Cell structure and function

Cells have evolved two basic architectural plans

1. Cells without a nucleus = Prokaryotes (can also be spelled procaryotes)
o includes the Bacteria and Archaea
o generally very small, unicellular
o the earliest and still most abundant life forms; evolved ~ 4 billion
years ago
o some species highly evolved pathogens: e.g. Borrelia burgdorferi,
the cause of Lyme disease.

2. Cells with a nucleus = Eukaryotes (eucaryotes)

o include the Animals, Plants, Fungi, and Protists
o some unicellular, some multicellular forms
o evolved ~ 1 billion years ago
o size ranges from tiny yeasts to giant sequoias, dinosaurs

Eucaryotic Cell Structure and Cell Components

 Diagram of a Eucaryotic cell -- see text for identification

Eucaryotic cells are organized into different compartments

 Compartments bounded by membranes (to be discussed in Ch. 8)
 Cytoplasm = central metabolic compartment, bounded by cell membrane.
Other compartments inside cytoplasm are called organelles
 Compartmentation allows specialized functions to be carried out in
different locations

The cytoplasm: site of protein synthesis and many metabolic events

 Contains many ribosomes = particles on which proteins are synthesized

o Ribosome size measured in Svedberg (S) units; derived from
 sedimentation in ultracentrifuge (used before electron microscopes
were available)
o Prokaryotes: ribosomes made of 30S and 50S subunits, assemble
into 70S ribosome
o Eukaryotes: ribosomes made of 40S and 60S subunits, assemble
into 80S ribosome
o In bacteria, ribosomes occupy 25% of cell volume, use 90% of cell
energy. Less in many specialized eukaryotic cells, but still the
dominant activity of almost all cells.
 Contains many enzymes for general metabolism
 Compartment in which foodstuffs enter and from which wastes leave cell
 Contains fiber of the cytoskeletal system, which organize cytoplasmic
structure
 Contains many different organelles

The Nucleus: locus of DNA & RNA synthesis and protein assembly
 Contains chromatin = DNA-protein complexes. Chromatin can condense
into chromosomes during cell division
 Site of RNA synthesis. 80% of RNA = ribosomal RNA. Remaining 20%
leaves nucleus as t-RNA & m-RNA, directs protein synthesis
 Contains nucleolus = assembly plant for ribosomes. Ribosomal proteins
are made in cytoplasm, must be transported back into nucleus. Ribosomal
RNA is made in nucleus. These two elements are integrated inside
nucleolus to create ribosomal subunits. These are then exported out of
nucleus through nuclear pores.
 Bounded by nuclear membrane = double layered structure. Contains
many nuclear pores, allow material to move in and out of nucleus
 Nuclear Pores have octagonal "doors" made of protein; open and close on
either side depending on specific signals. Pore has diameter of about 10
nanometers (10 x 10-9 m), smaller than diameter of a complete ribosome.
Pore can open up to as much as 26 nm in response to certain signals. Some
signals allow motion in but not out, other signals control reverse transport.

The Endomembrane system: moving materials into different

compartments
Endomembrane system = set of interconnected compartments: endoplasmic
reticulum (ER), Golgi body, lysosomes, cell membrane
 Endoplasmic Reticulum
o Rough ER: synthesizes proteins for export or movement to
different cell compartments (but not to cytoplasm).
o Signal hypothesis: certain mRNAs encode proteins designated for
export. These carry a peptide signal at growing end, causes growing
protein to move to ER ("docking"), insert peptide into membrane,
translocate growing polypeptide chain across ER membrane. When
protein synthesis is complete, polypeptide folds up inside ER, not in
cytoplasm.
 o Smooth ER (sER): synthesizes lipids, detoxifies drugs and poisons
(in liver).

 Golgi body
o functions as intracellular "post office" for sorting new proteins
made on rER.
o Vesicles containing protein pinch off from ER, fuse with cis face of
Golgi. Inside Golgi, oligosaccharide chains on proteins are
modified. Vesicles pinch off from trans face of Golgi, carry
proteins to several possible destinations: export (out of cell),
lysosomes, peroxisomes, cell membrane, etc.
 Lysosomes
o compartments to break down old proteins, foreign materials, many
wastes.
o Contain ~40 hydrolytic enzymes: lipases, proteases, nucleases, etc.
Break down organic polymers of all types.
o "Suicide bags" if opened up on cell itself = apoptosis.
o Lysosomes are used in phagocytosis, a process in which foreign
materials are brought into the cell and "chewed up".
 Cell membrane (aka plasma membrane)
 Vacuoles
o large membrane compartments (contrasted with small membrane
bags called vesicles).
o Plant cells have especially large vacuole called the central vacuole,
can occupy most of the volume of a plant cell. Stores pigments,
wastes, water, poisons, and more

Organelles involved in energy transformations are separate from the

endomembrane system
 "Energy organelles" have unique properties:
1. are enclosed by double membrane system
2. contain DNA and ribosomes (70S, not 80S like cytoplasmic
ribosomes)
3. make some of their own proteins
4. from their own genes
5. divide by binary fission (but not autonomous, cannot
6. grow or sustain life outside of cell)
 Mitochondria = centers for respiratory catabolism. Oxygen combined
with chemicals to break down foods, generate cell energy. Contain outer
and inner compartments, with many membranous cristae that "criss-cross"
the internal space. Found in virtually every eukaryotic cell. Small
structures similar to bacteria in some size.
 Chloroplasts = centers for photosynthetic anabolism. Belong to group
of plant organelles called plastids. Include chloroplasts (photosynthesis),
amyloplasts (store starch), chromoplasts (store pigments). Trap light,
convert energy to sugars (+ CO2, water). Contain stacks of thylakoids,
 where green pigmented chlorophyll is embedded in membrane to trap light.

Endosymbiont theory: All organelles seem to share many properties with

bacteria: contain 70S ribosomes (whereas rest of eukaryote cells contain 80S
ribosomes), divide by binary fission, contain circular DNA without nucleus, etc.
Lynn Margulis proposed endosymbiont hypothesis: that organelles derived from
ancient colonization of large bacteria (became the eucaryotic cell) by smaller
bacteria (became the mitochondria, chloroplast, etc.) Symbiosis = "living
together". Eventually, organelles lost ability to exist as separate organisms, cannot
be separated from cell. Recent evolutionary taxonomy by comparing ribosomal
RNA shows that this idea has lots of merit. Mitochondrial and plastid ribosomes
are very similar to current bacteria, very different from eukaryotes.
Build a cell (Campbell website activity)

Cytoskeletal system provides internal fibrous structure to cells

Cell is not "just a bag in a bubble". Lots of internal fibers = internal "skeleton".
Not rigid like bone; capable of being assembled, broken down in minutes. Allows
cell movement, cell division, internal motion of compartments.
 Microtubules
o Largest diameter fiber. Found in cytoplasm of all eukaryotes.
o Involved in many structures: cilia, flagella (9+2 arrangement);
spindle fibers that polymerize from centrioles during
mitosis/meiosis.
o Made of tubulin protein; polymerizes into hollow tubules 25 nm
diameter.
o View cell treated with anti-tubulin fluourescent antibody
o Cilia and flagella: organelles of locomotion. Contain 9 double
rings of microtubules, 2 central microtubules.
o View micrographs showing structure of cilia
o View SEM of cilia on surface of epithelial tissue
o Two motor proteins allow motion along microtubules
1. Motor protein 1 -- Dynein
 side arms allow one tubule to "walk up" along
neighboring tubule, cause bending (powered by
ATP).
 animation of microtubule motion (Campbell website
activity)
 Causes motion from positive (+) end of the
microtubule (where new tubulin is added to the
microtubule) toward the minus (-) end of the
microtubule.
 When coordinated, this causes rotating or whip-like
motion of entire cilium or flagellum --->motion.
 View animation of dynein pulling vesicle along a
microtubule
2. Motor protein 2 -- Kinesin
  Also powered by ATP, also allows protein to move
along microtubule
 Causes motion from negative (-) end of the
microtubule toward the positive (+) end of the
microtubule (where new tubulin is added to the
microtubule).
 pulls things toward outer reaches of cell
 Example: in nerve cells, kinesin pulls vesicles away
from center towards nerve endings.
 View animation of kinesin pulling vesicle along a
microtubule
 View more sophisticated animation of kinesin
moving along a microtubule. (Select either the
Quicktime or MPEG movie titled "Structural
Analysis of the Kinesin Motor Protein" ) )
 Microfilaments (= actin)
o another kind of fiber. Found in cytoplasm of most eukaryotes.
o nvolved in muscle contraction, cell support, pinching off of
daughter cells after mitosis (in animals), cytoplasmic streaming (in
plants).
o View TEM of microvilli    (protected) of the striated border of
epithelial tissue. Microfilaments within them can be seen extending
down into the terminal web, an aggregate of fine filaments lying in
the cell cytoplasm.
o View animation showing "treadmilling" of actin -- subunits are
added at one end, removed at the other, producing net migration of
the filament in one direction.
o View movie showing cell movement (left panel) and assembly of
actin (right panel) (2.8 Meg file)
 Intermediate filaments
o a third kind of fiber.
o Made from keratin subunits. Not so quickly assembled and
disassembled as microtubules or microfilaments.
o May be involved in resisting tension, reinforcing cell shape, fixing
location of nucleus.

