat
o Membrane-bound Sar1 continues to recruit COPII subunits to
the membrane until it forms a bud
Tethering of a vesicle to a target membrane

 Rab effector proteins (tethering proteins) interact with Rab proteins located both on
the target membrane and vesicle membrane to establish the first connection
between the two that are about to fuse
 Snare membrane-bound proteins (v-snare located on the vesicle and t-snare on the
target) intertwine to dock the vesicle to the target and catalyze the fusion of the two
bilayers
 Cargo is released into cell
Endocytosis of low-density lipoprotein (LDL)

 Clathrin-coated vesicles allow macromolecules from the extracellular fluid to be
taken up by the cell, in a process called receptor-mediated endocytosis
 Because cargo molecules are captured by specific receptors, large amounts of cargo
can be internalized by the cell, without taking in large volumes of extracellular fluid
(ie: a very efficient process)
 Cholesterol is transported in the blood in the form of lipid-protein particles known
as low-density lipoproteins (LDLs)
 LDL is taken up by cells for the synthesis of new membranes through receptor-
mediated endocytosis
o When the cell requires cholesterol, it produces and inserts LDL receptor
proteins into the plasma membrane

lOMoARcPSD|1377855
o The receptor proteins bind to both LDL (in the extracellular fluid) and
adaptor proteins (on the cytosol side)
o The adaptor proteins are bound to and help the formation of the clathrin coat
o Curvature commences and vesicles are formed, containing LDL particles
bound to LDL receptors
o After the clathrin coat is shed, vesicles deliver their cargo to the early
endosomes and then the lysosomes, where the LDL particles are released and
hydrolyzed to form free cholesterol (ready for membrane synthesis)
 If uptake of Cholesterol is blocked, cholesterol accumulates in the blood, causing the
formation of atherosclerotic plaques on the artery walls, which blocks blood flow,
leading to strokes and heart attacks
 The development of atherosclerosis:
o Atherosclerosis = condition in which the artery walls thicken, reducing
blood flow and leading to heart diseases
o Some individuals inherit defective genes
encoding LDL receptors- this means the uptake
of LDL is blocked and cholesterol will
accumulate in the blood
o This causes the formation of atherosclerotic
plaques on the artery walls (deposits of lipid
and fibrous tissue), blocking blood flow and
leading to strokes and heart attacks
The Dynamic Microtubules

The Cytoskeleton
 The cytoplasm of eukaryotic cells is spatially organized by a highly dynamic network
of filamentous fibers, collectively termed the cytoskeleton
 Types of fibers:
o Microtubules (25nm)
o Intermediate filaments (8-12nm)
– Keratins
– Vimentin and desmin
– Neurofilaments
– Lamin
o Actin microfilaments (7-8nm)
 NB: no intermediate filaments are found in plants
 Each filament type is polymerized (self-assembled) by their own unique subunit
type- tubulins, actins, lamin
 The diverse activities of mt and actin filaments are mediated by accessory proetins-
microtubule-associated proteins (MAPs) and actin-binding proteins (ABPs)
The Role of the Cytoskeleton

 Pivotal to cell division- it forms structures that pull chromosomes apart during
meiosis and mitosis to enable the cell to divide into two

lOMoARcPSD|1377855
 It drives and guides the intracellular traffic of organelles (eg: vesicles)- movement
powered by the action of actin filaments and their associated proteins
 It enables cells to swim, crawl and drives muscle contraction
 It shapes the growth of plant cells
 It functions in mRNA and protein synthesis, and in signal transduction
Microtubule Structure and Properties

 Microtubules are long, hollow polymers of tubulin
 13 tubulin proteins make up the mt ring (hollow lumen in the middle), rings stacked
on one another to form tube
 The smallest mt subunit = αβ-tubulin heterodimer
o Complex of α and β subunit- differ in the
way they interact with GTP
 α- GTP can NOT be hydrolyzed
 β- GTP can be hydrolyzed to GDP
o Subunit has polarity (due to difference
between α and β)- α– and β+
 Mt filament has structural polarity (due to αβ)
o Plus end (β) = fast growing/shrinking
o Minus end (α) = slow growing/shrinking
 Highly labile (when not stabilized) ie: continuously growing and shrinking
GFP: Green Fluorescent Protein

 A fluorescent protein found in the jellyfish Aequorea victoria
 GFP is used to locate proteins (and genes) of interest inside live cells
o The gene encoding GFP is inserted next to the gene for the protein of interest
using molecular biology
o DNA introduced into nuclei of living cells
o Cells will begin to express the protein with GFP attached
o Produces a ‘fruit bowl’ of colors
 Detecting GFP: fluorescence microscopes
o Lights of different wavelengths are shown on the sample
o Fluorescent tags will absorb light of a higher energy than they
emit- blue light = higher energy, red light = lower energy
o If we tag mt with GFP (GFP-tubulin) and shine blue light on
them, they will show up green
 Microtubules and FRAP
o FRAP = fluorescence recovery after photobleaching
o GFP labeled Mt are zapped with bright light until they are no longer
fluorescent
o Watch to see result- do they recover fluorescence?
 Microtubule plus ends
o Mt have associated proteins that help them to grow, shrink or become
stabilized- one of these is EB1 (end binding)
o When expressed as GFP-EB1, growing Mt ends will light up, showing speed
and motion of growth

lOMoARcPSD|1377855
Dynamic Instability
 Mt polymerize and depolymerize continually (half life = 5-10 min)
 Dynamic instability = mt are constantly changing (grow, shrink,
stable)
 Elongation occurs mainly at plus ends (β)
 Studying microtubule polymerization and de-polymerization in
vitro (in the lab):
o Ingredients = αβ-tubulin, Mg2+, GTP, 37oC, neutral pH
o At 37oC mt polymerize, at 4oC they depolymerize (cycles of
heating and cooling will produce a very pure sample of mt)
o In-vitro assembly requires a certain min αβ-tubulin conc (too
dilute = will not grow)
o At assembly/disassembly equilibrium, there is a population
of ~50% free tubulin
 GTP cap:
o Microtubule assembly requires the binding of GTP
to tubulin subunits, which is hydrolyzed to GDP
o In polymerization, the addition of new subunits
containing GTP is faster than GTP hydrolysis,
forming a protective GTP-cap that keeps the
molecule growing
o If addition of new subunits slows down, hydrolysis
of GTP exceeds addition, the GTP cap is lost and
the mt shrinks
 Different stages of growth:
o Growth = rapid polymerization with GTP-capped end
o Catastrophe = accidental loss of GTP cap
o Shrinkage = rapid de-polymerization
o Rescue = regain of GTP cap
o Growth =
 Dynamic instability can be utilized for specific purposes- eg: mitosis or cell
morphogenesis
o Localization of a mt-capping protein in a specific domain of the plasma
membrane will stabilize mts growing in that direction
Microtubule-nucleating complex
 Microtubule nucleation and elongation in living cells stems from γ-tubulin ring
complexes (γTuRCs)
o γ-tubulin forms ring complexes with 5-8 accessory proteins in stoichiometric
proportions (the smallest contains 2 γ-tubulins and 2 accessory proetins)
o γ-tubulin is a key component of Microtubule-organizing centers (MTOCs)
o In animal cells, γTuRCs aggregate into centrosomes (not in plant cells)

lOMoARcPSD|1377855
 Centrosomes produce microtubules

o Mts grow and shrink independently from γ-tubulin ring complexes of the
centrosome
o Minus end is stabilized in ring complex, plus ends grow and shrink as
required
Polymerization Prevention
 The chemical toxin Taxol binds to tubulin subunits, preventing polymerization
 Existing mts will hence depolymerize by dynamic instability
 Chemical used in certain chemotherapy treatment- stabilizes the mt to prevent cell
division
Microtubule Associated Proteins (MAPs)

 Bind to mts and modify their properties
 Stabilize mts against disassembly
 Mediate mts interaction with other mts or cellular components
MT Motors: Kinesins and Dyneins

From last time…
 Composed of α and β-tubulin heterodimers
 MTs are polar filaments- plus end (β) and minus end (α)
 In animal cells, MTs are nucleated in centrosomes from γ-tubulin ring complexes-
minus ends are linked to ring complexes while the plus ends grows out to the cell
periphery
Example: Pigment granules in a fish-scale cell

 When fish wants to change its colour (hormones released)- distribution of pigment
molecules changes from dispersed to aggregated
 Both dynein and kinesin associate with pigment granules- playing a tug-of-war
 Stronger kinesin wins, pulling granules outward along microtubules (dispersed)
 Upon stimulation (hormonal change causing a decrease in cAMP), kinesin is
inactivated, leaving dynein free to drag the granules towards the cell center, rapidly
changing the fish colour
 Bidirectional movement on MT: dynein = plus end  minus
end, kinesin = minus end  plus end

lOMoARcPSD|1377855
Motor proteins use MTs as train tracks to move cargo

 Kinesins
o Move towards the plus end
o Involved in organelle transport, mitosis, meiosis, movement of synaptic
vesicles along axons, cytoplasmic streaming
 Dyneins
o Move towards the minus end (center of cell)
o Types- ciliary and cytoplasmic dynein
o Involved in organelle transport (Golgi) and in mitosis
o NB: no dyneins in plants
Structure of Motor Proteins

 Composed of two heavy chains plus several light chains
 Each heavy chain is attached to a globular, ATP-binding head:
o Head binds to and ‘walks along’ MT
o Specifies the rate and direction of movement
 Tail at the end binds to the cargo
 Movement force is generated by ATP-hydrolysis
Kinesin movement
 A kinesin molecule “walks” along a MT
 Head binds to β-tubulin monomers
 Head-over-head action in 8nm steps towards plus end
o The binding of ATP allows one head to bind tight to MT
o Once ATP is hydrolyzed, the head dissociates
o Binding of ATP to other head causes a conformation change that swings head
over to bind to next β-tubulin monomer
 Rapid movement along MTs
 NB: only one head can be tightly bound to the MT at once

lOMoARcPSD|1377855
Motor Proteins distribute organelles within cells

 Maintains the position of organelles within the cell
 Kinesin molecules attach to ER, pulling it outward towards the edge of
the cell (along MTs) and stretching it like a net throughout the
cytoplasm
 Dynein molecules pull Golgi towards the nucleus (NB: interacts with
other proteins, such as dynactin)
Dynein movement
 Faster and smoother than kinesin movement
 Movement powered by ATP hydrolysis (bound to head)
Cilia and flagella organelles- axonemes

