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Intracellular Transport

Three basic modes of transport


1.

Gated transport

2. Transmembrane transport
3. Vesicular transport

A Simplified Roadmap of Intracellular Transport

Vesicular transport

Alberts

Overview of major protein sorting pathways in eukaryotic cells

Protein sorting
cytosol

nuclear
envelope

smooth
ER
lysosomes

nucleus
rough
ER

peroxisomes

mitochondria

Golgi

plasma
membrane

secreted

Mechanism 1: Gated Transport,


proteins enter the nucleus via nuclear pores
The nuclear
envelope is a
double membrane
Continuous with
the ER - both
compartments
share the same
lumen
Perforated by
nuclear pores

The Mechanism of Nuclear Transport

1. Proteins bind to
nuclear transport
receptors
2. Complex is guided to
the pore by filaments
3. Pore opens, receptor +
protein are transported
in (uses GTP)
4. Receptor is shuttled
back into the
cytoplasm

A Simplified Roadmap of Intracellular Transport

Vesicular transport

Alberts

Mechanism 2: Transmembrane Transport,


protein translocation from cytoplasm to
organelle
Proteins moving from the cytosol into
the ER, mitochondria, chloroplasts, or
peroxisomes
Protein movement is mediated by
specialized proteins termed protein
translocators
Unlike passage through nuclear pores,
translocation requires unfolding or cotranslational transport

Proteins are unfolded during translocation


into mitochondria

A Simplified Roadmap of Intracellular Transport

Vesicular transport

Alberts

Mechanism 3: Vesicular Transport


Vesicular transport delivers components between compartments in
the biosynthetic-secretory and endocytic pathways.

Vesicular transport
Vesicular transport delivers components between compartments in the biosyntheticsecretory and endocytic pathways.

Two Key Steps:


1. Sorting during vesicle formation
2. Targeting during vesicle fusion

Plasma membrane

recycling
endosome

late
endosome

secretory
vesicles

lysosome

Retrograde traffic

TGN
trans-Golgi

medial-Golgi
cis-Golgi

exocytosis (secretion)

endocytosis

sorting endosome

EGTC
endoplasmic reticulum
Nucleus

14

Cellular Membranes
Endomembrane Structure

% total membrane in % total in Pancreatic


Hepatocyte
Exocrine Cell

Endoplasmic Reticulum
(smooth)

16

<1

Endoplasmic Reticulum
(rough)

35

60

Golgi

10

Plasma membrane

Lysosomes

0.4

N.D.

Peroxisomes

0.4

N.D.

Endosomes

0.4

N.D.

Mitochondria

39

21

Nuclear envelope (inner


only)

0.2

0.7

What is the endomembrane system?


System of membrane-bound organelles in cells that
work cooperatively together to create secretory
proteins, membrane-bound proteins, or plasma
membrane proteins
The set of membranes that form a single functional and
developmental unit, either being connected directly, or
exchanging material through vesicle transport.

The endomembrane system regulates protein traffic


and performs metabolic functions in the cell

Also involved in assembly and transportation of lipids

Properties of endomembrane system


Present only in eukaryotic cells
Transport system for moving molecules through
interior of the cell
Made of lipid bi-layer with proteins attached to
either side or transversing them
Divides cell into organelles

The endomembrane system plays a key role in the


synthesis (and hydrolysis) of macromolecules in the
cell.
The various
components
modify
macromolecules
for their various
functions.
Fig. 7.16
Copyright 2002 Pearson Education, Inc., publishing as Benjamin Cummings

Endomembrane System
The endomembrane system consists of:

Endoplasmic reticulum(ER)

Golgi complex
Lysosome

secretory vesicles

The Endomembrane System

1. The endoplasmic reticulum manufactures membranes and performs many other


biosynthetic functions
2. The Golgi apparatus finishes, sorts, and ships cell products
3. Lysosomes are digestive compartments
4. Vesicles encloses and transport substances synthesized in the cell.

Vacuoles have diverse functions in cell


maintenance
Vacuoles are membrane-bound sacs with varied
functions.
Food vacuoles, from phagocytosis, fuse with
lysosomes.
Contractile vacuoles, found in freshwater protists,
pump excess water out of the cell.
Central vacuoles are found in many mature plant
cells.