Cell walls provide rigid structure around cells

 Found in plants, fungi, bacteria -- not in animal cells
 Allow cells to survive in plain water, rigid structure (tree towering 150 feet
high!)
 Thicker than cell membrane. Made from cellulose (plants and fungi), other
polysaccharides (bacteria).
 View plant cell walls    (protected)
 Cells maintain contact by plasmodesmata -- thin cytoplasmic connections,
lined by membrane, that pass across cell wall junctions.
 Cells don't end at their outer membrane; they possess an
extracellular matrix (ECM)
 Animal cells don't have walls, but do have ECM = meshwork of
macromolecules outside plasma membrane. Consists mainly of
glycoproteins (proteins with oligosaccharide chains), especially collagen.
 Some cells attached directly to ECM by bonding to collagen or fibronectin.
 View diagram of ECM   (protected)

Cells are joined by a variety of intracellular junctions

 In multicellular organisms, adajcent cells are held together by several types
of specialized junctions.
1. Tight junctions (found in animals): specialized "belts" that bind
two cells tightly to each other, prevent fluid from leaking into
intracellular space. View animation of tight junctions (Campbell
website activity)
2. Desmosomes (found in animals): intercellular "rivets" that create
tight bonds between cells, but allow fluids to pass through
intracellular spaces.
View animation of desmosomes (aka anchoring junctions)
(Campbell website activity)
3. Gap junctions (found in animals): formed by two connecting
protein rings embedded in cell membrane of adjacent cells. Allows
passage of water, small solutes, but not macromolecules (proteins,
nucleic acids).
View animation of gap junctions (aka communicating junctions)
(Campbell website activity)
4. Plasmodesmata (found in plants): channels connecting cells; allow
free passage of water and small solutes, but not macromolecules
(proteins, nucleic acids).
View figure showing plasmodesmata (Campbell website activity)

Self-quiz: Cells

Copyright 2002 Thomas M. Terry

The University of Connecticut
Instructions: You may answer as many or as few questions as you wish, in any order, and
you may take the test as often as you wish. When you are ready to check your
performance, scroll to the bottom of this document and click the 'Submit' button. All
multiple-choice questions you answered will be graded, and your score will be shown at
the bottom of the exam. Short answer questions will not be graded, but the instructor's
answers will be shown next to yours.
1) There are many small particles visible in electron micrographs of all cells. Chemical
analysis shows that they contain both RNA and protein. What are they?
a. chromatin
b. membrane fragments
c. ribosomes
d. centrioles
e. microtubules

2) Some of the proteins made by rough ER become part of:

a. ribosomes
b. the cytoplasm
c. membranes
d. the cytoskeleton
e. all of these

3) In the figure above, the nucleolus is represented by structure:

 a. a
b. n
c. j
d. l
e. d

4) With reference to the same figure, a mitochondrion would correspond to structure

____ :
a. a
b. n
c. j
d. l
e. d

5) Still refering to this figure, where would proteins be synthesized?

a. region 'o'
b. region 'g'
c. region 'l'
d. all of the above
e. both (a) and (b), but not (c)

6) An important function of lysosomes is:

a. phagocytosis
b. the synthesis of proteins for export from the cell
c. the transfer of information from the nucleus to the cytosol
d. the synthesis of chromatin
e. intracellular motion

7) Organelles such as mitochondria are different from elements of the endomembrane

system in that:
 a. mitochondria don't have membranes
b. mitochondria don't contain any proteins
c. mitochondria contain their own DNA and ribosomes
d. mitochondria are involved in the photosynthetic process
e. all of the above

8) One of the major functions of rough endoplasmic reticulum is

a. synthesis of phospholipids
b. synthesis of steroids
c. synthesis of secretory proteins
d. synthesis of cytoplasmic proteins
e. all of the above

9) In eucaryotes, ribosomes are assembled in/on:

a. the nucleolus
b. the cytosol
c. rough endoplasmic reticulum
d. smooth endoplasmic reticulum
e. mitochondria

10) The endomembrane system involves a process of membrane movement by pinching

off and fusion of membranous compartments in the cell. Which of the following
sequences would you expect to observe, if you could accurately follow a particular bit of
membrane as it traveled through the endomembrane system?
a. rER --> sER --> Golgi --> plasma membrane
b. Golgi --> rER --> sER --> plasma membrane
c. plasma membrane --> Golgi --> rER --> sER
d. rER --> sER --> plasma membrane --> Golgi
e. Golgi --> plasma membrane --> sER --> rER
 Submit Query Reset

Animal Cells
This schematic represents an idealized animal cell, e.g., a liver cell. The columns to the
left and right of the labels contain links to discussions of the particular structures.

Intermediate Golgi apparatus

filaments
Pinocytotic
Plasma vesicle
membrane
Actin filaments
Peroxisome
Glycogen
Vacuole granules

Lysosome Smooth
endoplasmic
Nucleolus reticulum

Centrioles Microtubules

Nucleus Ribosomes

Nuclear Mitochondrion
envelope
Rough
endoplasmic
reticulum

Cytosol
 Index to this page
Welcome&Next
Search  Actin Filaments
 Intermediate Filaments
 Microtubules
o Microtubule motors
o The Centrosome
 Centrosomes and Cancer
 Centrioles
o Cilia and Flagella

 Primary Cilia

The Cytoskeleton
Cells contain elaborate arrays of protein fibers that serve such functions as:
 establishing cell shape
 providing mechanical strength
 locomotion
 chromosome separation in mitosis and meiosis
 intracellular transport of organelles

The cytoskeleton is made up of three kinds of protein filaments:

 Actin filaments (also called microfilaments)
 Intermediate filaments and
 Microtubules

Actin Filaments
Monomers of the protein actin polymerize to form long, thin fibers. These are about 8 nm
in diameter and, being the thinnest of the cytoskeletal filaments, are also called
microfilaments. (In skeletal muscle fibers they are called "thin" filaments.) Some
functions of actin filaments:
 form a band just beneath the plasma membrane that
o provides mechanical strength to the cell
o links transmembrane proteins (e.g., cell surface receptors) to cytoplasmic
proteins
o anchors the centrosomes at opposite poles of the cell during mitosis
o pinches dividing animal cells apart during cytokinesis
 generate cytoplasmic streaming in some cells
 generate locomotion in cells such as white blood cells and the amoeba
 interact with myosin ("thick") filaments in skeletal muscle fibers to provide the
force of muscular contraction

Intermediate Filaments
 These cytoplasmic fibers average 10 nm in diameter (and thus
are "intermediate" in size between actin filaments (8 nm) and
microtubules (25 nm)(as well as of the thick filaments of
skeletal muscle fibers).

There are several types of intermediate filament, each

constructed from one or more proteins characteristic of it.

 keratins are found in epithelial cells and

also form hair and nails;
 nuclear lamins form a meshwork that stabilizes the
inner membrane of the nuclear envelope;
 neurofilaments strengthen the long axons of neurons;
 vimentins provide mechanical strength to muscle (and
other) cells.

Despite their chemical diversity, intermediate filaments play

similar roles in the cell: providing a supporting framework
within the cell. For example, the nucleus in epithelial cells is
held within the cell by a basketlike network of intermediate filaments made of keratins.
(photo at right)

In the photo (courtesy of W. W. Franke), a fluorescent stain has been used to show the
intermediate filaments of keratin in epithelial cells.

Different kinds of epithelia use different keratins to build their intermediate filaments.
Over 20 different kinds of keratins have been found, although each kind of epithelial cell
may use no more than 2 of them. Up to 85% of the dry weight of squamous epithelial
cells can consist of keratins.