 Cilia and flagella are hair-like appendages found in most animals, protozoa and some
lower plants
 Composed of axoneme (long structure of MTs) covered by a plasma membrane
 9 doublet MTs (doublet = half a
MT bound to a whole MT)
 Central MT pair
 Outer MTs have outer and inner
dynein arms to carry ciliry dynein
molecules (not cytoplasmic
molecules)
 Radial spokes connect outer and
center MTs
 Primary functions:
o Move fluid over cell surface (for feeding or locomotion)
o Sweep mucus in epithelial cells
o Sweep eggs along oviduct
o Swimming of algae and sperm
 Movement is generated by the bending of the axoneme
 Axoneme compostion is identical in almost all forms of cilia and flagella
 Examples in body:
o Cilia on the epithelium of human respiratory tract
o Flagellum propels a sperm using wavelike motion
Cilia and flagella movement

 Ciliary dynein drives axonemal bending
o Tail region bound to α tubule of MT doublet
(tightly bound)
o Head region hydrolyze ATP, moving along B tubule
of an adjacent MT doublet (able to walk along
molecule)
 Cross-link proteins between adjacent MT doublets allow
dynein movements to translate into axonomal bending

lOMoARcPSD|1377855
Development of axoneme structures

 Axonemal structures grow from basal bodies
 A basal body is composed of 9 sets of triplet (A, B, C) MTs, with other
proteins holding the MT array together
 Arrangement of MTs in a basal body and a centriole is similar, and the
two are functionally inter-convertible
 Centrioles usually arise by the replication of existing centrioles
 A centrosome consists of 2 centrioles perpendicular to each other-
before cell division, the two separate and a daughter centriole is formed
perpendicular to each original. The two pairs then move apart to form
the mitotic spindle
Actin and Actin-Binding Proteins

Actin Filaments
 Actin is an important cytoskeletal protein that:
o Allows cells to move
o Allows cells to change shape
o Aids cells to perform various functions eg: actin fibers form a contractile ring
during cell division to separate the two daughter cells
 Actin filaments (F-actin) are composed of uniformly oriented actin molecules (G-
actin)
 Two smaller protor filaments twist together to form an actin filament
 F-actin is thinner and more flexible than MTs ∴ can bend to form many shapes
 Actin forms remarkably different structures- stiff and permanent or dynamic and
unstable (influenced by ABPs)
 Polar polymers:
 Plus end = fast growing
 Minus end = slow growing
Actin Molecules (G-actin)

 Molecules of G-actin with ATP bound assemble into F-actin
(molecules contain ATP binding site in center)
 When ATP is hydrolyzed to ADP, the filament is destabilized
 Polar molecule- plus end and minus end

lOMoARcPSD|1377855
Actin Polymerization
 NB: similar process to MT polymerization
 Polymerization in vitro requires ATP, K+, Mg2+
 There needs to be a high conc of monomers in solution for filament to form
 The plus end polymerizes 10x faster than the minus end
 Polymerization is accompanied by the binding and hydrolysis of ATP  ADP (which
remains trapped in the molecule)
 F-actin is dynamic with a treadmilling motion (equal additions at plus end to losses
at minus end)
Treadmilling Motion of F-actin

 ATP bound actin molecules bind to the plus end of the filament
 ATP is hydrolyzed to ADP, causing a conformational change which destabilizes the
molecule
 ADP bound actin falls off filament and releases ADP, ready to be bound to ATP
 Net movement forward
Drugs and signals that affect the stability of actin

 Cytochalasins (fungal toxins)- bind to plus end of actin and prevent polymerization
(useful for studies of cell locomotion- leading edge of cell will stop moving and
retract)
 Phalloidins (toxins from amanita)- bind along f-actin, stabilizing the filament against
depolymerization
 Fluorescent phalloidins- bind to actin filaments in cell, staining them for
visualization in a fluorescent microscope
 Cell surface receptors- regulate the polymerization of actin
Actin-binding proteins
 Nucleation of actin filaments is promoted by actin-related proteins (ARPs)
o Their shape is very similar to the plus end of f-actin ∴ inducing filament
growth on their surface
 Actin filaments interact with various acting-binding proteins (ABPs) to form complex
networks that vary in length, stability and geometry
o Actin cross-linked with spectrin = mechanical support in plasma membrane
o Actin with fimbrin = parallel bundles at leading edge of cell
o Actin with α-actin = contractile bundles in stress fibers
o Actin with filamin = gel-forming proteins

lOMoARcPSD|1377855
Formation of two types of f-actin bundles- action of ABPs

 The length of cross-link proteins determines how close filaments can pack together.
Packing distance will either prevents or allows other proteins from entering the
bundle to interact with the filaments
o Actin cross-linked with α-actin = contractile bundle = myosin-II allowed to
enter bundle
o Actin cross-linked with fimbrin = parallel bundle = mysin-II prevented from

entering bundle
Movement of cell from leading edge

 Lamellipodium = protrusion from cell containing a high conc of actin filaments,
from which it moves forward (ie: the leading edge)
 Growing actin filaments (polymerization at plus end) push the edge of the cell
forward = protrusion
 ARP complexes will stem new filaments off existing filaments
 Destabilizing proteins depolymerize filaments at the minus end (away from leading
edge) so that actin molecules can be reused for polymerization
Myosin: Actin-binding motor protein

 Myosin interacts with actin filaments to drive cellular motility
 The action of actin and myosin are the basis of:
o Actomysin motor in contractile rings
o Stress fibers
o Cell motality
o Muscle contraction
o Cytoplasmic streaming
 Move to the plus end of filament (similar to kinesin)

lOMoARcPSD|1377855
Myosin I
 Structure:
o 1 globular head- binds to actin filaments
o 1 small chain (tail)- binds to membranes
 Hops along filament towards plus end- movement dependent on what it is bound to
 If bound to plasma membrane (fixed) filament will glide along membrane
(minus end first) as myosin hops along
 If bound to vesicle membrane, vesicle will move along filament towards plus
end
 Functions:
o Moving one acting filament relative to another
o Moving vesicles along filament
o Moving filament along membrane
Myosin II
 Structure:
o 2 globular heads (ATP binding site)- walk along filament
o 2 light chains, 2 heavy chains
 Skeletal muscle:
o Composed of bundles of thin actin filaments interlocked with thick myosin
filaments
o When myosin walks along plus end of actin (due to chemical signal), muscle is
contracted to a smaller structure
 Actomyosin motor drives muscle contraction:

(1) Ca2+ signals contraction
(2) Myosin head bound tightly to actin with no ATP
attached (rigid)
(3) ATP binds to head, making it detach from filament
(4) ATP is hydrolyzed to ADP, which causes a
conformational change in myosin that allows it to
weakly bind to the next actin molecule in the
filament
(5) ADP is released, allowing it to tightly bind again

lOMoARcPSD|1377855
Mitosis Recap
 Mitosis = the process in which two identical copies of every chromosome are
separated to allow accurate partitioning into every daughter cell
 Division of the nucleus
(1) Prophase:
 Chromatin (coiled strands of DNA) condense to
form visible chromosomes structures → a dense
compaction
 Each condensed chromosome consists of two
identical sister chromatids, joined at the
centromere
 Nucleolus disappear
 Centrosomes produces fibers (bundles of microtubules) termed
asters and move to opposite poles of the nucleus → forming
mitotic spindle
(2) Prometaphase:
 The nuclear envelope degenerates and the microtubules push
into the nuclear region
 Kinetochore MT fibers attach to each kinetochore of the
chromosome (a protein around the centromere)
 NB: non-kinetochore MT fibers = fibers of spindle not attached to
chromosomes
(3) Metaphase:
 Kinetochores are aligned on the metaphase plate, midway between
the poles- one sister chromatid on each side
 NB: takes a significant amount of time for cell to ensure that each
centromere is connected to the spindle and each chromosome has
reached the metaphase plate
(4) Anaphase:
 Sister chromatid are pulled apart by kinetochore microtubules →
becoming full chromosomes
 A = chromosomes move to opposite poles
 B = non-kin microtubules elongate spindle- pushing poles further
apart
(5) Telophase:
 Action of non-kin microtubules elongates cell further
 Chromosomes de-condense into chromatin
 Nuclear envelope forms around each set of chromosomes
 Nucleoli reforms
 Cytokinesis begins

lOMoARcPSD|1377855
Mitotic and Cytokinetic Apparatus

 Cells need to be able to divide exactly so that each daughter cell contains an identical
copy of the DNA
 Complicated, highly conserved evolutionary process
Mechanisms in the cell cycle

 Structural cycles:
o Nuclear cycle
o Microtubule cycle
o γ-tubulin cycle
o Actin filaments cycle
o Organellar cycle
 Regulatory proteins:
o Cyclins
o Cdc proteins
o Cell cycle kinetics
Phases of the eukaryotic cell cycle

 NB: cell division = mitosis + cytokinesis
 Interphase: cell at a resting state
o G1 (gap 1) phase- biosynthesis
 1st check-point- ensure environmental signals indicate that it is time to
divide
o G0 phase- if conditions do not favor mitosis, cell can remain in this face for
long periods of time.
o S (synthesis) phase- DNA replication
o G2 phase
– Before mitosis
– Centrosome duplicates
 2nd check-point
 Mitosis (M phase):
o Prophase
– Centrosomes move to opposite sides of the
nucleus
– MTs begin to shrink
– Chromatin condenses into chromosomes
o Prometaphase
– Nuclear envelope breaks down
– Centrosomes are at either side of the nucleus- begin to radiate out MTs
o Metaphase- chromosomes at equator
– Centrosomes produce spindle fibers (MTs)
– Chromosomes align at equator (spindle)
 3rd check-point
o Anaphase A/B
– Sister chromatids are split apart to form full chromosomes

lOMoARcPSD|1377855
o Telophase
– Chromosomes de-condense
 Cytokinesis: division of cell into two
Mitotic apparatus in animal cells

 At prophase, daughter centrosomes move (by motor proteins) to opposite sides of
the nucleus
 MT dynamic instability (plus end grows/shrinks faster than the minus end)
generates two half-spindles of opposite polarity, with the MT plus ends overlapping
at the equator
 At prometaphase, randomly probing MTs bind to kinetochores (center of
chromatids) by a ‘capture’ mechanism
 Three sets of spindle MTs are present at metaphase- astral, polar, kinetochore
 At anaphase A, kinetochores move towards the poles
 At anaphase B, a sliding force generated between overlapping polar MTs plus pulling
force on poles moves the poles and the chromatids further apart
Formation of a bipolar spindle in animal cells

