Cell Biology Chapter 9: The Endomembrane System and Membrane Trafficking Flashcards
The Endomembrane System Controls Molecular Transport In/Out Of Cell
True
vesicle-mediated transport
Existing membrane pinches off to form a vesicle
Vesicle-mediated Transport - 9 Steps
- Cargo selection
- Budding
- Scission
- Uncoating
- Transport
- Tethering
- Docking
- Fusion
- Disassembly
Exocytosis
Is a one way journey from ER —> Golgi —> to the plasma membrane
Cargo Selection
Sorting mechanisms are used to ensure only the right molecules are transported
Cytosolic tails of membrane proteins
Anterograde signal can consist of two leucines (-LL-)
Coat Proteins (Clathrin, COPI and COPII are examples) Cause receptors and other proteins with signal sequences to cluster into a patch on the surface of the donor compartment
Coat proteins
cause the receptors to cluster in a patch the surface of the donor compartment.
The 3 common Coat proteins are
Clathrin, COPI and COPII
Budding
Clustered coat proteins interact with cytosolic adaptor proteins
Together the coat and the adaptor proteins form a mesh-like lining. Makes the formation of vesicles energetically favorable
*Membranes would never pinch off vesicles by themselves
Scission
The budded vesicle must be cut from the membrane it originated from
Scission proteins surround the “neck” and tighten it until the membrane breaks
Uncoating
Coat proteins (like COPI and COPII) are no longer needed once the vesicle is free. They are recycled
Transport
Motor proteins that attach to microtubules or actin filaments (see Chapter 5) transport vesicles with cargo
Not absolutely required, but increases the efficiency of delivery
Tethering
Vesicles have small GTP-binding proteins called Rab proteins on them
Rabs bind tether proteins and this forms a temporary complex on the acceptor membrane bringing the vesicle closer to it
Like a search and rescue mission….Is it our vesicle?
Docking
After the “Rab” protein helps the vesicle find their targets, they have to dock.
t-SNARES (on target membrane)
v-SNARES (on vesicle membrane)
Are complementary and bind one another, stabilizing the temporary connection made between Rabs and tether proteins
Fusion
Several v-SNARE and t-SNARE interactions will lead to the fusion of the vesicle membrane with the acceptor compartment
Disassembly
This triggers uncoupling of the SNARE pairs by NSF and SNAP proteins
(they can be used again)
V-SNARES will be recycled back to the donor compartment in vesicles moving in the opposite direction
Exocytosis Begins In The ER
True
Exocytosis
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.
Peptidase
Signal Peptidase cleaves the amino-terminal signal sequence from most proteins that have one
N-oligosaccharide transferase
adds a core oligosaccharide to the side chain of Asn residues in an N-linked glycosylation signal
Patients with I-cell disease fail to form proper lysosomes, and thus accumulate inclusion bodies in their cells. In vitro (cell culture), cells from some I-cell patients can be “rescued” (restored to normal phenotype) by adding normal lysosomal enzymes to the culture medium. However, this treatment does not eliminate inclusions in cells from other I-cell patients. Which statement best explains this discrepancy?
) Cells that respond to the added enzymes have functional endosome proton pumps but faulty lysosomal enzymes; cells that do not respond have functional endosome proton pumps but faulty mannose-6-phosphate receptors.
What would happen if a cell had mutated COPI proteins
ER proteins that were accidentally carried to the Golgi complex would not return to the ER.
Choose the proper temporal sequence of events taking place for formation of a lysosome (first–> last).
Budding of vesicles from the trans Golgi network, dissociation of mannose-6-phosphate receptors from their ligands, activation of acidic hydrolases.
Recently, a t-SNARE was found in the outer membrane of mitochondria of a cell. What are the implications of this finding?
This suggests that vesicles budding from the Golgi fuse with mitochondria.
Which of the following is NOT a type of vesicle coat protein? rab, clathrin, COPI, COPII
rab
COPII-Coated Vesicles
Cells harbor either ER export sites or transitional ER
ER export signals are recognized by coat proteins called COPII
COPII proteins interact with adaptor proteins = budding
Scission mechanism is not well understood
Which is NOT a characteristic of receptor-mediated endocytosis?
A) Sorting B) Selectivity C) Ability to concentrate ligand D) Dynamin E) Ligand entry into the cell via a membrane pore
Ligand entry into the cell via a membrane pore
Uncoating
Sec23p in the COPII complex hydrolyzes GTP causing disassembly of the COPII coat
Transport
Sec23p binds dynactin, which can couple vesicles to dynein or kinesin
Tethering
Occurs when Sec23p interacts with at least two tethering proteins on cis-Golgi causing them to bind a Rab
For docking
The Bet1p v-SNARE
Disassembly
mediated by NSF and SNAP
Resident ER Proteins
Not all proteins that arrive at the CGN (cis Golgi network) are supposed to be there
Golgi-to-ER retrograde transport retrieves them
KDEL ER-retention signal
K-K-X-X or K-X-K dilysine ER retention signaL
THE Golgi apparatus is divided into
The Cis, Medial and Trans Cisternae
Vesicle trafficking within the Golgi occurs in both antergrade and retrograde directions
Each cisternae has a unique set of enzymes
Site for post-translational modification (PTM) like glycosylation
Which of the following are examples of posttranslational processing?