Microtubules
Microtubules
 are straight, hollow cylinders
 have a diameter of about 25 nm
 are variable in length but can grow 1000 times as long as they are thick
 are built by the assembly of dimers of alpha tubulin and beta tubulin.
 are found in both animal and plant cells

Microtubules
 grow at each end by the polymerization of tubulin dimers (powered by the
hydrolysis of GTP), and
 shrink at each end by the release of tubulin dimers (depolymerization)

However, both processes always occur more rapidly at one end, called the plus end. The
other, less active, end is the minus end.
 Microtubules participate in a wide variety of cell activities. Most involve
motion. The motion is provided by protein "motors" that use the energy of
ATP to move along the microtubule.

Microtubule motors
There are two major groups of microtubule motors:
 kinesins (most of these move toward the plus end of the microtubules) and
 dyneins (which move toward the minus end).

Some examples:
 The rapid transport of organelles, like vesicles and mitochondria, along the axons
of neurons takes place along microtubules with their plus ends pointed toward the
end of the axon. The motors are kinesins.
Charcot-Marie-Tooth disease. One cause of this rare disorder is an inherited
mutated gene for one of the kinesins. In these patients, axonal transport is
defective (which probably accounts for their muscle weakness first occurring in
muscles at the ends of the longer motor neurons).
 The migration of chromosomes in mitosis and meiosis takes place on
microtubules that make up the spindle fibers. Both kinesins and dyneins are used
as motors as we shall see below.

In plant cells, microtubules are created at many sites scattered through the cell. In animal
cells, the microtubules originate at the centrosome.

The Centrosome
The centrosome is
 located in the cytoplasm attached to the outside of the nucleus.
 Just before mitosis, the centrosome duplicates.
 The two centrosomes move apart until they are on opposite sides of the nucleus.
 As mitosis proceeds, microtubules grow out from each centrosome with their plus
ends growing toward the metaphase plate. These clusters of microtubules are
called spindle fibers.

The photo (courtesy of Tim Mitchison) shows microtubules growing in vitro from an
isolated centrosome. The centrosome was supplied with a mixture of alpha and beta
tubulin monomers. These spontaneously assembled into microtubules only in the
presence of centrosomes.

Spindle fibers have three destinations:

 Some attach to one kinetochore of a dyad with those growing from the opposite
centrosome binding to the other kinetochore of that dyad.
 Some bind to the arms of the chromosomes.
  Still others continue growing from the two centrosomes until they extend between
each other in a region of overlap.

All three groups of spindle fibers participate in

 the
assembly of
the

chromosomes at the metaphase plate at metaphase. Proposed mechanism (the

diagram shows only 1 and 2):
1. Microtubules attached to opposite sides of the dyad shrink or grow until
they are of equal length.
2. Microtubules motors attached to the kinetochores move them
 toward the minus end of shrinking microtubules (a dynein);
 toward the plus end of lengthening microtubules (a kinesin).
3. The chromosome arms use a different kinesin to move to the metaphase
plate.
 the separation of the chromosomes at anaphase.

o The sister kinetochores separate and, carrying their attached chromatid,

o move along the microtubules powered by minus-end motors, dyneins,
while the microtubules themselves shorten (probably at both ends).
o The overlapping spindle fibers move past each other (pushing the poles
farther apart) powered by plus-end motors, the "bipolar" kinesins.
o In this way the sister chromatids end up at opposite poles.

Other Functions of Centrosomes

In addition to their role in spindle formation, centrosomes play other important roles in
animal cells:
 signaling that it is o.k. to proceed to cytokinesis. Destruction of both centrosomes
with a laser beam prevents cytokinesis even if mitosis has been completed
normally.
  signaling that it is o.k. for the daughter cells to begin another
round of the cell cycle; specifically to duplicate their
chromosomes in the next S phase. Destruction of one centrosome
with a laser beam still permits cytokinesis but the daughter cells
fail to enter a new S phase.
 Segregating signaling molecules (e.g., mRNAs) so that they pass
into only one of the two daughter cells produced by mitosis. In this way, the two
daughter cells can enter different pathways of differentiation even though they
contain identical genomes. [Link to further discussion.]
 In at least some developing neurons, the position of the centrosome establishes
the point at which the axon will grow out.

Centrosomes and Cancer

Cancer cells often have more than the normal number (1 or 2 depending on the stage of
the cell cycle) of centrosomes . They also are aneuploid (have abnormal numbers of
chromosomes), and considering the role of centrosomes in chromosome movement, it is
tempting to think that the two phenomena are related.

Mutations in the tumor suppressor gene p53 seem to predispose the cell to excess
replication of the centrosomes.

Chromosome movement in mitosis also involves polymerization and depolymerization of

the microtubules. Taxol, a drug found in the bark of the Pacific yew, prevents
depolymerization of the microtubules of the spindle fiber. This, in turn, stops
chromosome movement, and thus prevents the completion of mitosis. Taxol is being used
with some success as an anticancer drug.

Centrioles
Each centrosome contains a pair of centrioles.

Centrioles are built from a cylindrical array of 9 microtubules, each of which has
attached to it 2 partial microtubules.

The photo (courtesy of E. deHarven) is an electron micrograph showing a cross section of

a centriole with its array of nine triplets of microtubules. The magnification is
approximately 305,000.

When a cell enters the cell cycle, and proceeds from G1 to S phase, each centriole is
duplicated. A "daughter" centriole grows out of the side of each parent centriole. Thus
centriole replication — like DNA replication (which is occurring at the same time) — is
semiconservative.

Once formed, most of the functions of the centrosomes can be accomplished without
centrioles. However,
  Centrioles appear to be needed to organize the centrosome in which they are
embedded.
 Sperm cells contain a pair of centrioles; eggs have none. The sperm's centrioles
are absolutely essential for forming a centrosome which will form a spindle
enabling the first division of the zygote to take place.
 Centrioles are also needed to make cilia and flagella.

Cilia and Flagella

Both cilia and flagella are constructed from microtubules, and both provide either
 locomotion for the cells (e.g., sperm) or
 move fluid past the cells (e.g., ciliated epithelial cells that line our air passages
and move a film of mucus towards the throat).

Both cilia and flagella have the same basic structure. If the cell has
 many short ones, we call them cilia or
 only one or a few long ones, we call them flagella.

Each cilium (or flagellum) is made of

 a cylindrical array of 9 evenly-spaced microtubules, each with a partial
microtubule attached to it. This gives the structure a "figure 8" appearance when
view in cross section.
 2 single microtubules run up through the center of the bundle, completing the so-
called "9+2" pattern.
 The entire assembly is sheathed in a membrane that is simply an extension of the
plasma membrane.

This electron micrograph (courtesy of Peter Satir) shows the 9+2 pattern of microtubules
in a single cilium seen in cross section.

Motion of cilia and flagella is created by the microtubules sliding past one another —
Link. This requires:

 motor molecules of dynein, which link adjacent microtubules together, and

 the energy of ATP.

Each cilium or flagellum grows out from, and remains attached to, a basal body
embedded in the cytoplasm. Basal bodies are identical to centrioles and are, in fact,
produced by them.

Primary Cilia
Motile, "9+2", cilia are found only on certain cells in the vertebrate body, e.g., the
epithelia lining the airways.
But almost every cell in vertebrates has — or had — a single primary cilium. These do
not beat because they lack the central pair of microtubules; that is they are "9+0".

Where functions have been identified, they all involve sensory reception.

Some examples:

Mechanoreceptors
A primary cilium extends from the apical surface of the epithelial cells lining the kidney
tubules and monitors the flow of fluid through the tubules. Inherited defects in the
formation of these cilia cause polycystic kidney disease.

Chemoreceptors
We detect odors by receptors on the primary cilium of olfactory neurons. [Link]

Photoreceptors

The outer segment of the rods in the vertebrate retina is also derived from a primary
cilium. [View]

Welcome&Next Search

Cell Membranes
One universal feature of all cells is an outer limiting membrane
called the plasma membrane.

In addition, all eukaryotic cells contain elaborate systems of

internal membranes which set up various membrane-enclosed
compartments within the cell.

Cell membranes are built from lipids and proteins.

The Plasma Membrane

The plasma membrane serves as the interface between the machinery in the interior of the
cell and the extracellular fluid (ECF) that bathes all cells.

The lipids in the plasma membrane are chiefly phospholipids like phosphatidyl
ethanolamine and cholesterol. Phospholipids are amphiphilic with the hydrocarbon tail of
the molecule being hydrophobic; its polar head hydrophilic. As the plasma membrane
faces watery solutions on both sides, its phospholipids accommodate this by forming a
phospholipid bilayer with the hydrophobic tails facing each other.
 Integral Membrane Proteins
Many of the proteins associated with
the plasma membrane are tightly
bound to it.
 Some are attached to lipids in
the bilayer.
 In others - the
transmembrane proteins -
the polypeptide chain actually
traverses the lipid bilayer. The
figure shows a transmembrane
protein that passes just once through the bilayer and another that passes through it
7 times. All G-protein-coupled receptors (e.g., receptors of peptide hormones, and
odors each span the plasma membrane 7 times.