LM

Vacuole filling with water

LM

Vacuole contracting

Central vacuole

Colorized TEM

(a) Contractile vacuole in Paramecium

(b) Central
vacuole in a plant cell
Laura Coronado Bio 10 Chapter 4

Figure 4.17

Endoplasmic Reticulum

ENDOPLASMIC
RETICULUM (ER):
An extensive
tubovesicular network
where proteins and
lipids are made.
Rough ER: studded with
ribosomes, site of protein
biosynthesis
Smooth ER: site of lipid
biosynthesis

Endoplasmic Reticulum
There are two distinct regions of ER
Smooth ER, which lacks
ribosomes
Rough ER, which contains
ribosomes
Rough ER

Smooth ER

Functions of ER
Smooth

The smooth ER
Synthesizes lipids
Metabolizes
carbohydrates
Stores calcium (In
muscle cells, these
trigger contractions)
Detoxifies poison

Rough

The rough ER
Has bound ribosomes
Produces proteins and
membranes, which are
distributed by
transport vesicles

The synthesis of phosphatidylcholine in the ER membrane

Rough ER Functions
Protein and Membrane synthesis
Ribosomes covering Rough ER secrete
proteins
Folded into lumen
Later transported by vesicles

Ex. Insulin

Membranes made for itself are later


transported to other endomembrane
systems

Synthesis of secretory proteins - review


1. N-terminal signal sequence is
synthesized
2. Signal bound by SRP, complex
docks with SRP receptor on ER
membrane
3. Signal sequence binds to
translocon, internal channel
opens, inserted into translocon
4. Polypeptide elongates, signal sequence cleaved
5. ER chaperones prevent faulty folding, carbohydrates added to specific
residues
6. Ribosomes released, recycle
7. C-terminus of protein drawn into ER lumen, translocon gate shuts, protein
assumes final conformation

Synthesis of secretory proteins

Synthesis of integral membrane protein


1. internal signal sequence
bound by SRP

2.

SRP-protein-ribosome

complex docks with SRP


receptor, C-terminal portion
of protein cotranslationally
inserted into lumen of ER
3. Mature protein transverses ER bilayer forming integral membrane protein
NOTE: Orientation of protein within membrane dependent upon cluster of
charged residues adjacent to internal signal sequence
4. Polypeptide elongates, carbohydrates added to specific residues
5. Ribosomes released, recycle, integral membrane protein produced that
forms transmembrane domain

Synthesis of membrane proteins

PROTEIN GLYCOSYLATION IN THE ROUGH ER: During


translation, a signal sequence on membrane and secretory
proteins directs the nascent protein into the ER lumen. After the
protein has entered the ER, the glycosylation process begins.

A PRE-FORMED
PRECURSOR
OLIGOSACCHARIDE
IS TRANSFERRED
EN BLOC
TO PROTEINS
IN THE ER

Alberts

PROTEIN GLYCOSYLATION IN THE ROUGH ER

Alberts

SOME PERIPHERAL MEMBRANE PROTEINS


AQUIRE A COVALENTLY ATTACHED
GLYCOPHOSPHATIDYLINOSITOL (GPI)
ANCHOR IN THE ER

Alberts

Transport from the ER through the


Golgi apparatus

Golgi Apparatus
Made of cisternae
Cis and Trans faces
Cis serves as bridge w/ER
Trans makes vesicles for transport to other cell regions
Modifies proteins
Cisternae between cis and trans faces
Works in partnership with the ER
Receives, refines, stores, and distributes
chemical products of the cell

Function

1. First modification of lipids and proteins


(Modifies the N-linked oligosaccharides and adds O-linked
oligosaccharides)

2. Storage and packaging of materials that


will be exported from the cell.

12/28/2014

41

OLIGOSACCHARIDE CHAINS ARE PROCESSED


IN THE GOLGI APPARATUS

Production of complex oligosaccharides

Modification of the N-linked oligosaccharides is done by


enzymes in the lumen of various Golgi compartments.

Two possible models


explaining the organization
of the Golgi apparatus and
the transport of proteins from
one cisterna to the next

Transport from the trans Golgi


nextwork to Lysosomes

The Lysosomes

The structure of the lysosome:


Discovered in 1950 by Rene . De .Duve, a Lysosome is a tiny
membrane-bound organelle found in the cytoplasm of all
eukaryotic cells containing various acid hydrolytic enzymes
that can digest every kind of biological molecule.

The structure of the lysosome


Lysosomes are highly heterogeneous
Shape and size
But all have acid hydrolases
The stomach of a cell

Lysosomes is common in
animal cells but rare in plant.
Marker enzyme: acid
phosphatase.

The structure of the lysosome


Lysosome membrane:
1.H+-pumps:
internal proton is kept high H+concentration by H+-ATPase
2.Glycosylated proteins:
may protect the lysosome from
self-digestion.

3.Transport proteins:
transporting digested materials.