 Daughter centrosomes move to opposite poles of the nucleus
 Action of dynamic instability- MTs will grow out (plus ends)
from centrosomes, trying to latch onto a chromosome
 Inter-polar MTs overlap along the equator, forming the structure
of the spindle
o Kinesin-5 proteins connect overlapping MTs
 Kinetochore MTs of the mitotic spindle attach to chromsomes
o Specialized MT binding region = the kinetochore
o Kinetochore proteins located around the centromere
region of the two sister chromatids
o NB: MTs must be attached to both sides of the
chromosome (if only one is attached, MT will be released)
 Astral MTs grow out into the cytoplasm
o Set up the position of the spindle inside the cell (can bind to plasma
membrane)
Alternative models of kinetochore generated polar force

(1) ATP-driven motor protein drives both chromosome movement and MT disassembly
(2) MT disassembly drives chromosome movement towards spindle

lOMoARcPSD|1377855
Forces that separate chromatids at anaphase A and B

 Anaphase A: chromosomes are pulled pole-ward
o Kinetochore MTs shrink
o Cohesion proteins binding sister
chromatids weaken
o Forces are generated at kinetochores to
move daughter chromosomes toward their spindle poles
 Anaphase B: poles are pushed and pulled apart
o A sliding force is generated between overlapping inter-polar
MTs to push them apart
o Plus ends of inter-polar MTs grow while still bound by kinesin-5
o Kinesin-5 slides growing ends along each other, pushing poles apart
o A pulling force acts directly on the poles to move them apart
Cytokinesis in animal cells: Cleavage

 The plane of cell division is determined by the position of
the mitotic spindle
 A cleavage furrow begins to form during anaphase,
perpendicular to the long axis of the spindle
 A contractile ring of actin filaments and myosin
constricts the plasma membrane until two separate cells
are formed
Cytokinesis in plant cells: phragmoplast and cell plate

 Preprophase band of MTs leaves a molecular memory at the future division site in
the cell cortex, irrespective of spindle orientation
 Two sets of antiparallel MTs, with their plus ends inter-digitating, form a cytokinetic
phragmoplast centrally between the daughter nuclei
 The MT array then expands centrifugally while MTs in center disappear, forming a
ring that builds the edges of a new cell wall (cell plate)
 Phragmoplast MTs work in cooperation with actin filaments, partner proteins and
Golgi-derived vesicles

lOMoARcPSD|1377855
Tools in Contemporary Cell Biology

 NB: animal cells are colorless and translucent
 Microscopy relies on the performance of the microscope as well as preparing the
sample correctly
Compound Light Microscope

 Light source = LED
 Light is focused on the specimen by lenses in the condenser
 A combination of objective lenses and eyepiece lenses are
arranged to focus an image of the illuminated specimen in
the eye
 Between light source and condenser = field diaphragm
 Condenser sharpens edges of black circle
 NB: light waves do not follow the idealized straight path,
instead they travel slightly different routes and interfere with
each other
 If two waves reaching the same point by different
paths are in phase, light = bright and clear
 If two waves reaching the same point by different
paths are out of phase, light = dim and bad resolution
Preparing cells to view under a light microscope

 Fixing = chemical added to cell, which stabilizes and stops any enzymatic function
by cross-linking proteins
o Fixative examples: glutaraldehyde (highly toxic), osmium tetroxide (non-toxic
alternative)
o NB: take precaution when using highly toxic chemicals- read safety label,
search online database (chemalert) for material safety data sheet
 Embedding = soft and fragile tissues are embedded in a supported medium
o Usually embedded in resin or a cryogenic chemical that solidifies when cooled
o Block can then be readily sectioned
 Sectioning = cutting sample into very thin slices using a microtone so that individual
cells can be examined directly at a high resolution
 Staining = colouless and translucent cells are stained in order to visualize
o Example: H and E stain- nuclei blue, everything else pink (non-specific)

lOMoARcPSD|1377855
Fluorescence Microscopy
 Individual proteins labeled with different colours
 NB: fluorescent molecules will emit light in a different wavelength than absorbed
(eg: excited by blue light, will reflect green)
 Molecule labeled with fluorescently tagged anti-body
Scanning electron microscope

 Much higher resolution than a light
microscope- clearer and closer images
 Beam of electrons directed from electron gun
source through sample
 Magnetic coils (instead of lenses) focus e beam
on the specimen
 Specimen must be placed in a vacuum ∴ not
alive
 Detector measures the quantity of electrons
scattered by specimen to produce a 3D
computer images

lOMoARcPSD|1377855
Comparing light and electron microscopes

NB: the overall design is similar but e microscope is upside down
Light microscope Electron microscope
Source LED light Electron gun
Lenses (NB: lenses in Light focused with glass lenses E beam focused with magnets
both have the same
name)
Specimen Can be living Non-living, placed in a vacuum
Viewing Image viewed directly with Image viewed on screen or
eyes photographic filn
Preparing cells to view under an e microscope

 Fixation = kill and preserve living tissue (same as above)
 Sectioning = cutting sample into extremely thin slices so that e can penetrate
through
o Specimen dehydrated and permeated with monomeric resin
o Solidified block cut by fine glass or a diamond blade in a microtone
 Plating = thin, water-free sections are placed on a small circular metal grid for
viewing
 Metal shadowing = a thin film of heavy metal (eg: platinum or gold) is evaporated
onto the dry specimen (done at an angle to create “shadows”)
o Reveals the surface of the specimen- creates a 3D picture

lOMoARcPSD|1377855
Cell Culture
 In vitro = (in glass) growing cells in glass or plastic, outside a living organism
 Important for scientists to communicate ideas and discoveries with the general
public- in order for understanding and not scare
Applications
 Used in contemporary medicine to generate tissue for patients (eg: burn victims) so
that bodily rejection dose not occur (ie: patient tissue used as donor tissue)
o Epithelial cells taken from patient and grown in culture
o Cells will form confluent mass (uniform layer of cells)
o Cells will form matrix (keratin) around mass
o Tissue then able to be used as a graft over burns (autograph)
 Used to observe cell function- different cell types placed in culture to see how they
respond (individual cells observed as well as their interaction with other cells)
 Used to model diseases (visualization)
 Used to generate immortal cell lines
o Commercially available
o Keep frozen early passage cells
o Examples: BY2 tobacco cells (plant), HeLa (animal)
Source of Cells
 Source = any cell from an organism (any cell can be used as long as it is prepared
correctly
 Organ homogenized (broken down)
 Cell isolated- treated enzymatically to release cells from extracellular matrix and cell
wall (plant cells only)
 Cell purified with buffer and centrifuged to separate into layers- the least dense
component will float on top of mass and can be extracted
 Example: collagenase used to break down the placenta (extracellular matrix) and the
trophoblast cells plated out
Culture Vessel
 Different cell types require different growing conditions
 Many cells will fall to the bottom of plastic and stick (confluent growth)
 If cells grow on flat surface- need a matrix to encourage growth eg: collagen or a
component of extracellular matrix
Aseptic Conditions
 Sterile conditions must be used to prevent the growth of microbes on the cell culture
o Bench cleaned with 70% ethanol
o Hands washed in 70% ethanol and gloves worn
 Experiment must be carried out under a Laminar flow or Class II hood to prevent
microbe contamination (air rotates within chamber)

lOMoARcPSD|1377855
Growth Requirements
NB: Conditions dependent on the particular cells
 Growth medium:
o Used to bath and feed cells
o Usually red in colour- pH indicator
o Usually contains (animal cells):
– Glucose (or other organic nutrients)
– Growth factor (serum)
– Vitamins
– Inorganic salts (Na+, K+)
– Antibiotic and antifungal agents
 Keeping the cells warm (animal cells):
o 1 hr at 37o allows the cells to attach
o 5% CO2
o Plants require light
Cellular Membrane Proteins

Plasma Membrane
 Entire cell and intracellular compartments are enclosed by plasma membranes
(phospholipid bilayer)
 Hydrophobic phospholipid tails face inside the membrane while the hydrophilic
sugar heads remain on the outside
 Sugar heads are able to interact with water (hydrophilic)
 Barriers and compartments allow evolving cells to have specialization (higher
complexity)
 The cell is jam-packed with proteins, which all attract water ∴ the inside of the cell is
a very viscous gel
 Due to the high viscosity, cells with holes will leak very slowly- holes can be
punctured by microinjection, electrical barrage or protease (enzyme that destroys
membrane proteins)
 Functions of the membrane:
o Receives cell signals eg: endocrine, hormones
o Regulates the import/export of molecules eg: proteins, hormones
o Maintains shape and size
o Allows movement and expansion
The Lipid Bilayer

 Proteins are jam-packed into membrane
 Alpha helices give proteins rigidity
 Hydrophobic regions lock into membrane while
hydrophilic regions interact with heads
 Linker proteins connect membrane-bound proteins to
ensure they remain close together (not embedded in
membrane ∴ may be entirely hydrophilic)
 Proteins not linked are mobile and fluid

lOMoARcPSD|1377855
 Types of membrane lipid:

o Phospholipid
– Polar head, hydrophobic fatty acid tails
– Double bonds will kink fatty acid chains, preventing close
packing
o Glycolipid
o Cholesterol
– OH top has polarity (H is ionized off to form O-)
– Ring structures and side chains are hydrophobic
– Cholesterol slots into spaces (created by fatty acid kinks)
between phospholipids to produce more efficient packing
 OH head will interact with polar phospholipid head
 Hydrophobic end embedded in membrane
– Side chain is cleaved by cytochrome P450
 Double bonds in fatty acid tails influence the thickness of the
membrane
o Unsaturated fatty acid chains are more spread apart,
causing thinner membranes
o Exclusively saturated fatty acid chains are fully
extended, causing thicker membranes
 Lipid raft = a region of saturated fatty acid tails causing a thick membrane section
 Membranes will form in the most energetically favourable configuration ie: to hide
hydrophobic tails from water
Membrane Proteins
 Transporters allow molecules to pass through the membrane- often powered by ATP
 Linker proteins connect membrane-bound proteins together
 Receptors receive extracellular and intracellular signals
 Enzymes catalyze the conversion of substrate to product

lOMoARcPSD|1377855
Different ways proteins can slot into membrane

(1) Transmembrane/lipid anchor
(2) Triple (or more) pass transmembrane
(3) β sheet barrel
o Example: Porin
– Evolved in bacteria
– Allows the passage of ions through membrane
– Present in mitochondria and bacteria
(4) Non-penetrating α helix
Connected membranes
 Anchor proteins join membranes
 Proteins in the outer membrane can be linked to proteins in the inner membrane:
 By adapter (anchor proteins)
 Directly
Excitable Cells
 Excitable cell = a cell with the capacity to depolarize upon stimulation ie: changes
ionic gradient- more negative on the inside, more positive on the outside (eg:
neurons and muscle cells)
The membrane has channels and pumps