In all these cases, the portion within the lipid bilayer consists primarily of hydrophobic
amino acids. These are usually arranged in an alpha helix so that the polar -C=O and -NH
groups at the peptide bonds can interact with each other rather than with their
hydrophobic surroundings.

Those portions of the polypeptide that project out from the bilayer tend to have a high
percentage of hydrophilic amino acids. Furthermore, those that project into the aqueous
surroundings of the cell are usually glycoproteins, with many hydrophilic sugar residues
attached to the part of the polypeptide exposed at the surface of the cell.

Some transmembrane proteins that span the bilayer several times form a hydrophilic
channel through which certain ions and molecules can enter (or leave) the cell. [Example]

Peripheral Membrane Proteins

These are more loosely associated with the membrane. They are usually attached
noncovalently to the protruding portions of integral membrane proteins.

Membrane proteins are often restricted in their movements.

A lipid bilayer is really a film of oil. Thus we might expect that structures immersed in it
would be relatively free to float about. For some membrane proteins, this is the case. For
others, however, their mobility is limited:
 Some of the proteins exposed at the interior face of the plasma membrane are
tethered to cytoskeletal elements like actin microfilaments.
 Some proteins are the exterior face of the plasma membrane are anchored to
components of the extracellular matrix like collagen.
 Integral membrane proteins cannot pass through the tight junctions found between
some kinds of cells (e.g., epithelial cells).

Welcome&Next Search
 Index to this page
 The Nuclear Envelope
 Chromatin
 Nucleosomes
 Histone Modifications
The Nucleus  Histone Variants
The nucleus is the hallmark of  Euchromatin versus Heterochromatin
eukaryotic cells; the very term  Nucleosomes and Transcription
eukaryotic means having a "true  The Nucleolus
nucleus".  Nuclear Pore Complexes (NPCs)
 Import into the nucleus
 Export from the nucleus
The Nuclear Envelope
 "Nucleoplasm"
The nucleus is enveloped by a pair of
membranes enclosing a lumen that is continuous with that of the endoplasmic reticulum.
The inner membrane is stabilized by a meshwork of intermediate filament proteins called
lamins.

The nuclear envelope is perforated by thousands of nuclear pore complexes (NPCs) that
control the passage of molecules in and out of the nucleus.

Chromatin
The nucleus contains the chromosomes of
the cell. Each chromosome consists of a
single molecule of DNA complexed with
an equal mass of proteins. Collectively, the
DNA of the nucleus with its associated
proteins is called chromatin.

Most of the protein consists of multiple copies of 5 kinds of histones. These are basic
proteins, bristling with positively charged arginine and lysine residues. (Both Arg and
Lys have a free amino group on their R group, which attracts protons (H+) giving them a
positive charge.) Just the choice of amino acids you would make to bind tightly to the
negatively-charged phosphate groups of DNA.

Chromatin also contains small amounts of a wide variety of nonhistone proteins. Most
of these are transcription factors (e.g., the steroid receptors) and their association with the
DNA is more transient.

Nucleosomes
Two copies of each of four kinds of histones
 H2A
 H2B
 H3 and
 H4
 form a core of protein, the nucleosome core. Around this
is wrapped about 147 base pairs of DNA.

From 20–60 bp of DNA link one nucleosome to the next.

Each linker region is occupied by a single molecule of
histone 1 (H1).

The binding of histones to DNA does not depend on

particular nucleotide sequences in the DNA but does
depend critically on the amino acid sequence of the histone. Histones are some of the
most conserved molecules during the course of evolution. Histone H4 in the calf differs
from H4 in the pea plant at only 2 amino acids residues in the chain of 102.

This electron micrograph (courtesy of David E. Olins and Ada L. Olins) shows chromatin
from the nucleus of a chicken red blood cell (birds, unlike most mammals, retain the
nucleus in their mature red blood cells). The arrows point to the nucleosomes. You can
see why the arrangement of nucleosomes has been likened to "beads on a string".

The formation of nucleosomes helps somewhat, but not nearly enough, to make the DNA
sufficiently compact to fit in the nucleus. In order to fit 46 DNA molecules (in humans),
totaling over 2 meters in length, into a nucleus that may be only 10 µm across requires
more extensive folding and compaction.

 Interactions between the exposed "tails" of the core histones causes nucleosomes
to associate into a compact fiber 30 nm in diameter.
 These fibers are then folded into more complex structures whose precise
configuration is uncertain and which probably changes with the level of activity
of the genes in the region.

Histone Modifications
Although their amino acid sequence (primary structure) is unvarying, individual histone
molecules do vary in structure as a result of chemical modifications that occur later to
individual amino acids.

These include adding:

 acetyl groups (CH3CO-) to lysines
 phosphate groups to serines and threonines
 methyl groups to lysines and arginines

Although 75–80% of the histone molecule is incorporated in the core, the remainder — at
the N-terminal — dangles out from the core as a "tail" (not shown in the figure).

The chemical modifications occur on these tails, especially of H3 and H4. Most of theses
changes are reversible. For example, acetyl groups are
  added by enzymes called histone acetyltransferases (HATs)(not to be confused
with the "HAT" medium used to make monoclonal antibodies!) and
 removed by histone deacetylases (HDACs).

More often than not, acetylation of histone tails occurs in regions of chromatin that
become active in gene transcription. This makes a kind of intuitive sense as adding acetyl
groups neutralizes the positive charges on Lys thus reducing the strength of the
association between the highly-negative DNA and the highly-positive histones.

But there is surely more to the story.

 Acetylation of Lys-16 on H4 prevents the interaction of their "tails" needed to

form the compact 30-nm structure of inactive chromatin. But this case involves
interrupting protein-protein not protein-DNA interactions.
 Methylation, which also neutralizes the charge on lysines (and arginines), can
either stimulate or inhibit gene transcription in that region.
o Methylation of lysine-4 in H3 is associated with active genes while
o methylation of lysine-9 in H3 is associated with inactive genes. (These
include those imprinted genes that have been permanently inactivated
in somatic cells. Link to discussion.)
 And adding phosphates causes the chromosomes to become more — not less —
compact as they get ready for mitosis and meiosis.

In any case, it is now clear that histones are a dynamic component of chromatin and not
simply inert DNA-packing material.

Histone Variants
 We have genes for 8 different varieties of histone 1 (H1). Which variety is found
at a particular linker depends on such factors as
o the type of cell,
o where it is in the cell cycle, and
o its stage of differentiation.

In some cases, at least, a particular variant of H1 associates with certain

transcription factors to bind to the enhancer of specific genes turning off
expression of those genes.

 Some other examples of histone variants:

o H3 is replaced by CENP-A ("centromere protein A") at the nucleosomes
near centromeres. Failure to substitute CENP-A for H3 in this regions
blocks centromere structure and function.
o H2A may be replaced by the variant H2A.Z at the boundaries between
euchromatin and heterochromatin.
o All the "standard" histones are replaced by variants as sperm develop.
In general, the "standard" histones are incorporated into the nucleosomes as new DNA is
synthesized during S phase of the cell cycle. Later, some are replaced by variant histones
as conditions in the cell dictate.

Euchromatin versus Heterochromatin

During interphase, little can be seen of chromatin structure (except for special cases like
the polytene chromosomes of Drosophila and some other flies). However, the density of
the chromatin (that is, how tightly it is packed) varies throughout the nucleus:
 dense regions are called heterochromatin
 less dense regions are called euchromatin.

Heterochromatin
 is found in parts of the chromosome where there are few or no genes, such as
o centromeres and
o telomeres
 is densely-packed;
 is greatly enriched with transposons and other "junk" DNA;
 is replicated late in S phase of the cell cycle;
 has reduced crossing over in meiosis.
 Those genes present in heterochromatin are generally inactive;
that is, not transcribed and show
o increased methylation of the cytosines in CpG islands
of the DNA [Link];
o decreased acetylation of histones and
o increased methylation of lysine-9 in histone H3,
which now provides a binding site for
heterochromatin protein 1 (HP1), which blocks access by the
transcription factors needed for gene transcription.