A) glycosylation B) protein folding C) 5' CAP D) signal sequence removed E) all but choice c
E) all but choice c
The Transmembrane Region of Resident Golgi Proteins Serve as a Retention Signal
Golgi retention sequences
For Transmembrane proteins this includes the signal anchor sequence that initially targets it to the ER
Two models proposed to explain how these sequences work and neither require a receptor!
Kin recognition model
Bilayer thickness model
Which of the following could be an ER retention signal?
A) arg-Arg (R-R) B) lys-asp-glu-leu (KDEL) C) lys-x-lys (KXK) D) answers A and B E) answers A, B and C
E) answers A, B and C
ER retention signals are _______
- Lys-Asp-Glu-Leu (KDEL)
- Lys-Lys-X-X or Lys-X-Lys (KKXX/KXK)
- Arg-Arg or Arg-X-Arg (RR/RXR)
Resident ER Proteins in yeast
In Yeast, a protein that functions as a t-SNARE during ER-Golgi transport also functions as a v-SNARE for retrograde transport
These receptors interact with COPI (not COPII) coat complex
Vesicles are transported back by kinesin on MTs
The Golgi Apparatus Is Subdivided Into _______
Cis, Medial, And Trans Cisternae.
Each cisternae has a unique set of enzymes
Site for post-translational modification (PTM) like glycosylation
What serves in Resident Golgi Proteins Serve as a Retention Signal
The Transmembrane Region.
Golgi retention sequences
For Transmembrane proteins this includes the signal anchor sequence that initially targets it to the ER
Golgi retention sequences models
Kin recognition model
Bilayer thickness model
Kin recognition model
Protein recognize each other via their membrane spanning domains, cluster to prevent forward movement.
Trans Golgi Network (TGN)
Sorts Proteins Exiting The Golgi Apparatus
Bilayer thickness model
Proteins move along the pathway (cis—> medial—-> trans) until membrane spanning domain matches the thickness of the bilayer.
Sorting Mechanisms at the TGN
Post-translational modifications can act as sorting signals
N- and O-linked glycosylation, phosphorylation and ubiquitination
Protein aggregation
Traditional signal-receptor method
Lipid raft hypothesis
TGN may assemble patches of cholesterol and lipid that some proteins have a very high affinity for
Budding Of Vesicles At The TGN
3 different mechanisms have been proposed
Curvature-inducing proteins
Modification of membrane phospholipids by Phospholipase A2
Phospholipid asymmetry that could arise from flippases or lipid transfer proteins
Scission of Vesicles Budding from the TGN
Often Requires Microtubule Motor-Based Forces that Provide Traction for Curvature
- Budding is an energy dependent process. Dynamin uses GTP to regulate scission of a vesicle from a donor compartment
2 Mechanisms For Controlling The Final Steps Of Exocytosis
Constitutive secretion
- constant
Regulated secretion
- Controlled by signaling proteins
- Secretory vesicles (zymogen granules)
- Condensing vacuole
Endocytosis Begins _______
at the Plasma Membrane.
Endocytosis removes material from the extracellular space
Can destroy dangerous objects (like viruses, foreign cells)
A method to remodel the cell surface
Clathrin
Stabilizes The Formation Of Endocytic Vesicles
Endocytic sorting
requires multiple compartments unlike exocytosis
Separation and sorting occurs in early and late endosomes
The Endosome Is Subdivided
Into Early And Late Compartments
during endocytosis Specific interacting molecules ensure correct vesicles fuse with vesicles from TGN or early endosomes
EEA-1,Syntaxin,Rab 5, PIP3
pH during endocytosis
pH of 6.4-6.8 allows cargo to dissociate from receptors (maintained by proton pumps)
Proton Pumps
Transmembrane proton pumps move protons to the interior of the vesicle interior
Pumps accumulate in the membrane over time as vesicles continue to fuse and the vesicle becomes increasingly more acidic
When pH drops below 6.0 the early endosome becomes a late endosome
Fig.9.14
Endocytosis Ends At _______
The Lysosome
Mamo-6-phosphate
lysosome retrieval sequence
TGN
At the TGN a mannose-6-phosphate “tag” is recognized as a lysosomal sorting signal
At the lysosome, the mannose 6-phosphate receptors are then sorted and returned to the TGN, while the hydrolases and pumps remain there
The pumps are responsible for triggering the drop in pH in the endosomes.
Hydrolases are only active at these low pHs (protective)
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
Ub-mediated Degradation Occurs In MVBs
Ubiquitin tags proteins for degradation by cytosolic proteosomes
Also: Ub-tags on the cytosolic portion of membrane proteins in the early endosome helps trigger formation of a multivesicular body (MVB)
MVBs fuse with lysosomes for digestion