 Proteins slot into membrane (hydrophobic within membrane, hydrophilic poke out)
to regulate the passage of molecules in/out of cells
 Channel proteins have specificity- allow only certain molecules to pass
through membrane
 Dog door theory (now discredited) = specificity due to size

lOMoARcPSD|1377855
 Many substances can be transported through membranes via channels and pumps:
o NB: fats require endocytosis (membrane to escort molecules in)
Passive and Active Transport

 Passive transport = movement through membrane that does not require energy
o Movement down concentration gradient- ie: molecule is pushed through
membrane from a region of high conc to low conc
o Conformation change in transport protein induced by molecule binding, not
the hydrolysis of ATP
 Active transport = movement through membrane requiring energy
o The hydrolysis of ATP used to push substances through membrane against
their concentration gradient
Conformational Change- moving down conc gradient

 Molecule is picked up by protein on one side of the membrane
 Binds to the solute-binding site, inducing a conformational change in the protein that
allows it to open on the other side of the membrane
 Molecule is released on the other side of the membrane
 Solute-binding site empty, allowing protein to resume original conformation

lOMoARcPSD|1377855
Powering transporters- moving against conc gradient

 NB: when moving down conc gradient the electrochemical gradient pushes the
molecule through the membrane but when moving against conc gradient, another
source of energy is required
 Coupled transport = one molecule moves through membrane down conc gradient
and ‘piggy backs’ the other against its conc gradient
 ATP-driven pump = energy from the hydrolysis of ATP  ADP (ADP then carried to
the mitochondria to be phosphorylated back to ATP)
 Light-driven pump = light energy used to open pump
The Sodium Potassium Pump

 Located in neurons (excitable cells- depolarized)
 Anti-porter = pushes Na and K in opposite directions against their conc gradient
 Depolarizes cell- 3 Na+ pumped out, 2 K+ pumped in- making cell more positive on
the outside and more negative on the inside (due to Cl- and negative proteins)
 Hydrolysis of ATP required for energy
 Response to stimulus:
o Neurotransmitter (eg: ACh) released by one neuron into synapse
o Binds to membrane-bound receptor on target neuron
o Receptor-coupled channel opens, causing Na (high conc of Na on outside of
cell) to rush in
o Na attracted towards negative charges inside cell, generating an electric
current which opens channels along axon
o More Na rushes into cell
 Pump mechanics:
o Na binds to specific Na-binding sites in
channel
o Pump is phosphorylated (ATP  ADP)
causing a conformational change that
closes the cytosol opening and opens
the extracellular opening
o Na is released into extracellular space
o K on outside binds to specific K-binding
site in channel
o Phosphate is released, causing channel
to resume original conformation
o K is released into the cytosol

lOMoARcPSD|1377855
Types of transport
 Uniport = one molecule transported through membrane
 Symport = two molecules transported through membrane in the same direction
 Antiport = two molecules transported through membrane in opposite directions
Calcium pump
 NB: alpha helices provide rigidity to proteins
 Ca pumped from cytosol into the lumen of the sarcoplasmic reticulum (ER in the
skeletal muscle)
 Ca down into channel (between alpha helices) until it cannot move any further
 Aspartic acid is phosphorylated (ATP  ADP) causing a conformational change that
opens up the alpha helices at the bottom of the channel, allowing Ca to pass through
Epithelial Cells
 Epithelium = the thin tissue forming the outer layer of a body's surface and lining
the alimentary canal and other hollow structures
o The only water-proof cells
o Act as a barrier
o Packed with proteins that regulate the transport of molecules in/out
Specificity Filter (bacterial K+ channel)

 NB: ions in solution are surrounded by water molecules (hydration)- contributes to
the physical size of the ion
 Oxygen molecules (negative) that surround the pore will
interact with cations (positive)
 Oxygen molecules are positioned EXACTLY so that they
interact perfectly with K and not Na
o K will have a charge interaction with all 4 O
o Na will not (slightly smaller)- only 2 O

lOMoARcPSD|1377855
 NB: ion will swap water molecules in favor of molecules in interactive selectivity
loop
Gated Channels
 Eg: voltage dependent channels (present in nervous tissue)
Receptor coupled ion channel

 The binding of ligand to receptor opens up channel
Patch Clamping
 Patch Clamping = a technique used to study voltage-gated channels
 Cell held against a small vacuumed pipette and a small piece of
membrane is taken off
 Current flowed through membrane to study the opening/closing of
channels
 Produces patch clamp trace:

lOMoARcPSD|1377855
Mitochondria and Steroid Hormones

Basic Morphology
 NB: mitochondria is more complex than basic
morphology diagrams suggest
 Folding membrane = large surface area
 Mitochondria and closely associated with
microtubules- used a vesicle train tracks that
carry cholesterol to the mit
 Join well with other mit
 Contains its own DNA
Fission and Fusion

 Fission = mitochondria splitting in half
 Used in the process of reproduction
 Promoted by fzo1 (fuzzy onion gene)
 Fusion = mitochondria joining together
 Forms networks- filamentous, tubular
structures
 Promoted by dynam GTP binding protein
 Conformation changes induced by different
chemical/hormonal signals
Steroidal Tissue
 Steroid hormones are only produced in certain tissues:
o Brain
o Gonads
o Adrenal
o Placenta
o Retina
Cholesterol Metabolism
 Steroid Hormone = a ring structure hormone formed in the body from cholesterol
(eg: estrogen, testosterone, cortisone, and aldosterone)
 Progesterone = a steroid hormone released by the corpus luteum that stimulates
the uterus to prepare for pregnancy
 Cholesterol is used as steroid hormone substrate- brought into cell by LDL receptor
as LDL, covered in a clathrin coated vesicle
 Free cholesterol (broken down in lysosomes) is carried to the mitochondrion
 StAR protein (steroidogenic acute regulatory protein) then transports hydrophobic
cholesterol from the outer to the inner mitochondrial membrane
 Cytochrome P450scc cleaves the side chain off cholesterol to form pregnenolone,
which is then converted to progesterone in the smooth ER
 ACTH stimulates new protein synthesis that enables steroidogenesis

lOMoARcPSD|1377855
StAR Transporter Protein

 Aids the transport of cholesterol from the lumen into the inner mitochondrial
membrane
 START hydrophobic region able to bind to cholesterol
 Accelerates cholesterol transfer
 Protein import occurs at the contact site between the outer and inner
mitochondrion- proteins form a bridge between mit membranes to join them
together
The Effect of Dihydrotestosterone on the reproductive system

 Dihydrotestosterone = the active form of testosterone
 If StAR is mutated, then patient can not produce steroid hormones
o Genitals unable to develop from vaginal pouch during development-
intermediate sex
o Congenital adrenal hyperplasia (mutation of hormone gene)
o Loss of control over salt and carb balance

lOMoARcPSD|1377855
Plant Cells and Development

The Totipotency of Plant Cells
 Totipotency = the ability the produce an entire new organism (totipotent is the
adjective)
 Fertilized eggs of both plants and animals are totipotent
 Animal cells loose their totipotency early in development, while plant cells
remain totipotent
 Determination = an embryonic cell that has become committed to a particular
specialized path of development (ie: fixed fate)
Differentiation = process by which the cell undergoes change into a specialized cell
type
o Determination always precedes differentiation and can occur without
differentiation taking place (happens early on in development in animals and
is generally irreversible and stable)
o As animal cells loose their totipotency, they become determined to develop
into a specific type of cell that becomes fixed
 Animal cells have more than 200 different cell types while plants have only 14 major
cell types (structures are simpler)
o Example: Vascular, Ground, Dermal
 In plants, the process of determination is very weak/less stable and can vary rapidly
according to the situation
 Stable determination is possible, but rare. Examples:
o Phase changes in woody plants (some have juvenile and adult forms)
– Juvenile and adult forms have different structures
– Juvenile forms cannot make flowers (not sexually mature)
– A cutting from a juvenile plant will generate a juvenile while an adult
cutting will generate an adult form
– An adult will revert to juvenile form following meiosis
– Example: Ivy
o Apex of Osmundia fern

– Shows transition from pinnate juvenile leaves to phyllodes (leaf stalk)
in the adult form
– When dissected off and cultured, the primordial (bumps) closest to the
apex will form shoots, closest to the ground will form leaves and the
ones between will vary (as they are not yet determined)

lOMoARcPSD|1377855
 Unstable determination is more common in plant cells- will change developmental

pathway more rapidly to form new cells, shoots or entire new organisms (ie: are
more flexible). Examples of totipotency:
o Plant tissue culture
– A section can be taken from the tip of an old tree (adult plant) and then
cultured to form an entire new plant
– Tissue culture is used to produce transgenic plants
o Regeneration after wounding
– If cut, sections will grow back
Life Cycles
 Plants have alternation of generations
 Haplotype and diplophase stages
Potential Immortality of Plant Cells

 Plants don’t separate germ cells (gamete) from somatic cells- all are the same
 In animals, somatic cells are terminally differentiated and only sex cells have the
potential for immortality
 At any time or place, plants can vegetatively reproduce (ie: cuttings to form a whole
new plant)- potential immortality
 Consequences of somatic variation
Structure of the Plant Body

 Simple structure
 Modular structure of the plant as repeated units
 Determinate verses indeterminate growth
Size of the Body

 Plant body as a leaf support scaffold to hold out leaves to sunlight
 Aim to produce large surface area for photosynthesis
 Can be huge or small (variety)
Growth Habit
 Plants are immobile (fixed in one place)
o Able to make complex organic nutrients from inorganic sources via
photosynthesis (autotrophy)

lOMoARcPSD|1377855
o Limited nutrient supply- can only access what is in their immediate

environment
 Changes form in response to environmental changes (phenotypic plasticity)- eg:
wind, water, gravity, location of nutrients, etc
 Capacity for repair (can’t run away from danger)
Photosynthetic Ability
 Contain chloroplasts and light harvesting machinery
 Produce their own nutrients
Reproduction and Dispersal

 Vegetative reproduction (cuttings from one plants used to produce another)
 Sexual reproduction- pollen dispersal
 Fruits with seeds- animals will eat and disperse seeds
Genetic Systems
 Local adaptation- asexual reproduction
 Wide range of environments in which zygotes (eg: seed dispersal) are deposited- no
selection in migration
Internal Communication
 Routes
o Via extracellular compartments- cell walls, intercellular spaces and xylem
o Via continuity of cytoplasm through intercellular connections (the symplast)
o Phloem
 Signals
o Receptor serine/threonine kinases function as cell-surface receptors in plants
o Plant hormones act as long-distance signals
Model Plant: Arabidopsis thaliana