Euchromatin
 is found in parts of the chromosome that contain many genes;
 is loosely-packed in loops of 30-nm fibers.
 These are separated from adjacent heterochromatin by insulators.
More on insulators
 The loops are often found near the nuclear pore complexes. (This would seem to
make sense making it easier for the gene transcripts to get to the cytosol, but there
is evidence that as gene transcription proceeds, the active DNA actually moves
into the interior of the nucleus.)
 The genes in euchromatin are active and thus show
o decreased methylation of the cytosines in CpG islands of the DNA
[Link];
o increased acetylation of histones and
o decreased methylation of lysine-9 in histone H3.
The diagram represents a hypothetical model of how euchromatin and heterochromatin
may be organized during interphase in a vertebrate cell.

Nucleosomes and Transcription

Transcription factors cannot bind to their promoter if the promoter is blocked by a
nucleosome. For at least some genes, one of the first functions of the assembling
transcription factors is to slide the nucleosome father along the DNA molecule exposing
the gene's promoter so that the transcription factors can then bind to it.

The actual transcription of protein-coding genes is done by RNA polymerase II (RNAP

II). In order for it to travel along the DNA to be transcribed, a complex of proteins
removes the nucleosomes in front of it and then replaces them after RNAP
II has transcribed that portion of DNA and moved on.

The Nucleolus
During the period between cell divisions, when the chromosomes are in
their extended state, 1 or more of them (10 in human cells) have loops
extending into a spherical mass called the nucleolus. Here are synthesized
three (of the four) kinds of RNA molecules (28S, 18S, 5.8S) used in the
assembly of the large and small subunits of ribosomes.

28S, 18S, and 5.8S ribosomal RNA is transcribed (by RNA polymerase I) from hundreds
to thousands of tandemly-arranged rDNA genes distributed (in humans) on 10 different
chromosomes. The rDNA-containing regions of these 10 chromosomes cluster together
in the nucleolus.

(In yeast, the 5S rRNA molecules — as well as transfer RNA molecules — are also
synthesized (by RNA polymerase III) in the nucleolus.)

Once formed, rRNA molecules associate with the dozens of different ribosomal proteins
used in the assembly of the large and small subunits of the ribosome.

But all proteins are synthesized in the cytosol — and all the ribosomes are needed in the
cytosol to do their work — so there must be a mechanism for the transport of these large
structures in and out of the nucleus. This is one of the functions of the nuclear pore
complexes.

Nuclear Pore Complexes (NPCs)

The nuclear envelope is perforated with thousands of pores.

Each is constructed from a number (30 in yeast; probably around 50 in vertebrates)

different proteins called nucleoporins.
The entire assembly forms an aqueous channel connecting the cytosol with the interior of
the nucleus ("nucleoplasm"). When materials are to be transported through the pore, it
opens up to form a channel some 25 nm wide — large enough to get such large
assemblies as ribosomal subunits through.

Transport through the nuclear pore complexes is active; that is, it requires

 energy
 many different carrier molecules each specialized to transport a particular cargo
 docking molecules in the NPC (represented here as colored rods and disks).

Import into the nucleus

All proteins are synthesized in the cytosol and those needed by the nucleus must be
imported into it through the NPCs. Probably each of these proteins has a characteristic
sequence of amino acids — called a nuclear localization sequence (NLS) — that targets
it for entry.

They include:
 all the histones needed to make the nucleosomes
 all the ribosomal proteins needed for the assembly of ribosomes
 all the transcription factors (e.g., the steroid receptors) needed to turn genes on
(and off)
 all the splicing factors needed to process pre-mRNA into mature mRNA
molecules; that is, to cut out intron regions and splice the exon regions.

Export from the nucleus

Molecules and macromolecular assemblies exported from the nucleus include:
 the ribosomal subunits containing both rRNA and proteins
 messenger RNA (mRNA) molecules (accompanied by proteins)
 transfer RNA (tRNA) molecules (also accompanied by proteins)
 transcription factors that are returned to the cytosol to await reuse

Both the RNA and protein molecules contain a characteristic nuclear export sequence
(NES) needed to ensure their association with the right carrier molecules to take them out
to the cytosol.

"Nucleoplasm"
The term "nucleoplasm" is still used to describe the contents of the nucleus. However, the
term disguises the structural complexity and order that seems to exist within the nucleus.
For example, there is evidence that DNA replication and transcription occur at discrete
sites within the nucleus.
Welcome&Next Search

The Golgi Apparatus

The Golgi apparatus is a cell structure mainly devoted to processing the proteins
synthesized in the endoplasmic reticulum (ER).
 Some of these will eventually end up as integral
membrane proteins embedded in the plasma
membrane.
 Other proteins moving through the Golgi will end up
in lysosomes
 or be secreted by exocytosis (e.g., digestive
enzymes).
Link to discussion of the paths taken by proteins when they leave the Golgi.
 The major processing activity is glycosylation: the
adding of sugar molecules to form glycoproteins.
 In some cells, e.g., mucus-secreting cells in
epithelia, the amount of carbohydrate so far exceeds
that of the protein that the product is called a
mucopolysaccharide (also known as a
proteoglycan).
 In plant cells, the Golgi secretes the cell plate and
cell wall.
 Small peptides, e.g., some hormones and
neurotransmitters, are too small to be synthesized
directly by ribosomes. Instead, the ribosomes on the
ER synthesize a large precursor protein that is later
cut up into small peptide fragments as it traverses
the Golgi.

Example: proopiomelanocortin (POMC) — a polypeptide of 265 amino acids

from which is cut

o ACTH
o alpha and beta MSH
o beta-endorphin
o and others.

The Golgi consists of a stack of membrane-bounded cisternae located between the

endoplasmic reticulum and the cell surface.

Many different enzymes (proteins) are present in the Golgi to perform its various
synthetic activities. So there must be mechanisms
 to sort out the processed proteins and send them on to their destinations while
 reclaiming processing proteins (e.g., glycosylases) for reuse.
All the details are far from worked out, but these are some of the features for which there
is considerable experimental evidence.

The Outbound Path

 Transition vesicles pinch off from the surface of the endoplasmic reticulum
carrying
o integral membrane proteins
o soluble proteins awaiting processing
o processing enzymes
 Pinching off requires that the vesicle be coated with COPII
(Coat Protein II)
 The transition vesicles move toward the cis Golgi on
microtubules.
 As they do so, their COPII coat is removed and they may
fuse together forming larger vesicles.
 These fuse with the cis Golgi
 Sugars are added to proteins in small packets so many
glycoproteins have to undergo a large number of sequential
steps of glycosylation, each requiring its own enzymes.
 These steps take place as shuttle vesicles carry the proteins from cis to medial to
the trans Golgi compartments.
 At the outer face of the trans Golgi, vesicles pinch off and carry their completed
products to their various destinations.

The Inbound Path

The movement of cisternal contents through the stack means that essential processing
enzymes are also moving away from their proper site of action.

Using a variety of signals, the Golgi separates the products from the processing enzymes
that made them and returns the enzymes back to the endoplasmic reticulum.

This transport is also done by pinching off vesicles, but the inbound vesicles are coated
with COPI (coat protein I)

How does a vesicle recognize its correct target?

This involves pairs of complementary integral membrane proteins
 v-SNAREs = "vesicle SNAREs" — on the vesicle surface;
 t-SNAREs = "target SNAREs" — on the surface of the target membrane.

v-SNAREs and t-SNAREs bind specifically to each other thanks to the complementary
structure of their surface domains.
Binding is followed by fusion of the two membranes.

Another mechanism of Golgi traffic?

There is evidence (in yeast) that in addition to the pinching off and fusing of shuttle
vesicles (and perhaps more important), the cisternae of the Golgi actually migrate
themselves; that is, the cis Golgi gradually migrates up the stack becoming a medial and
finally a trans Golgi (depicted in the top figure with red arrows).

The Golgi is not a static cell organelle

 The Golgi Index to this page
breaks up
and  Pathways Through the Endoplasmic Reticulum (ER)
disappears  The Signal Sequence
at the onset  Chaperones
of mitosis.  Destinations of proteins synthesized within the ER
 By  The Signal Recognition Particle (SRP)
telophase of  Destinations of Proteins Synthesized By Free Ribosomes
mitosis, the o Nucleus
Golgi o Mitochondria
reappears. o Chloroplasts
How it is
recreated is o Peroxisomes
still
uncertain.

Welcome&Next Search

Protein Kinesis: Getting Proteins to Their Destination

Proteins are the major building blocks of life. Eukaryotic cells synthesize proteins for
thousands of different functions. Some examples:

 to build the components of the cytosol (e.g. microtubules, glycolytic enzymes);

 to build the receptors and other molecules exposed at the surface of the cell
embedded in the plasma membrane;
 to supply some of the components of the mitochondria and (in plant cells)
chloroplasts;
 proteins secreted from the cell to supply the needs of other cells and tissues (e.g.
collagen to support cells, hormones to signal them).

