Extra credit for Midterm 2
Send all textbook corrections (any chapter, any
type of correction) by Monday April 9th. The
more corrections, the more points the class will
earn.
Fundamentals of Cell Biology
Chapter 9: The Endomembrane System and
Membrane Trafficking
Chapter Summary: The Big Picture (1)
• Chapter foci:
– Define the endomembrane system and introduce
the general concepts of vesicle-mediated traffic
– Understand the temporal sequence of events
beginning with ER and ending at PM
– Explore details of Golgi function
– Understand Golgi to PM pathway
– The role of the endosome in sorting
– The significance of the lysosome
Chapter Summary: The Big Picture (2)
• Section topics:
– The endomembrane system is a network of
organelles in eukaryotic cells
– Exocytosis begins in the endoplasmic reticulum
– The Golgi apparatus modifies and sorts proteins in
the exocytic pathway
– Exocytosis ends at the plasma membrane
– Endocytosis begins at the plasma membrane
– The endosome sorts proteins in the endocytic
pathway
– Endocytosis ends at the lysosome
The endomembrane system is a network
of organelles in eukaryotic cells
• Key Concepts (1):
– The endomembrane system is a set of
interconnected organelles that readily exchange
materials.
– The primary functions of the endomembrane
system are to control the export (exocytosis) and
import (endocytosis) of materials to/from the
extracellular space.
The endomembrane system is a network of
organelles in eukaryotic cells
• Key Concepts (2):
– Membrane-bound compartments called vesicles
shuttle between organelles in the endomembrane
system and are responsible for carrying material
from one organelle to another.
– The creation, transport, targeting, and fusion of
vesicles occurs in nine steps.
The endomembrane system controls
molecular transport in/out of cell
• Endocytic and exocytic
pathways
• Endomembrane system
Figure 09.01: The endocytic and
exocytic pathways.
Vesicles shuttle material between organelles in the
endomembrane system
• Donor/acceptor
compartments
• Vesicle-mediated
transport
• Anterograde/retro
grade transport =
vesicle shuttling
• Exocytosis,
endocytosis, and
fusion of vesicles
Figure 09.02: In vesicle-mediated
transport, a membrane-bound
vesicle buds from one compartment
and fuses with another.
Vesicle-mediated transport - 9 steps
1)
2)
3)
4)
5)
6)
7)
8)
9)
Cargo selection
Budding
Scission
Uncoating
Transport
Tethering
Docking
Fusion
Disassembly
Figure 09.03: Vesiclemediated transport
occurs in nine steps.
Vesicle-mediated transport - 9 steps
1)
2)
3)
4)
5)
6)
7)
8)
9)
Cargo selection
Budding
Scission
Uncoating
Transport
Tethering
Docking
Fusion
Disassembly
Figure 09.04: The three main
types of coats involved in
vesicle-mediated transport
are COPI, COPII, and clathrin
coats.
Mneumonic Contest
Best mneumonic for the 9 steps wins 2 points on
their choice of midterm exams (1, 2 or 3)
C B S U T T D FD D
Exocytosis begins in the ER
• Key Concepts:
– Newly synthesized endomembrane proteins are
modified in the ER.
– Signaling sequences in the newly-made proteins
signal their secondary destinations. Some have
retention signals that keep them in the ER.
– Proteins that leave the ER enter the cis-Golgi
network via COPII-coated vesicles.
– Retrograde vesicle transport returns ER-resident
proteins from the Golgi apparatus to the ER.
Job of exocytic pathway – sort/distribute
new proteins in endomembrane system
• Newly synthesized proteins begin post-translational
modification as ER resident proteins help them fold
properly
• COPII-coated vesicles shuttle proteins from the ER
to the Golgi apparatus
• Resident ER proteins are retrieved from the Golgi
apparatus
Exocytic pathway
Figure 09.05: Cargo proteins are
incorporated into nascent COPII
vesicles by different mechanisms.
Figure 09.06: ER-resident proteins
are occasionally carried to the CGN
and must be retrieved.
The Golgi apparatus modifies and sorts
proteins in the exocytic pathway
• Key Concepts (1):
– The Golgi apparatus is organized into discrete
compartments called cisternae. The cisternae are
stacked on top of one another, and are classified
as cis, medial, or trans according to their relative
location within the overall Golgi structure.
– Golgi-resident proteins are primarily responsible
for modifying proteins undergoing exocytosis.
They are retained in the Golgi apparatus by
transmembrane Golgi retention sequences.
The Golgi apparatus modifies and sorts
proteins in the exocytic pathway
• Key Concepts (2):
– The extreme ends of the Golgi apparatus are
elaborated into long, tubular structures called the
cis Golgi network and trans Golgi network.
– Both Golgi networks sort proteins into vesicles
targeted to different locations. The trans Golgi
network is especially effective at sorting a large
number of proteins into many distinct vesicle
types.
The Golgi apparatus is subdivided into cis, medial,
and trans cisternae
• Site for post-translational
modification (PTM)
• Golgi proteins retained
by retention sequence
Figure 09.08: Proteins leaving the TGN can
travel to at least five different destination
compartments.
The trans Golgi network (TGN) sorts
proteins exiting the Golgi apparatus
Figure 09.09: Dynamin uses GTP to regulate scission of a vesicle from a donor compartment.