 Small genome (26,000 genes)
 DNA sequence completely known
 Range of molecular tools available
 Short life cycle (6 weeks)
 Mutational analysis
Plant Cell Walls and Polarity

Structure of Plant Cell Walls (extracellular matrix)
 Primary cell wall:
o Primary cell wall = a thin, flexible and extensive layer formed while the cell is
growing and developing
o Matrix composed of microfibrils (60 cellulose molecules), hemicellulose,
pectins and glycoproteins
o Recently deposited microfibrils form parallel arrays

lOMoARcPSD|1377855
 Lamella:
o Lamella = the middle layer between primary and secondary walls
o Formed first from the cell plate during cytokinesis
 Secondary cell wall:
o Secondary cell wall = a thick layer formed inside the primary cell wall after
the cell is fully grown
o NB: not all cells have a secondary wall
o Material is invested into developing the second wall, rather than developing
the cell
o Composed of a range of additional compounds that modify their mechanical
properties and permeability
Formation of New Cell Walls

 New cell walls are formed between 2 daughter nuclei during cytokinesis
o Microtubules deliver vesicles containing cell wall material to the mid-plane
between daughter nuclei
o Vesicles fuse to form an outwardly expanding cell plate
o Plate fuses with parent cell walls, dividing the cell in 2
o The plane of division is precisely predicted by a preprophase band of MTs
(disperses before division but serves as a memory site)
 Additional material is added to the cell wall via:

o Cellulose synthetase complexes inserted into the cell membrane
o The secretion of golgi derived vesicles containing cell wall materials
Consequences of the Cell Wall for Growth and Development

 Plant cells are fixed in position relative to their neighbors ie: can’t migrate (whereas
animal cells are able to migrate around the body)
 The cell wall dictates the shape of the cell (protoplasts are spherical without cell
walls). The resultant shape of the plant depends on:
o Directed cell expansion
o Precise planes of division
 Turgor pressure (high conc of water inside cell due to high conc of solutes) pushes
cell wall outwards
 Because the microfibrals (MF) are precisely aligned and inelastic, turgor pressure
causes cells to form an axis
 MT are involved in the parallel deposition of MF (but exactly how is unknown)
 If the alignment of MF is disrupted (eg: by drug), cell cannot organize itself to form
an axis
Example: the disruption of MT in Arabidopsis root inhibits elongation and stimulates
root swelling (seen in prac)

lOMoARcPSD|1377855
Plant Polarity
 Polarity in plants first develops in zygotes and embryos
 The hormone Auxin is transported down the plant (from cell to cell) in a polar
fashion (from apex to base), stimulating the formation of stems and roots
 High concs of Auxin in a cell stimulate the formation of roots while low concs of
Auxin stimulate the formation of shoots
 Polarity is innate and ∴ plant cuttings will retain polarity (ie: shoots will form
shoots), regardless of gravity
The Polar Transport of Auxin

 Cells are arranged to transport auxin in a polarized fashion (apex  base)
 Cell wall pH = 5, cytoplasm pH = 7 (difference maintained by a proton pump)
 Inside cell wall, auxin is in IAAH form (auxin associated with a proton)- it is
hydrophobic and ∴ can freely diffuse across membrane through the influx
transporter AUX
 Once inside the cytosol, the pH changes from 5  7, causing IAAH to ionize to form
IAA- (hydrophilic) meaning it is trapped in the cell
 H is pumped back into cell wall
 At basal end, PIN transporter protein channel exports IAA- through membrane
where it re-associates with H (due to lower pH)
 Examples of changes in plant polarity:
o Lateral branch formation
o Primordia formation
o Wound response
Chemical Signals in Plants

 Plants contain many chemical signals that control their growth and development
Short range chemical signals between plant cells

 Receptor Serine/Threonine Kinases function as cell-surface receptors in plants (eg:
LRR receptor kinases- leucine rich repeat)
 Plants have a huge variety of serine/threonine kinases (unlike animal cells)
 These receptors have a cytoplasmic domain and an extracellular ligand binding
domain (like animal cells)
 Example of LRR receptor kinase complex: Clavata1/Clavata2 complex (Clv1/Clv2)
o Clv1/Clv2 complex is activated by the binding of Clv3
o Cells in the outer layer of the growing tip secrete the small peptide Clv3
o Cells in a more central region of the growing tip express the Clv1/Clv2
receptor complex
o Once Clv3 has bound, Clv1 phosphorylates associated receptor proteins,
activating Rho family GTPases
o This signaling pathway suppresses meristem growth by inhibiting cell
division or stimulating cell differentiation- leading to large flower and shoot
meristems

lOMoARcPSD|1377855
Long distance chemical signals in plants: plant hormones

 Involved in the coordination of developmental events in separate parts of the plant
 Hormones may be produced in a variety of organs at various stages of development
(not produced by specialized glands)
 May be transported some distance to the site of action or be produced within
affected cells
 A large variety of responses are produced in different tissues and at different stages
of development
 There are 6 groups of plant hormone:
(1) Gibberellin
(2) Auxin
(3) Ethylene
(4) Cytokinin
(5) Absciscic acid (ABA)
(6) Brassinosteriod
Auxins (eg: IAA- Indole Acetic Acid)

 Aux conc influences response in plant:
 In stems, low conc of auxin stimulates cell elongation.
 In roots, high conc of auxin inhibits elongation
 Transported through plant in a polar fashion
 Affect on growth (hypothesis):
o Auxin causes cells to pump hydrogen ions into cell wall (explained above-
linked to change in pH)
o Increase in H causes the pH in the cell wall to decrease, activating enzymes
that break cross-links between cellulose fibers in the cell wall
o Cellulose fibers loosen and allow the cell to expand as turgor pressure inside
the cell pushes against the cell wall
 Auxin stimulating pathway:
o In the absence of auxin:
– A transcriptional repressor protein (Aux/IAA), binds to auxin-
response factor (ARF), preventing the transcription of auxin genes

lOMoARcPSD|1377855
o In the presence of auxin:

– Auxin binds to the auxin receptor protein
– The receptor-auxin complex recruits the ubiquitin ligase complex,
which degrades Aux/IAA
– ARF is then left free to activate transcription of auxin target genes
 Auxin controls phototropism

o Phototropism = the orientation of the plant towards (positive) or away from
(negative) light
o Auxin accumulates on the shaded side of stems, stimulating elongation and
bending towards light
o Auxin efflux inhibitor blocks phototropism and auxin redistribution
 Auxin controls gravitropism
o Gravitropic bending of a root or shoot results from differential growth on
upper and lower sides of the root or shoot
o In shoots, high auxin conc stimulates elongation ∴ that the bottom side grows
faster and the shoot bends upwards
o In roots, high conc of auxin inhibits elongation ∴ bottom side grows slower
and roots bend down
 Other affects of auxin:

o Defers leaf fall
o Auxin stimulates root formation (rooting power)
o Apical dominace- inhibits growth of axillary buds

lOMoARcPSD|1377855
Gibberellins
 Plant hormones that promote growth, seed germination and leaf expansion
 There are >100 different gibberellins, although individual species only produce a few
each
 The active compound gibberellin acid (GA1), is the endogenous (internal origin)
active gibberellin that causes stem elongation in many plants
 Dwarfed varieties of plants usually lack the genes for the synthesis of gibberellins
 Gibberellins and seed germination
o The endosperm = a region of the seed that stores nutrients (protein and
carbohydrates) for the developing plant embryo)
o Gibberellins break seed dormancy and stimulate germination
o GA1 stimulates the release of hydrolytic enzymes, which breaks down
starches and proteins in the endosperm
Cytokinins
 Plant hormones that stimulate cell division/cytokinesis
 Delays leaf senescence (loss of ability to grow and develop- age)
o Opposing action of auxin- direct application of cytokinin promotes the growth
of auxiliary buds
 Stimulates stomatal opening
Abscisic acid (ABA)

 A growth inhibitor
 Often antagonistic to action of other hormones (eg: GA in seed germination)
 Assists in the toleration/avoidance of adverse conditions eg: drought, salinity or low
temp
 ABA and drought resistance:
o In drought, ABA closes stomata to reduce water loss
o Efflux of K+ from guard cells followed by osmotic loss leads to a decrease in
guard cell turgor closure of stomata
 ABA and breaking of seed dormancy (inaction):
o ABA levels increase during seed maturation, enabling embryo to survive
dehydration
o The breaking of dormancy is associated with a decline in the level of ABA
o Dormancy only broken by specific environmental cues, ensuring that seeds
germinate only under suitable conditions of moisture, light and temp
Ethylene
 The only gaseous plant hormone (C2H4)
 Involved in plant responses to environmental stresses (eg: flooding, drought,
infection, wounding, mechanical pressure)
 Influences a wide range of developmental processes (eg: shoot elongation, flowering,
seed germination, fruit ripening, leaf abscission and senescence)
 Ethylene stimulates MT to reorient from transverse to longitudinal- changes the
direction of cell expansion

lOMoARcPSD|1377855
Chemical signals between plants

 Volatile compounds may be released from plants on wounding, particularly insect
herbivory
 Volatile compounds may stimulae defence responses in distant parts of that plant or
even adjacent plants
 Volatile compounds my attract specific predators of plant herbivores
 Inhibitors of germination may be released to limit competition for resources
Phytochrome & blue light receptors

Plants and light
 Plants are unable to move ∴ need to be able to adapt to environmental conditions
 The most important environmental influence on plants is light:
o Used as an energy source (photosynthesis)
o Used to regulate their orientation and development
 Plants have evolved a set of light-sensitive proteins to monitor the quality, direction
and duration of light- photoreceptors
 Photo-proteins sense light through a light-absorbing chromophore that causes a
conformational change in the protein
 Examples:
o Phytochrome- responds to red and far-red light
o Blue light receptors- phototropin, chryptochromes and zeaxanthins
The Phytochrome Pigment

 Phytochrome = a light-sensitive protein kinase responsible for many light
responses in plants.
 Phytochrome stimulated responses:
o Stem elongation to reach light- plants in dark environment (eg: buried in soil)
will elongate until they reach light (above ground)
o Seed germination- allows plant to germinate only when light environment is
optimal for seed survival
o Timing of flowering- to allow maximum flower exposure to light
o Shade avoidance- stimulates shaded plants to elongate away from a shaded
position or to grow away from a potential shading source
 Pigment exists in two inter-convertible forms:
(1) Pr (red light absorbing)
o Absorbs at 660nm
o The inactive form ie: does not activate genes
o Present in dark-grown seedlings
o Exposure to red light will convert Pr  Pfr
(2) Pfr (far-red absorbing)
o Absorbs at 730nm
o The active form ie: activates genes
Eg: germination genes
o Exposure to far-red light will convert Pfr  Pr

lOMoARcPSD|1377855
 Example: Arabidopis mutants

o Wild-type stem elongation is inhibited by exposure to white light, leaves
begin to develop at apex
o Mutants grow tall in white light with no leaves
o Mutants are deficient in one or more functional phytochrome, preventing
seedlings from responding to white light
Phytochrome: mechanism of action