All proteins are synthesized by ribosomes using the information encoded in molecules of
messenger RNA (mRNA). This process is called translation and is described in Gene
Translation: RNA -> Protein. Our task here is to explore the
ways that these proteins are delivered to their proper
destinations.

The various destinations for proteins occur in two major sets:

 one set for those proteins synthesized by ribosomes
that remain suspended in the cytosol, and
 a second set for proteins synthesized by ribosomes
that are attached to the membranes of the
endoplasmic reticulum (ER) forming "rough
endoplasmic reticulum" (RER). This electron
micrograph (courtesy of Keith Porter) shows the RER
in a bat pancreas cell. The clearer areas are the
lumens.

So the first decision that must be made as a ribosome begins to translate a mRNA into a
polypeptide is whether to remain free in the cytosol or to bind to the ER.

Pathways Through the Endoplasmic Reticulum (ER)

The decision to enter the ER is dictated by the presence of a signal sequence on the
growing polypeptide.

The Signal Sequence

The signal sequence consists of the first portion of the elongating polypeptide chain (so
the signal sequence occurs at the amino terminal of the polypeptide). Typical signal
sequences contain 15 - 30 amino acids. The precise amino acid sequence varies
surprisingly from one protein to the next, but all signal sequences include many
hydrophobic amino acids.

The 1999 Nobel Prize in Physiology or Medicine was awarded on October 11, 1999 to Dr.
Günter Blobel for his discovery of the signal sequence and other intrinsic signals that
enable proteins to reach their proper destinations.

If a signal sequence is present,

 translation ceases after it has been synthesized

 the signal sequence is recognized by and is bound by a signal recognition
particle (SRP)
 the complex of ribosome with its nascent polypeptide and the SRP binds to a
receptor on the surface (facing the cytosol) of the ER.
 the SRP leaves and translation recommences
 the growing polypeptide chain is extruded through a pore in the ER membrane
and into the lumen of the ER.
  the signal sequence is usually clipped off the
polypeptide unless the polypeptide is to be retained as
an integral membrane protein.
 other proteins, called molecular chaperones, present
in the lumen of the ER, bind the growing polypeptide
chain and assist it to fold into its correct tertiary
structure.
 sugar residues may be added to the protein. The
process is called glycosylation and often is essential
for proper folding of the final product, a glycoprotein.

Destinations of proteins synthesized within the

ER
Proteins synthesized within the ER are transported to the
Golgi apparatus. Portions of the ER are pinched off,
forming transport vesicles. These carry their load of proteins to the Golgi apparatus. The
membrane of the transport vesicle fuses with the membrane of the Golgi apparatus,
merging their contents. Further steps of glycosylation may occur within the Golgi
apparatus. The exact pattern of glycosylation determines the final destination of the
proteins. There are two options.
 proteins glycosylated with residues of mannose-6-phosphate will leave the Golgi
in transport vesicles that eventually fuse with lysosomes (path 2 in the figure).
 proteins that do not receive this marker, leave in transport vesicles that eventually
fuse with the plasma membrane (path 1 in the figure). These are:
o integral membrane proteins that become exposed at the surface of the cell
(forming receptors and the like) and
o proteins in solution within the transport vesicle. These are discharged from
the cell. This secretory process is called exocytosis.

The Signal Recognition Particle (SRP)

The signal recognition particle in mammalian cells is made from:
 a single small (7S) molecule of RNA
 six different molecules of protein

It contains binding sites for:

 the signal sequence
 the ribosome, and
 an SRP receptor, also called the docking protein, on the cytosol face of the
membranes of the ER.

[Return to signal sequence]

Destinations of Proteins Synthesized By Free Ribosomes

Ribosomes synthesizing a protein without a signal sequence do not bind to the ER and
continue synthesis until the polypeptide is completed. Chaperones are also present in the
cytosol that help the protein assume its final three-dimensional configuration. Some of
the important destinations for these proteins are:
 The cytosol itself. Such proteins as
o the enzymes of glycolysis
o tubulins for making microtubules
o actin for making microfilaments

are simply released from the ribosome and go to work.

 The nucleus. Many proteins - histones, transcription factors, and ribosomal

proteins are notable examples - must move from the cytosol into the interior of the
nucleus. They are targeted to the nucleus by their nuclear localization sequence,
a sequence of 7 - 41 amino acids of which the basic amino acids lysine and
arginine are characteristic members. These proteins are actively transported
through pores in the nuclear envelope into the interior.
 Mitochondria. Although the mitochondrion has its own genome [View] and
protein synthesizing machinery, most of the proteins used by mitochondria are
o encoded by genes in the nucleus of the cell
o synthesized in the cytosol,
o must be imported into the mitochondrion.

Proteins destined for mitochondrion contain a characteristic signal sequence. This

is recognized and bound by a chaperone called mitochondrial stimulation
factor (MSF). MSF targets the protein to a receptor embedded in the outer
membrane of the mitochondrion. Other factors and receptors shepherd proteins
through the intermembrane space to the inner mitochondrial membrane (e.g. some
proteins of the electron transport chain) and the matrix.

External Link
Animation
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 Chloroplasts. Chloroplasts, like mitochondria, have their own genome [View]
and their own protein-synthesizing machinery. But also like mitochondria, most
of the proteins used in chloroplasts are encoded by genes in the nucleus of the
cell, are synthesized by ribosomes in the cytosol, and must then be imported into
the chloroplast. Proteins destined for chloroplasts are recognized by their
characteristic transit sequence. Chaperones are also needed to get them to their
final destination: stroma, thylakoid membrane, etc.
 Peroxisomes. Proteins destined for peroxisomes are synthesized with a
peroxisomal targeting signal (PTS) that binds to a receptor molecule that takes
the protein into the peroxisome and then returns for another load.

Two peroxisomal targeting signals have been identified:

 o a 9-amino acid sequence at the N-terminal of the protein;
o a tripeptide at the C-terminal.

Each has its own receptor to take it to the peroxisome.

Welcome&Next Search

Index to this page

 Mitochondria
 The Citric Acid Cycle
 The Electron Transport Chain
 Chemiosmosis in Mitochondria
 How many ATPs?

 Mitochondrial DNA (mtDNA)

Cellular Respiration
Cellular respiration is the process of oxidizing food molecules, like glucose, to carbon
dioxide and water. The energy released is trapped in the form of ATP for use by all the
energy-consuming activities of the cell.

The process occurs in two phases:

 glycolysis, the breakdown of glucose to pyruvic acid
 the complete oxidation of pyruvic acid to carbon dioxide and water

In eukaryotes, glycolysis occurs in the cytosol. (Link to a discussion of glycolysis). The

remaining processes take place in mitochondria.

Mitochondria
Mitochondria are membrane-enclosed organelles distributed through the cytosol of most
eukaryotic cells. Their number within the cell ranges from a few hundred to, in very
active cells, thousands. Their main function is the conversion of the potential energy of
food molecules into ATP. Mitochondria have:
 an outer membrane that encloses the entire structure
 an inner membrane that encloses a fluid-filled matrix
 between the two is the intermembrane space
 the inner membrane is elaborately folded with shelflike cristae projecting into the
matrix.
 a small number (some 5–10) circular molecules of DNA

The number of mitochondria in a cell can

  increase by their fission (e.g.
following mitosis);
 decrease by their fusing together.

The Outer Membrane

The outer membrane contains many
complexes of integral membrane proteins
that form channels through which a variety
of molecules and ions move in and out of the
mitochondrion.

The Inner Membrane

The inner membrane contains 5 complexes
of integral membrane proteins:
 NADH dehydrogenase (Complex I)
 succinate dehydrogenase (Complex II)
 cytochrome c reductase (Complex III; also known as the cytochrome b-c1
complex)
 cytochrome c oxidase (Complex IV)
 ATP synthase (Complex V)

The Matrix

The matrix contains a complex mixture of soluble enzymes that catalyze the respiration
of pyruvic acid and other small organic molecules.

Here pyruvic acid is

 oxidized by NAD+ producing NADH + H+

 decarboxylated producing a molecule of
o carbon dioxide (CO2) and
o a 2-carbon fragment of acetate bound to coenzyme A forming acetyl-CoA

The Citric Acid Cycle

 This 2-carbon fragment is donated to a molecule of oxaloacetic acid.
 The resulting molecule of citric acid (which gives its name to the process)
undergoes the series of enzymatic steps shown in the diagram.
 The final step regenerates a molecule of oxaloacetic acid and the cycle is ready to
turn again.