TGN sorting mechanisms are complex
• Many different “cargo selection” mechanisms
– PTM
– Protein aggregation
– Signal receptor
– Lipid raft
Budding of vesicles at the TGN likely
occurs by several different mechanisms
• 3 different mechanisms have been proposed
– Curvature-inducing proteins
– Modification of membrane phospholipids
– Phospholipid asymmetry
Scission of budding vesicles requires
microtubule motor-based forces
• Dynamin - GTPase
Figure 09.10: Examples of the golgi cisternae, rough ER (RER), and various stages of
secretory granule formation. Ly = lysosome.
Exocytosis ends at the plasma membrane
• Key Concepts:
– Cells regulate the last stage of exocytosis
(fusion) for most exocytic vesicles, to control
when and how much material is released into the
extracellular space and to control the delivery of
membrane-associated proteins to the plasma
membrane.
– Controlled secretion is also called regulated
secretion, and is under the control of signaling
pathways.
2 mechanisms for controlling the final
steps of exocytosis
• Constitutive secretion
– constant
• Regulated secretion
– Controlled by
signaling proteins
– Secretory vesicles
(zymogen granules)
– Condensing vacuole
Figure 09.11: Transmission electron
micrograph of clathrin-coated pits and
vesicles at the oocyte surface.
Exocytosis ends at the plasma membrane
• Key Concepts:
– Cells regulate the last stage of exocytosis
(fusion) for most exocytic vesicles, to control
when and how much material is released into the
extracellular space and to control the delivery of
membrane-associated proteins to the plasma
membrane.
– Controlled secretion is also called regulated
secretion, and is under the control of signaling
pathways.
Endocytosis begins at the plasma
membrane
• Key Concepts:
– The onset of endocytosis is most often indicated
by the clustering of cargo receptors on the
plasma membrane, accompanied by the
assembly of a clathrin coat on the cytosolic face
of the cluster. In micrographs, this structure
resembles a pit in the membrane, so it is often
called a coated pit.
– Coated pits complete the nine steps of vesicle
transport and deliver the vesicle to an organelle
called the endosome.
Clathrin stabilizes the formation of
endocytic vesicles
Figure 09.12: The role of clathrin in
endocytosis.
Figure 09.13: Clathrin-coated vesicles are
uncoated and tethered to early endosomes
via EEA1, which binds to
phosphatidylinositol-3-phosphate and Rab5.
The endosome sorts proteins in the
endocytic pathway
• Key Concepts (1):
– The endosome is formed by the fusion of
endocytic vesicles with specific vesicles that bud
from the TGN.
– The endosome sorts materials arriving from the
plasma membrane; cargo receptors are recycled
to the plasma membrane, while cargo remains in
the endosome.
The endosome sorts proteins in the
endocytic pathway
• Key Concepts (2):
– The lumen of the endosome is slightly acidic
relative to the extracellular space, and this acidity
is key to the sorting mechanism.
– This sorting mechanism is very different from the
sorting mechanisms used in the Golgi apparatus.
The endosome is subdivided into early
and late compartments
• Specific interacting
molecules ensure correct
vesicles fuse with
vesicles from TGN or
early endosomes
– EEA-1,Syntaxin,Rab 5,
PIP3
• Proton pump proteins
play central role in
sorting/activation of
endosomal contents
Figure 09.14: Maturation of endosomes to form
lysosomes. Sorting of cell surface receptors and M6P
receptors requires acidic environment provided by
proton pumps.
Endocytosis ends at the lysosome
• Key Concepts (1):
– Complete degradation of endocytosed materials
takes place in an organelle called the lysosome.
– The lysosome is likely generated from the
endosome in several ways, and requires fusing a
vesicle from the TGN that contains essential
proton pump proteins and digestive enzymes with
the endosome.
Endocytosis ends at the lysosome
• Key Concepts (2):
– The digestive enzymes and proton pumps are
carried to the endosome by receptors that
recognize a mannose 6-phosphate tag on
proteins in the TGN. The mannose 6-phosphate
receptors are sorted and returned to the TGN,
while the hydrolases and pumps remain in the
endosome. The pumps are responsible for
triggering the drop in pH in the endosomes.
Endocytosis ends at the lysosome
• Key Concepts (3):
– Once the plasma membrane cargo receptors and
mannose 6-phosphate receptors have been
removed from the endosome, it becomes
progressively more acidic; when it reaches pH 5,
shape changes in the hydrolases are triggered,
which activate the endosomes and allow them to
begin digesting cargo.
– Once digested, the cargo building blocks (sugars,
nucleosides, amino acids, etc.) are transported
into the cytosol for reuse.
Maturation from endocytic vesicle to
lysosome
Figure 09.15: Assembly of the mannose 6-phoshate sorting signal.
Endogenous proteins destined for
lysosome are tagged and sorted by
Golgi
Figure 09.16: Digestion of cargo
molecules in the lysosome.
pH is used in 3 ways to control endocytosis
1) the acidic environment in endosomes helps sort
cargo from receptors
2) the relatively neutral pH of the ER and Golgi
apparatus keeps the hydrolytic enzymes from
digesting these organelles
3) the enzymes’ requirement for a strong acid
environment protects the endomembrane system
from digesting itself
Digestion in the lysosome
•
•
•
•
Proteases
Glycosidases
DNaseII
Phosphatases
Figure 09.17: Ubiquitin-mediated degradation occurs in
multivesicular bodies, which are late endosomal
compartments of the endocytic pathway.
Lysosomes can also degrade some
resident organelles
• “Autophagy” by
autophagosome