 Red light stimulates a conformational change that converts Pr  Pfr
o Molecule autophosphorylates
o Change exposes the nuclear localization sequences (NLS)
 NLS sends most Pfr into the nucleus, where it is able to phosphorylate other proteins
to regulate gene expression
 Some phytochrome stays in the cytoplasm where it mediates rapid response
 A cascade of events is stimulated by light
 Far red light will convert Pfr back to Pr, causing it to stop regulating genes and exit
the nucleus
 Example:
o Reach optimal light source = exposure to red light = stop elongation
o Shaded from light = far-red light = elongate to reach light
Other examples of phytochrome controlled responses

 Chloroplasts may move/spin to face dim light to maximize photosynthesis or shy
away from strong light to protect pigments
 Regulates sleep patterns of leaves
 NB: some phytochrome responses involve ion fluxes eg: movement
o Movement generated at the base of leaflets (pulvini)
o K and Cl are pumped from one side of leaf to another by phytochrome
o Water follows movement of ions (osmosis)
o Turgor pressure pushes pulvini open/closed based on water distribution (due
to ions)

lOMoARcPSD|1377855
Blue light receptors

 Phototropin
o Stimulates photoprism (eg: phot 1)
– A serine/threonine kinase
– Stimulates plant to grow towards blue light
o Stimulates chloroplast migration
– Movement driven by actin
– In low light, chloroplasts accumulate on the surface of cell towards the
light
– In high light, chloroplasts migrate to the sides of cells
 Cyptochrome
o Inhibits stem elongation
– Signals to cells that optimal light source has been reached- time stop
elongation and exert energy into flowering and germination etc
 Zeaxanthin
o Induces stomatal opening
– Stomata open in the light and close in the dark (due to photosynthesis
in guard cells but also due to blue light)
– Blue light applied on top of a saturating red light also leads to
additional opening
– Blue light stimulates ion uptake into guard cells
– Without Zea, blue light induced stomatal opening is lost
Cell Junctions
Types of Cell Junctions
Junction Type Animal Cells Plant Cells
Anchoring = including  Adherens junctions (c-c)  Protoplast wall
both cell-cell (c-c) and cell-  Desmosomes (c-c) connections
matrix (c-m) adhesions  Focal adhesions (c-m)
 Hemidesmosomes (c-m)
Occluding = seal the gaps  Tight junctions  Apoplastic barriers
between epithelia cells to  Septate junctions (endodermis)
create a semi-permeable or
impermeable barrier
Communicating = create  Gap junctions  Plasmodesmata
passageways linking the
cytoplasm’s of adjacent
cells

lOMoARcPSD|1377855
Anchoring Junctions: Animal cells

 Anchoring junctions are present in tissues subject to mechanical stress (eg: heart
muscle or skin epithelium)
 Two types:
(1) Cell-cell adhesion = connect cytoplasm of a cell to its neighbor’s
(2) Cell-matrix adhesion = connect cytoplasm of cell to the extracellular matrix
 Adherens Junctions
o An actin-linked, cell-cell adhesion
o Bundles of actin filaments inside two adjacent cells are connected by
transmembrane proteins (cadherins)
o Protruding from each cell- cadherin repeats, joined by flexible hinge regions,
form chains that arrange in parallel lines
o Ca2+ binds to flexible hinge regions to prevent flexing and promote straight,
parallel formation
o Adhesion between the two is formed by the N-terminal cadherin repeat-
generally bind homophilically
o Cadherins are linked to actin filaments (inside the cell) via β-catenin,
p120-catenin and other anchor proteins
o In epithelial cells they form a continuous belt below the tight junction
o Contraction of actin filaments at adhesion belts orchestrates the
formation of epithelial tubes
 Desmosomes
o An intermediate filament-linked (eg: keratin), cell-cell adhesion
o Cadherin adhesion proteins (desmoglein and desmocollin) form parallel
chains between adjacent cells
o Just inside each plasma membrane sits a dense plaque of anchor proteins,
which is attached to intermediate filaments
o Cadherin tails protrude through the plasma membrane and connect to anchor
proteins plakoglobin and plakophillin
o Anchor proteins connect to desmoplakin, which connects to intermediate
fillaments

lOMoARcPSD|1377855
 Hemidesmosomes
o An intermediate-filament linked, cell-matrix adhesion
o Integrin and collagen XVII proteins span the membrane
o Integrin is linked to intracellular anchor proteins dystonin and plectin and
extracellular laminin and collagen
o Anchor proteins bind to intermediate filament keratin
o NB: in epithelial cells, intermediate filament networks are connected to one
another via desmosomes and to the basal lamina via hemidesmosomes
 Focal Adhesions
o An actin-linked, cell-matrix adhesion
o Uses transmembrane integrins
Summary

lOMoARcPSD|1377855
Anchoring Junctions: Plant cells

 It is not clear whether similar junctions exist in plant cells due to the structural role
of the cell wall
 HOWEVER plasmolysed cells stay connected to the cell wall via hechtian strands
Occluding Junctions: Animal cells

 Tight Junctions (vertebrates)
o An extracellular molecules on one side of an epithelium is prevented from
crossing by the tight junction that seals adjacent cells
o They are impermeable to macromolecules and are variably permeable to
small molecules
o Transporter proteins are specific to nutrients and are confined to different
regions of the cell membrane- permits a directional transfer of nutrients
across the epithelium without entry of others
o Sealing strands consist of long rows of transmembrane proteins that are
embedded in the plasma membrane of both adjacent cells
o Example: gut epithelial cells- help contents within gut lumen whilst allowing
nutrients to be transported into the blood
 Septate Junctions (invertebrates)

o Adjacent membranes are connected by parallel rows of junctional proteins
Occluding Junctions: Plant cells

 Apoplastic barriers:
o Plants have barriers within the cell wall, which have a similar function to tight
junctions
o These barriers consist of bands of impermeable material, suberin, that is
deposited within cell walls
o In roots, there is a permeability barrier in the cell wall between the cortex and
central vascular tissue- prevents the diffusion of any compound in the soil
into the xylem and up into the plant

lOMoARcPSD|1377855
Communication Junctions: Animal cells

 Gap Junctions:
o The plasma membrane of each is penetrated by protein assemblies called
connexons (composed of 6 subunits)
o Two connexons join across the intercellular gap ells to form a continuous
aqueous channel that connects the two cells
o NB: connexins = monomer, connexon = 6 connexions, intercellular channels =
2 connexons
o Rapidly closed (reversible) by high cytoplasmic Ca, changes in pH and
intercellular voltage differences
Communication Junctions: Plant cells

 Plasmodesmata:
o The cytoplasm of adjacent cells is continuous through plasmodesmata across
the cell wall
o Surrounded by a tube of cell membrane
o ER is continuous through plasmodesmata (desmotubule)
o Formation of Plasmodesmata:
– Primary- formed at cytokinesis as vesicles containing wall material
fuse and entrap ER
– Secondary- form in existing walls via a close association of the ER with
the cell membrane
o Function of Plasmodesmata:
– Transports water, ions, metabolites and larger molecules (such as
proteins, transcription factors and RNA) from cell to cell
– NB: more plasmodesmata = more cells are electrically connected
o Composition of Plasmodesmata:
– Actin, myosin, tropmyosin, centrin, cellulose, HSP70
o Regulation of Plasmodesmata transport:
– Closed by high cytoplasmic Ca
– NOT closed by pH changes
– Closed by rapid turgor pressure changed
– Closed by callose deposited between the wall and membrane

lOMoARcPSD|1377855
Cellular Endocrinology
 Pituitary = a pea sized endocrine gland, attached to the base of the brain, that is
responsible for secreting hormones involved in growth, development and the
functioning of other hormones
 Hypothalamus = a region of the forebrain below the thalamus which coordinates
both the autonomic nervous system and the activity of the pituitary. It controls body
temperature, thirst, hunger, and other homeostatic systems, and is involved in sleep
and emotional activity
 Hormones released by the brain (and other organs) trigger cellular signals eg: CRH
 NB: the only aa’s that can be phosphorylated in proteins are- tyrosines, serines and
thronines
Hormones Trigger Cell Signals: α-interferon vs TGF-β

 α-interferon (IFNα)
o Blocks viral infection in cells by triggering resistance proteins
o NB: uses a similar signaling system to prolactin, erythropoietin and growth
hormone
o Signaling pathway (JAK-STAT pathway):
– The binding of INFα to its cell-surface receptor activates the JAK-STAT
intracellular signaling pathway
– Once INFα is bound, receptor subunits dimerize, bringing the
associated JAKs close enough to cross-phosphorylate each other
– Activates JAKs the phosphorylate receptors (on tyrosines)
– This attracts STAT1 and 2 to dock on to the receptor where they are
phosphorylated by JAK
– Phosphorylated STATs dissociate from receptors and dimerize
– STAT dimer travels into the nucleus where it binds to DNA and other
regulatory proteins to activate gene transcription

lOMoARcPSD|1377855
 TGF-β (transforming growth factor β group)

o A superfamily of hormones/mediators
o Regulates genes and proteins involved in development, extracellular matrix
development and differentiation
o Receptor has two types (type I and II) which are structurally similar
homodimers
o Signaling pathway (Smad-dependent):
– TGF- β binds to a type-II receptor, causing it to recruit and then
phosphorylate a type-I receptor
– Phosphorylated type-I attracts and phosphorylates Smad 2 or 3
– Smad 2 or 3 opens up, allowing it to dimerize with Smad 4
– The Smad complex enters the nucleus where it recruits other gene
regulatory proteins to activate the transcription of specific genes
 Major differences between α-interferon and TGF-β signaling pathways:

o α-interferon triggers viral infection resistance proteins whereas TGF-β
triggers developmental proteins
o α-interferon receptor requires JAK to phosphorylate whereas TGF-β receptor
has built in phosphorylation
o α-interferon binds to two receptors whereas TGF-β binds to one (which then
recruits the other)
Corticotrophin Releasing Hormone (CRH)