Summary:
 Each of the 3 carbon atoms present in the pyruvate that entered the mitochondrion
leaves as a molecule of carbon dioxide (CO2).
  At 4 steps, a pair of electrons (2e-) is removed and transferred to
NAD+ reducing it to NADH + H+.
 At one step, a pair of electrons is removed from succinic acid and
reduces FAD to FADH2.

The electrons of NADH and FADH2 are transferred to the electron

transport chain.

The Electron Transport Chain

The electron transport chain consists of 3 complexes of integral
membrane proteins
 the NADH dehydrogenase complex (I)
 the cytochrome c reductase complex (III)
 the cytochrome c oxidase complex (IV)

and two freely-diffusible molecules

 ubiquinone
 cytochrome c

that shuttle electrons from one complex to the next.

The electron transport chain accomplishes:

 the stepwise transfer of electrons from NADH (and FADH2) to oxygen molecules
to form (with the aid of protons) water molecules (H2O);

(Cytochrome c can only transfer one electron at a time, so cytochrome c oxidase

must wait until it has accumulated 4 of them before it can react with oxygen.)

 harnessing the energy released by this transfer to the pumping of protons (H+)
from the matrix to the intermembrane space.
 Approximately 20 protons are pumped into the intermembrane space as the 4
electrons needed to reduce oxygen to water pass through the respiratory chain.
 The gradient of protons formed across the inner membrane by this process of
active transport forms a miniature battery.
 The protons can flow back down this gradient, reentering the matrix, only through
another complex of integral proteins in the inner membrane, the ATP synthase
complex (as we shall now see).

Chemiosmosis in mitochondria
The energy released as electrons pass down the gradient from NADH to oxygen is
harnessed by three enzyme complexes of the respiratory chain (I, III, and IV) to pump
protons (H+) against their concentration gradient from the matrix of the mitochondrion
into the intermembrane space (an example of active transport).

As their concentration increases there (which is the same as saying that the pH
decreases), a strong diffusion gradient is set up. The only exit for these protons is through
the ATP synthase complex. As in chloroplasts, the energy released as these protons flow
down their gradient is harnessed to the synthesis of ATP. The process is called
chemiosmosis and is an example of facilitated diffusion.

One-half of the 1997 Nobel Prize in Chemistry was awarded to Paul D. Boyer and John
E. Walker for their discovery of how ATP synthase works.

External Link
Animations of the electron transport chain and the workings of ATP synthase
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How many ATPs?

It is tempting to try to view the synthesis of ATP as a simple matter of stoichiometry (the
fixed ratios of reactants to products in a chemical reaction). But (with 3 exceptions) it is
not.

Most of the ATP is generated by the proton gradient that develops across the inner
mitochondrial membrane. The number of protons pumped out as electrons drop from
NADH through the respiratory chain to oxygen is theoretically large enough to generate,
as they return through ATP synthase, 3 ATPs per electron pair (but only 2 ATPs for each
pair donated by FADH2).

With 12 pairs of electrons removed from each glucose molecule,

 10 by NAD+ (so 10x3=30); and

 2 by FADH2 (so 2x2=4),
 this could generate 34 ATPs.

Add to this the 4 ATPs that are generated by the 3 exceptions and
one arrives at 38.

But

 The energy stored in the proton gradient is also used for

the active transport of several molecules and ions through
the inner mitochondrial membrane into the matrix.
 NADH is also used as reducing agent for many cellular
reactions.

So the actual yield of ATP as mitochondria respire varies with conditions. It probably
seldom exceeds 30.

The three exceptions

A stoichiometric production of ATP does occur at:
 one step in the citric acid cycle yielding 2 ATPs for each glucose molecule. This
step is the conversion of alpha-ketoglutaric acid to succinic acid.
 at two steps in glycolysis yielding 2 ATPs for each glucose molecule.

Mitochondrial DNA (mtDNA)

The human mitochondrion contains 5–10 identical, circular molecules of DNA. Each
consists of 16,569 base pairs carrying the information for 37 genes which encode:

 2 different molecules of ribosomal RNA (rRNA)

 22 different molecules of transfer RNA (tRNA) (at least one for each amino
acid)
 13 polypeptides

The rRNA and tRNA molecules are used in the machinery that synthesizes the 13
polypeptides.

The 13 polypeptides participate in building several protein complexes embedded in the

inner mitochondrial membrane.
 7 subunits that make up the mitochondrial NADH dehydrogenase
 3 subunits of cytochrome c oxidase
 2 subunits of ATP synthase
 cytochrome b

Each of these protein complexes also requires subunits that are encoded by nuclear genes,
synthesized in the cytosol, and imported from the cytosol into the mitochondrion. Nuclear
genes also encode ~900 other proteins that must be imported into the mitochondrion.
Mutations in mtDNA cause human diseases.
A number of human diseases are caused by mutations in genes in our mitochondria:
 cytochrome b
 12S rRNA
 ATP synthase
 subunits of NADH dehydrogenase
 several tRNA genes

Although many different organs may be affected, disorders of the brain and muscles are
the most common. Perhaps this reflects the great demand for energy of both these organs.

Some of these disorders are inherited in the germline. In every case, the mutant gene is
received from the mother because none of the mitochondria in sperm survives in the
fertilized egg. Other disorders are somatic; that is, the mutation occurs in the somatic
tissues of the individual.

Example: exercise intolerance

A number of humans who suffer from easily-fatigued muscles turn out to have a
mutations in their cytochrome b gene. Curiously, only the mitochondria in their muscles
have the mutation; the mtDNA of their other tissues is normal. Presumably, very early in
their embryonic development, a mutation occurred in a cytochrome b gene in the
mitochondrion of a cell destined to produce their muscles.

The severity of mitochondrial diseases varies greatly. The reason for this is probably the
extensive mixing of mutant DNA and normal DNA in the mitochondria as they fuse with
one another. A mixture of both is called heteroplasmy. The higher the ratio of mutant to
normal, the greater the severity of the disease. In fact by chance alone, cells can on
occasion end up with all their mitochondria carrying all-mutant genomes — a condition
called homoplasmy (a phenomenon resembling genetic drift).

Why do mitochondria have their own genome?

Many of the features of the mitochondrial genetic system resemble those found in
bacteria. This has strengthened the theory that mitochondria are the evolutionary
descendants of a bacterium that established an endosymbiotic relationship with the
ancestors of eukaryotic cells early in the history of life on earth. However, many of the
genes needed for mitochondrial function have since moved to the nuclear genome.

The recent sequencing of the complete genome of Rickettsia prowazekii has revealed a
number of genes closely related to those found in mitochondria. Perhaps rickettsias are
the closest living descendants of the endosymbionts that became the mitochondria of
eukaryotes.

Further discussion of the evolutionary implications of mtDNA.

Welcome&Next Search

Ribosomes
Ribosomes are the protein-synthesizing machines of the cell.

They translate the information encoded in messenger RNA (mRNA) into a polypeptide.

Link to a description of the process

Ribosomes are
 roughly spherical.
 With a diameter of ~20 nm, they can be seen only with the electron microscope.
 They can make up 25% of the dry weight of cells (e.g., pancreas cells) that
specialize in protein synthesis. (A single pancreas cell can synthesize 5 million
molecules of protein per minute.)

In eukaryotes,
 Ribosomes that synthesize proteins for use within the cytosol (e.g., enzymes of
glycolysis) are suspended in the cytosol.
 Ribosomes that synthesize proteins destined for:
o secretion (by exocytosis)
o the plasma membrane (e.g., cell surface receptors)
o lysosomes

are attached to the cytosolic face of the membranes of the endoplasmic

reticulum. As the polypeptide is synthesized, it is extruded into the interior
(lumen) of the endoplasmic reticulum.

Then, before these proteins reach their final destinations, they undergo a series of
processing steps in the Golgi apparatus.

Link to discussion of Protein Kinesis: the pathways taken by proteins synthesized

on the endoplasmic reticulum.
 Ribosomes that synthesize 13 of the proteins destined for the inner membrane of
mitochondria are found within the mitochondrion itself and are quite different in
structure from the others.

Link to discussion of the evolution of mitochondria.

The ribosomes of bacteria, eukaryotes, and mitochondria differ in many details of their
structure.
 This table gives some of the data. (S values are the
sedimentation coefficient: a measure of the rate at which the particles are spun down in
the ultracentrifuge. S values are not additive. nts = nucleotides.)