Biogenesis of Lysosomes

1. A phosphate attached to the mannose residue.


2.This mannose-6 phosphate forms a sorting signal that moves through the
cisternae to the trans region where it binds to a specific receptor.
3.After it binds to the receptor, it begins to bud and a coat made of clathrin
forms around the bud (to strengthen it).
4.It moves away to fuse with a late endosome .
5.The phosphate is removed and hydrolase is dissociated from the receptor.
6.The receptor is then recycled back to the Golgi complex .

Biogenesis of Lysosomes
Late endosome

Lysosomal hydrolase precursor


Addition of phosphate
Mannose-6-phosphate(M-6-P)

Receptor-dependent
transport

ATP

ADP+Pi

From
RER
Clathrin coat
Binding to
M6P receptor

Dissociation at
acidic pH

RER

Cis golgi
network
Golgi apparatus

H+

H=5
P
Removal of
phosphate

Mature
lysosomal
hydrolase

Trans golgi
network

M6P receptor in
budding vesicle

Mature lysosomes

The types of lysosomes


Primary lysosome are newly formed by
budding from the Golgi complex,and
therefore have not yet encountered
Primary Lys.
substrate for digestion and with acid
Hydrolytic enzymes inactive.
Secondary lysosomes result from the
repeated fusion of primary lysosomes
with a variety of membrane bounded
substrates and active hydrolytic enzymes
within the lysosomes. The bounded
substrates may be food bacteriumor
worn organalles and so on.
Second Lys Phagosome is a kind of secondary
lysosomes licked up food or bacterium.
Autophagosome is a kind of secondary
lysosomes licked up ageing organelles.
The secondary lysosomes digest the contents of phagocytic or autophagic vesicles to
form residual bodies that either undergo exocytosis or are retained in the cell as
lipofuscin granules.

The functions of Lysosomes


All major classes of macromolecules are degraded
in lysosomes

Lysosomes are involved in four major


D. The Functions of Lysosomes
cell functions:
1.Heterophagy;
2.Autophagy;
3.The extracellular digest;
4. Autocytolysis;

The functions of Lysosomes


1. Heterophagy
Digestion of materical of
extracellular origin.
Lysosomes pick up foreign invaders
such as bacteria, food and break them
into small pieces that can hopefully be
used again. If they pick up a really
harmful invader, they will eat it up
and expel what is left of it out of the
cell so that the debris can be removed
from the body.
2.Autophagy
Digestion of materical of intracellular
origin.
Lysosomes also play a key role in
destroying old organelles within the
cell and thus allow them to be
replaced with fresher, more effective
ones.This process is known as
autophagy and is accomplished in two
stages.

The functions of Lysosomes


Autophagy
Firstly, a membrane is
donated by the
endoplasmic reticulum.
This membrane then
surrounds the old organelle.
Secondly ,a lysosome fuses
with this membrane to
form an autophagic vacuole.
The lysosome can safely
enter it's enzyme contents
into this vacuole and
destroy the old organelle.
The electron micrograph
shows a lysosome in the
process of destroying a
membrane bound
mitochondria.

The functions of Lysosomes


3.The extracellular
digest :Another function of
lysosome in the human
occurs during fertilization
of the egg by the sperm.
The head of the sperm cell
contains a package of
lysosomal material called
the acrosome.
The enzymes from this are
released when the sperm
makes contact with an egg
and they effectively bore a
hole through the cell
membrane of the egg
allowing the sperm to enter.

The functions of Lysosomes

4.Autocytolysis :Lysosomes may also be important in


development. For instance , they are responsible for the
breakdown of a tadpoles tail as the tadpole develops into a
frog. In the process, the lysosome releases hydrolases to
cytoplasm to digest the cell of oneself.

Transport into the cell from the


plasma membrane endocytosis

Endocytosis
Endocytosis: process of taking in liquids or larger
molecules into a cell by engulfing in a vesicle;
requires energy

Phagocytosis by a macrophage

Phagocytosis by a neutrophil
pseudopods

One macrophage and two red blood cells


Pinocytosis: cell drinking; Phagocytosis: cell eating
Phagocytosis: large particle, >250nm
Pinocytosis: fluid, liquid, 100 nm

ENDOCYTOSIS IS IMPORTANT FOR


CELLS TO:
1. Import selected extracelluar molecules (i.e.
receptor-mediated endocytosis).

2. Regulate levels of membrane proteins on the


cell surface (i.e. receptor down-regulation).

Transport from the trans Golgi network to the


cell exterior: exocytosis

Exocytosis: fusion of vesicles


with the plasma membrane

Exocytosis
Exocytosis: process of releasing substances out of
a cell by fusion of a vesicle with the membrane

Exocytosis of
secretory vesicles

Two mechanisms of secretion

Three pathways of protein sorting in the trans Golgi network