 Hormone produced in the hypothalamus
 Contains 41 aa
 Has evolutionary homology to proteins in fish and frog eggs
 Involved in stress response and pregnancy
 Brief signaling pathway:
o Stress causes hypothalamus to release CRH

lOMoARcPSD|1377855
o CRH travels down the hypophyseal portal system

(blood vessels connecting hypothalamus and
pituitary) to the pituitary gland
o ACTH is released at the pituitary
o Travels through the blood stream to the adrenal gland
(located above the kidney)
o Cortisol (made in the cortex of the adrenal medulla) is
then released, causing the body to break down fat and
protein supplies as a mechanism to deal with stress
o Cortisol then inhibits the release of more CRH
 POMC (proopiomelanocortin) is the pro-hormone (pro-curser) from which ACTH
and a variety of other active molecules are produced
NB: ACTH and β-endorphin are always co-secreted
Increased CRH levels in pregnant women

 CRH is also secreted by the placenta during pregnancy
 Pregnant women’s pituitary becomes desensitized to CRH due to high levels,
resulting in high concs of CRH but not as high concs of ACTH and cortisol
 CRH acts as a time-clock- as [CRH] increases, it begins to stimulate time-regulating
proteins (stimulates MPAK signaling pathway)
Example: contraction/relaxation of uterus smooth muscle
 At 6 months uterus opening is contracted to hold baby in while stomach
muscle is relaxed to allow baby space to grow and move
 At 9 months stomach muscle becomes contracted to push baby out while
uterus muscle becomes relaxed to allow exit
Cyclin Dependent Protein Kinases

 M phase = mitosis + cytokinesis
 S phase = synthesis of DNA
 G1 = (gap 1) growth and
development
 G2 = more growth and development
 G0 = a state of no progression
(cyclins disappear)- can be
permanent, semi-permanent or
temporary
 NB: many checkpoints along the way

lOMoARcPSD|1377855
Checkpoints
 One event does NOT trigger the next event in line- central contract ensures that the
cell is ready to take the next step (eg: DNA replicated, enough protein, big enough,
etc)
 Checkpoints:
o G2 checkpoint (before M begins)
– Is all DNA replicated?
– Is cell big enough?
o M checkpoint (trigger anaphase and proceed to cytokinesis)
– Are all chromosomes attached to the spindle?
– Has DNA divided exactly between two daughter cells?- start
cytokinesis
o G1 checkpoint (before S begins)
– Is cell big enough?
– Is environment favorable? (eg: enough glucose, temp, not crowded)
– Is DNA damaged?
 NB: no G1 and G2 in embryonic cells- rapidly divides and synthesizes to put down a
scaffold of cells
Cyclin Dependent Protein Kinases

 Cyclin phosphorylates Cdks, causing a conformation
change that exposes Cdk’s active site to form an active
complex
 Without cyclin, Cdk is inactive
 Complex drives the cell cycle- phosphorylates other
proteins to stimulate progression through the cycle
M-phase Promoting Factors (MPFs)

 Push cells into mitosis
 A protein kinase-cyclin complex
 Phosphorylates lamin proteins (located under nuclear membrane to reinforce it) in
the nuclear envelope, causing them to break down (phosphorylated lamin
monomers lose attraction to one another, falling apart)

lOMoARcPSD|1377855
 MPF Activity:
o Causes chromosomes to condense
o Causes MTs to form spindle
o Phosphorylates lamins- nuclear envelope breaks down
o NB: requires cyclin to be active
 Found by a bioassay (any assay that studies a biological action)
 Relationship between cyclin and MPFs- protein expression in embryonic cells

o Cyclin and MPF conc linked
o Critical conc of cyclin reached to drive mitosis
Phosphorylation
 Phosphorylation = a reaction in which a phosphate molecule is covalently added to
another
 Proteins are activated by phosphorylation- causes a conformation change
 Cyclin activity increases steadily during interphase
 NB: Phosphates can sometimes be inhibitory
Activation of MPF
 Cdk will associate with M-cyclin as the
levels of M-cyclin gradually rise
 This conformation change will expose
Cdk’s activating site
 The inactive M-Cdk complex is then
phosphorylated by both Cdk-activating
kinase (CAK) (activating phosphate)
and Cdk-inhibitory kinase (Wee1)
(inhibitory phosphate)
NB: it is the inhibitory phosphate that prevents the M-Cdk from becoming active
 The inactive M-Cdk complex is then activated at the end of G2 by the phosphatase
Cdc25, which removes the inhibitory phosphate
 Cyclins are then degraded by proteases and signal disappears (temporary signal)

lOMoARcPSD|1377855
Cdpk Heterogenity
 There are different Cdks and different cyclins for different parts of the cell cycle (eg:
S-Cdk complex initiates the synthesis of DNA in S phase)
Arresting the cycle

 Cdk inhibitor proteins (eg: p27, p21, p16) inactivate Cdk complexes to stop
progression
 Binds to both the cyclin and Cdk, covering the active site of the Cdk and preventing
activity
 Allows the breaks to put on the cell cycle rapidly when there is a problem eg:
environment not favorable or DNA is damaged (a much faster mechanism than
relying on cyclin conc to decrease)
 Production of inhibitory proteins:
o If DNA is damaged, p53 will be activated
o Active p53 binds to the regulatory region of thee p21 gene
o P21 is transcribed and translated, then inactivates Ckd-complex
o Cell cycle stops
Cell Control Checkpoints

lOMoARcPSD|1377855
Control of Cellular Proliferation

Normal Cells
 Most normal cells are mortal- will reproduce N times in their lifespan
 Once cell has reproduced N times, apoptosis is initiated
 Some cells are immortal- eg: cancer cells, cell lions (man made)
 Cell senescence = a period where the cell is alive but no longer able to divide
 Cell able to keep track of N by telomeres (get smaller with every replication)
 The balance of signals will determine the cell’s fate- demolition (apoptosis) vs repair
(keep dividing)
The production of Immortal Cells (Monoclonal Antibodies)

 Immortal cells are very useful:
o Never die- can be tested and retested
o Share majority of characteristics with normal cells
 Monoclonal antibody = fusion of antibody producing cell with tumor like cell
 Specimen (eg: mouse) immunize to stimulate antibody production (antibody =
protein produced in response to a specific antigen)
 Antibody forming cells are isolated from the spleen and broken down into single
cells (with collagenase and protease)
 Fused with tumor-like cell (ie: is immortal) to form hybrid
 Hybrids screened for antibody production- antibody producing are cloned
Apoptosis
 Apoptosis = controlled cell suicide, which is a normal part of the cell’s growth and
development process
 Allows neat, orderly, sequential degradation of the cell with minimal damge to
neighboring cells
 Stages:
(1) Cell reduces in size
(2) Cytoskeleton and nucleus contents break down and cell becomes ill-shapen
(blebbing)
(3) Cell contents packaged into apoptotic bodies
(4) Bodies engulfed and digested by phagocytes (phagocytized)
 Can be used to sculpt tissue:
o Example: finger formation- fetus’ have webbed fingers 5 weeks post
conception
o Webs were sculpted away by apoptosis of web cells to produce individual
fingers
o Requires a balance of signals- keep tissue vs sculpt away
 “Trimming the fat after covering the bases”
o Embryonic cells have no G1 and G2 stages- continuous replication
o Allows embryo to produce all the necessary cells quickly
o Overcompensates the number of cells needed, allowing embryo to only retain
the best developed cells and sculpt away tissue

lOMoARcPSD|1377855
Proteolysis
 Proteases = enzymes that breakdown proteins into amino acids (process termed
proteolysis)
 Caspases = any group of protease that mediates apoptosis
 An important part of the apoptotic mechanism (allows the cell to be broken down)
 Activation of proteolysis cascade:
o Each caspase is initially made in the
inactive pro-protein form
o Inactive pro-domain is cleaved to form
the active protease
o The first activated protease (initiator)
will then cleave and activate many more
(amplification cascade)
o Each level of the cascade has a specific
region of the cell that it will degrade (eg:
cytolistic proteins or nuclear lamin)
Activation of Intrinsic Apoptotic Pathway

 Cytochrome C (CC) is located in the mitochondrion inter-membrane space
 In the absence of an intrinsic apoptotic signal, Bcl-2 binds to BH123 transporter
proteins, preventing the release of CC
 In the presence of an apoptotic signal, Bcl-2 becomes inactive, allowing BH123
proteins to aggregate and release CC into the cytosol
 CC binds to and activates Apoptosis acting factor (Apaf-1), which cleaves and
activates caspase 9, initiating cascade
Activation of Extrinsic Apoptotic Pathway

 Fas ligand on the surface of a killer lymphocyte (small white blood cell) activates Fas
death receptors in the surface of the target cell
 The cytolistic tail of the Fas receptor then recruits the adaptor protein FADD, which
binds to and activates caspase 8 or 10
 Cascade is then initiated
Confluence and Contact Inhibition

 Confluence = when cells have grown over the entire surface of a plate, forming a
confluent layer of cells that can no longer proliferate
 Contact inhibition is stimulated and cells go into a state of senescence

lOMoARcPSD|1377855
Cancer
Cancer Cells
 Cancer = a disease caused by the uncontrolled division of abnormal cells in one (or
more) part of the body
NB: can not occur in bacteria- bacterial cells grow and divide rapidly
 Cells have an enlarged nucleus- potentially containing too many chromosomes
 Cells ignore the social etiquette of normal cells (ie: disobey contact inhibition- keep
dividing)
Benign and Malignant Cancers

 Benign = a disease which is not harmful in effect
o Could be slow growing
o Adenoma = connective tissue surrounding cancer cells- easily contained in
the body (wont enter bloodstream) and removed surgically
o Example: a mole (although if cancer cells break through basal lamina, mole
may turn malignant)
 Malignant = an uncontrollable and harmful disease
o Cells make break free from adenoma, spreading around the body
 Metastasis = the ability for cancer cells to detach, enter the bloodstream and
develop secondary growths at other regions of the body
o Cancer cells express surface proteases (eg: type-IV collagenase) which break
down basal lamina and allow motility
Proliferation
 Factors that slow normal cell proliferation:
o Growth control- checkpoints, less Cdks produced or inactivated
o Activate apoptosis
o Activate senescence
 NB: growth control, apoptosis control and senescence control must be lost for cell to
become cancerous

lOMoARcPSD|1377855
 Phases of cancerous development:

o Dysplasia (cell moving and growing abnormally)
o Adenoma (cancer cells encased in connective tissue)
o Non-infiltrating carcinoma (cells attached to each other and not moving)
o Infiltrating carcinoma (express surface enzymes which break the connective
tissue, allowing cells to leak into the blood stream)  metastatic
 Mutations can accumulate
Proto-oncogenes and Oncogenes