Comparison of Ribosome Structure in Bacteria, Eukaryotes, and

Mitochondria
  Bacterial (70S) Eukaryotic (80S) Mitochondrial (55S)
Large Subunit 50S 60S 39S
23S (2904 nts) 28S (4700 nts) 16S (1560 nts)
rRNAs
5S (120 nts) 5S (120 nts)
(1 of each)  
  5.8S (160 nts)
Proteins 33 ~49 48
 
Small Subunit 30S 40S 28S
rRNA 16S (1542 nts) 18S (1900 nts) 12S (950 nts)
Proteins 20 ~33 29

But despite these differences, the basic operations of bacterial, eukaryotic, and
mitochondrial ribosomes are very similar.

Welcome&Next Search Index to this page

 Phagocytosis
 Pinocytosis
 Receptor-Mediated Endocytosis
o The LDL Receptor

 Games Parasites Play

Endocytosis
In endocytosis, the cell engulfs some of its extracellular fluid (ECF)
including material dissolved or suspended in it. A portion of the
plasma membrane is invaginated and pinched off forming a
membrane-bounded vesicle called an endosome.

Phagocytosis
Phagocytosis ("cell eating"):
  results in the ingestion of particulate matter (e.g., bacteria) from the ECF.
 The endosome is so large that it is called a phagosome or vacuole.
 Phagocytosis occurs only in certain specialized cells (e.g., neutrophils,
macrophages, the amoeba), and
 occurs sporadically.

This electron micrograph (courtesy of Dr. Robert J. North) shows a guinea phagocyte
ingesting polystyrene beads. Several beads are already enclosed in phagosomes while the
others are in the process of being engulfed.

In due course, phagosomes deliver their contents to lysosomes. The membranes of the
two organelles fuse. Once inside the lysosome, the contents of the phagosome, e.g.
ingested bacteria, are destroyed by the degradative enzymes of the lysosome.

Games parasites play!

Phagocytic cells, like macrophages and neutrophils, are an early line of defense against
invading bacteria. However, some bacteria have evolved mechanisms to avoid destruction
even after they have been engulfed by phagocytes.

Two examples:

 Salmonella enterica is a bacterium that causes food poisoning in humans. Once

engulfed by phagocytosis, it secretes a protein that prevents the fusion of its
phagosome with a lysosome.
 Mycobacteria (e.g., the tubercle bacillus that causes tuberculosis) use a different
trick.
o When the phagosome is first pinched off from the plasma membrane, it is
coated with a protein called "TACO" (for tryptophan-aspartate-containing
coat protein).
o This must be removed before the phagosome can fuse with a lysosome.
o Mycobacteria taken into a phagosome are able, in some way, to keep the
TACO coat from being removed.

o Thus there is no fusion with lysosomes and the mycobacteria can continue
to live in this protected intracellular location.

Pinocytosis
In pinocytosis ("cell drinking"), the drop engulfed is relatively small.

Pinocytosis
  occurs in almost all cells
 occurs continuously

A cell sipping away at the ECF by pinocytosis acquires a representative sample of the
molecules and ions dissolved in the ECF. But pinocytosis also provides a much more
elegant method for cells to pick up critical components of the ECF that may be in scant
supply.

Receptor-Mediated Endocytosis
Some of the integral membrane proteins that a cell displays at its surface are receptors for
particular components of the ECF. For example, iron is transported in the blood
complexed to a protein called transferrin. Cells have receptors for transferrin on their
surface. When these receptors encounter a molecule of transferrin, they bind tightly to it.
The complex of transferrin and its receptor is then engulfed by endocytosis. Ultimately,
the iron is released into the cytosol. The strong affinity of the transferrin receptor for
transferrin (its ligand) ensures that the cell will get all the iron it needs even if transferrin
represents only a small fraction of the protein molecules present in the ECF. Receptor-
mediated endocytosis is many thousand times more efficient than simple pinocytosis in
enabling the cell to acquire the macromolecules it needs.

Another Example: the Low-Density Lipoprotein (LDL) Receptor

Cells take up cholesterol by receptor-mediated endocytosis. Cholesterol is an essential

component of all cell membranes. Most cells can, as needed, either synthesize cholesterol
or acquire it from the ECF. Human cells get much of their cholesterol from the liver and,
if your diet is not strictly "100% cholesterol-free", by absorption from the intestine.

Cholesterol is a hydrophobic molecule and

quite insoluble in water. Thus it cannot pass
from the liver and/or the intestine to the cells
simply dissolved in blood and ECF. Instead
it is carried in tiny droplets of lipoprotein.
The most abundant cholesterol carriers in
humans are the low-density lipoproteins or
LDLs.

LDL particles are spheres covered with a

single layer of phospholipid molecules with
their hydrophilic heads exposed to the
watery fluid (e.g., blood) and their
hydrophobic tails directed into the interior.
Over a thousand molecules of cholesterol are
bound to the hydrophobic interior of LDL
particles. One molecule of a protein, called apolipoprotein B-100 (Apo B-100) is
exposed at the surface of each LDL particle.

The first step in acquiring LDL particles is for them to bind to LDL receptors exposed at
the cell surface. These transmembrane proteins have a site that recognizes and binds to
the apolipoprotein B-100 on the surface of the LDL. The portion of the plasma membrane
with bound LDL is internalized by endocytosis. A drop in the pH (from ~7 to ~5) causes
the LDL to separate from its receptor. The vesicle then pinches apart into two smaller
vesicles: one containing free LDLs; the other containing now-empty receptors. The
vesicle with the LDLs fuses with a lysosome to form a secondary lysosome. The
enzymes of the lysosome then release free cholesterol into the cytosol. The vesicle with
unoccupied receptors returns to and fuses with the plasma membrane, turning inside out
as it does so (exocytosis). In this way the LDL receptors are returned to the cell surface
for reuse.

People who inherit two defective (mutant) genes for the LDL receptor have receptors that
function poorly or not at all. This creates excessively high levels of LDL in their blood
and predisposes them to atherosclerosis and heart attacks. The ailment is called familial
(because it is inherited) hypercholesterolemia.

Mutations in the Apo B-100 gene cause another form of inherited hypercholesterolemia.

Other small hydrophobic molecules are also transported in the blood while bound to
soluble proteins:
 the retinoid vitamin A (retinol) bound to the retinol-binding protein
 the steroids
o 25[OH] vitamin D3 bound to the vitamin D binding protein
o cortisol bound to the corticosteroid binding globulin
o testosterone and estrogens bound to the sex hormone binding globulin

and there is growing evidence that, like cholesterol, they are taken into the cell by
receptor-mediated endocytosis.

More Games Parasites Play

Some intracellular parasites exploit receptor-mediated endocytosis to sneak their way
into their host cell.

They have evolved surface molecules that serve as decoy ligands for receptors on the
target cell surface. Binding to these receptors tricks the cell into engulfing the parasite.

Some examples:
 Epstein-Barr Virus (EBV). This virus causes mononucleosis and is a
contributing factor in the development of Burkitt's lymphoma, a cancer of B
lymphocytes. It binds to a receptor present on the surface of B cells [Link].
  Influenza virus. The hemagglutinin on the surface of the virus binds to
carbohydrate on the surface of the target cell tricking the cell into engulfing it
[More].
 Listeria monocytogenes. This food-borne bacterium can be dangerous to people
with defective immune systems as well as to pregnant women and their newborn
babies. It has two kinds of surface molecules each a ligand for a different receptor
on the target cell surface.
 Streptococcus pneumoniae. Epithelial cells like those in the nasopharynx have
receptors that are responsible for transporting IgA and IgM antibodies from the
blood to the cell surface. The pneumococcus exploits this receptor for a return trip
into the cell.

This is the organism that led to the discovery that genes are DNA. Link to a
discussion.

Coming full circle.

Endocytosis removes portions of the plasma membrane and takes them inside the cell. To
keep in balance, membrane must be returned to the plasma membrane. This occurs by
exocytosis.

Welcome&Next Search

Cytosol
The fluid in which the organelles of the cytoplasm are suspended. Also called the
ground substance of the cell.

Plant Cells
vs.
Animal Cells
 
Some differences between Plant cells and Animal cells are:
 Plant cells have cell walls as their outermost layer
  Plant cells have chloroplasts that contain chlorophyll
for pigmentation
 Plant cells have larger vacuoles (part of the
endomembrane system used for storage) than Animal
cells
 Animal cells contain centrioles that play a role in
mitosis
 Animal cells have flagellum connected to the cell
membrane which aids in movement of the cell
Cell Walls, chloroplasts, larger vacuoles, centrioles, and
flagellum are all part of the uniqueness that distinguish
Plant cells from Animal cells.
Source: Biology/Neil A. Campbell-4th edition
Copyright 1996