 Proto-oncogenes = any gene that regulates the cell cycle (normal gene)
 Oncogenes = a mutated proto-oncogene, which can transform a cell into a tumor cell
 Example:
o Ras- monomeric GTP binding protein with its own GTPase activity- splits
gamma phosphate GTP  GDP
o Function- switches on other proteins, which turn on genes that regulate cell
growth, differentiation and survival eg: MAP-kinase, which regulates
proliferation
o Inactive when GDP is bound, active when GTP is bound
o If Ras is mutated (locked in on position and not able to turn self off) it would
stimulate uncontrollable cell growth ie: proto-oncogene  oncogene
Tissues
Tissue Types
 Epithelium = covering
 Connective tissue = support
 Muscle = movement
 Nervous tissue = control
 NB: the gut contains all classes of
tissue

lOMoARcPSD|1377855
Connective Tissue
 Connective tissue = any tissue that connects, supports, binds or separates other
tissues or organs. Consists of relatively few cells embedded in extracellular matrix
(matrix = fibers + ground substance)
 Requirements:
o Ground substance- often proteoglycans (carbohydrates and proteins) and
interstitial fluid
o Cells
o Fibers
 Includes- bone, cartilage and loose connective tissue
 Slowest healing- lower levels of vacuolization
Nervous Tissue
 Composed of neurons (impulse-conducting cells able to send and receive signals)
 Excitable- able to be polarized (Na/K channel)
 Three main parts:
o Cell body (soma)- contains the nucleus, site of
protein synthesis
o Dendrites- signals are received
o Axon- neurotransmitters are released at the axon
terminal into the synapsis
Muscle
 Specialized for contraction
 Three types:
o Smooth (cross fibers)
o Skeletal (parallel fibers)
o Cardiac (parallel fibers)
Epithelium
 Epithelium (plural epithelia) = the sheet of cells covering the outer surface of a
structure or lining a cavity
 Epithelial membrane = a multicellular sheet composed of epithelium with a
connective tissue underlay (basal lamina)
 Epidermis = the epithelial layer covering the outer surface of the body (NB:
different structures in different animal groups)
 Layer types:
o Simple (single layer)
o Stratified (multiple layers)
o Pseudo stratified (one layer of cells that appears to be 2- due to 2 rows of
nuclei)

lOMoARcPSD|1377855
 Cell shapes:
o Columnar
o Cuboidal
o Squamous
 Tight junctions between cells (cross-linking proteins) prevent water and other
molecules from passing through the barrier
 Cells are polarized (basicalapical)- due to tight junctions
 Cells are susceptible to damage ∴ have multiple mitochondria to produce ATP to
drive processes
Mammalian Skin
 Fibroblasts- lay down fibrin tissue (for growth and repair)
 Collagen- aids with elasticity of skin
 Dermis = the thick layer of connective below the epidermis, containing blood
capillaries, nerve endings, sweat glands, hair follicles, and other structures (rich in
collagen)

lOMoARcPSD|1377855
Changes in tissue
 Sports injury- cartilage (connective tissue) often heals slowly because it has no blood
vessels
 Cartilage becomes less elastic with age
 Tissue repair- when skin is wound fibroblasts lay down new fibrin tissue (fibrosis)-
the mix and amount depends on the severity of the wound. Macrophages then eat
away the excess tissue
 Scar tissue- contains lots of connective tissue (strong, not as flexible), too much
fibrin laid down for macrophages to eat away
 Basal cell carcinoma form in the dermis
The Extracellular Matrix (ECM) and

Connective Tissue
Extracellular Matrix
 Extracellular matrix = fibers and ground substance that surround the cell (NB:
matrix = fibers + ground substance)
 Composed of glycoproteins, proteoglycans and collagen fibers
 In connective tissue, the main stress-bearing component is the ECM (ie: ECM is the
defining feature of connective tissue)
 Most important while animal is an embryo- supplies nutrients, provides support, fills
up space left by moving cells, provides a scaffold for new cells to grow, attracts
growth factors and hormones
 Contains many negative charges (due to proteoglycans), which attract water-making
the ECM a gel-like substance
Collagen Fibers = strength

 Fibroblasts = proteins found in connective tissue that lay down fibrin, used to
construct new and repair wounded tissue
 Collagen = a fibrous protein rich in glycine and proline that provides strength to the
ECM in animals (surrounded by gel-like substance)

lOMoARcPSD|1377855
 Type I: skin, tendon and bone, type II: cartilage, type IV: basal laminae
 Composition- makes collagen STRONG
o Single-stranded polypeptide chain = glycine, proline (X) and hydropoline (Y)
o Three chains would into triple helix termed a fibril
o Bundles of fibrils make up a fiber
o Fibers are arranged at right angles to each other
 Collagen is cross-linked with other proteins to enhance strength and limit stretch-
hyper-extensible skin is caused by the improper collagen crosslinking
 Adaptor proteins attach actin filaments (just inside the plasma membrane) to
collagen fibers in the extracellular matrix
o Integrin: goes through (integrated) the plasma membrane
o Adaptor protein: joins actin-integrin
o Fibronectin: joins integrin-collagen
Proteoglycans
 Fill up spaces between collagen
 Negatively charged
 Composition (bristles on a brush):
o Long GAG chain
o Linker proteins attach core proteins
perpendicular to chain
o More GAGs stem off core proteins

lOMoARcPSD|1377855
 Glycosaminglycan (GAG) = a carbohydrate polymer composed of repeated

disaccharide (2 sugars) monomers
NB: COO- contains the negative charge, which attracts the salt (and ∴ water)
 Glycoprotein vs proteoglycan- both are proteins with carbohydrates attached BUT
glycoproteins have higher % protein whereas proteoglycans have higher % carb (ie:
sugar)
 Types of proteoglycans
o Heparan Sulfate- used in developmental signal proteins
o Elastin- providing elastic fibers
Development
The Development of a Multicellular Organism
 An animal or plant begins its life as a single cell (that contains all its genetic
information) and develops into a multicellular organism (that contains many
specialized cells)
 Development involves 4 essential processes
(1) Cell proliferation = producing many cells from one
(2) Cell specialization = creating cells with different characteristics at different
positions
(3) Cell interactions = coordinating the behavior of one cell with that of its
neighbors
(4) Cell movement = rearranging the cells to make structured tissues and organs
NB: in a developing embryo, all these processes occur at once in different ways, in
different parts of the cell
 s
Gastrulation
 Gastrulation = the stage of embryogenesis during which the embryo is transformed
from a ball of cells into a structure with a gut
 Layer of embryonic tissue and what they become:
o Ectoderm = embryonic epithelial tissue that is the precursor of the epidermis
and nervous system (skin)

lOMoARcPSD|1377855
o Endoderm = embryonic tissue that is the precursor of the gut and associated
organs (gut)
o Mesoderm = embryonic tissue that is the precursor to muscle, connective
tissue, skeleton and many internal organs (everything in between)
o NB: unlike humans, in sea urchin the ectoderm does not form the CNS
 Basic anatomical scheme of development:
o One cell (fertilized egg) divides to form many smaller cells
o These cohere to create an epithelial sheet facing the external medium
o Most of the sheet remains external, forming the ectoderm
o A part of the sheet tucks into the interior, forming the endoderm
o Another group of cells move into the space between the ectoderm and
endoderm, forming the mesoderm
o This process by which a ball of cells develops into a structure with a gut is
called gastrulation
Determination
 Determination = an embryonic cell that has become committed to a particular
specialized path of development (eg: become a neuron or a liver cell)
NB: proceeds differentiation
 A cell’s state of determination can be tested by transplanting it into different
environments and observing the outcome- a determined cell will remain on the same
developmental pathway whereas a non-determined cell will develop as a new cell
Morphogens
 Morphogen = an embryonic signal molecule that can impose a pattern on a field of
cells by causing cells in different places to adopt different fates
 Induction = a change in the developmental fate of one tissue caused by an
interaction with another tissue (eg: the passing of a morphogen from one cell to
another)
 A group of cells at one end of the embryo become specialized as a signaling center
and will secrete morphagens
 This protein will spread out from its source, creating a morphagen gradient
 The responding cells will adopt different cell fates in accordance with their position
in the gradient
 Example: the influence of Bicoid protein in drosophila embryogenesis (fruit fly)
o The bicoid protein a morphagen which influences the formation of the head
and tail
o Embryonic cells at the tip of the embryo that are determined to become head
cells will secrete bicoid (signaling center)
o The protein is dissipated from cell to cell via gap junctions, creating a bicoid
gradient (ranging from a high conc in head region to a low conc in the tail
region)
o Cells will develop differently depending on their position in the gradient

Notes For Biology

Uploaded by

Document Information

Original Title

Copyright

Available Formats

Share this document

Share or Embed Document

Sharing Options

Did you find this document useful?

Is this content inappropriate?

Copyright:

Available Formats

Notes For Biology

Uploaded by

Copyright:

Available Formats

lOMoARcPSD|1377855

Complete lecture notes for BIOL2016

Cell Biology (University of Sydney)

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

How Cells Communicate

 Extracellular signal molecules bind to receptor molecules and initiate a response

Signal molecules bind to specific receptors

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

o When bound to by a signal molecule, the receptor becomes activated and

Forms of Intercellular signaling

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

G Proteins and Second Messengers

The Plasma membrane

G-protein-coupled receptors (GPCRs)

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

GTP binding proteins

The GPCR Signaling complex

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

o The activated cat-subunits can then phosphorylate specific target proteins

How to detect new G Proteins in the cell- mitochondria

Phospholipase and Calcium Signaling

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

How does hypothalamus stimulate pathway in pituitary cell

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

The Synthesis of PIP2

Phospholipase C (PLC) cleaves PIP2 into IP3 and DAG

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

G Protein Coupled Receptor Activates Phospholipase C (PLC)

Effects of forskolin on β-endorphin/ACTH release

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

Ca2+ Function as a Ubiquitous Intracellular Mediator

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

Evolution of the Nucleus

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

Vesicular Transport- budding and fusion

Gated Transport- nuclear pores

Transmembrane Transport- nuclear import receptors

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

Ran GTPase imposes directionality on transport

Ran’s influence on nuclear import receptors

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

Protein import into the mitochondrion by TIM and TOM

Intracellular Vesicular Transport

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

Road map of the cell

 Red arrows indicate inside  out (exocytosis)

Vesicle Coat Proteins

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

 Three types of protein, each used for different transport step

Pulse Chase Experiments

Clathrin Coat Structure

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

Assembly and Disassembly of a Clathrin Coat

Dynamin pinches off budding vesicles

Formation of a COPII-Coated Vesicle

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

process of membrane curvature

Tethering of a vesicle to a target membrane

Endocytosis of low-density lipoprotein (LDL)

Distributing prohibited | Downloaded by Michelle Lee (thecutemango@gmail.com)

The Dynamic Microtubules

The Role of the Cytoskeleton