Week 8 pt II - And Exocytosis Flashcards
COPI vesicles
Recruitment of GTPase called SARI into ER membrane
What happens when there is GTP to GDP exchange?
It sticks out amphiphilic alpha helix that is integrated into the membrane
Starts the membrane curvature
What are examples of COP proteins?
COP16
COP24
What is the outer code?
COP30
COP31
What occurs after the outer code?
The vesicles will bud off
It has to lose the protein coat before it can fuse with the target membrane (occurs by GTP hydrolysis)
What are COPI vesicles?
Vesicles that bring back from the Golgi to the ER
What does COPI vesicles bring?
Stuff from Trans Golgi back to Cis Golgi
What are the enzymes that are necessary for the Golgi to perform its function?
Manase
Glycosyl transferase
How are Manase and glycosyl transferase recruited?
Through receptors and binding proteins
What is Arf1?
GTPases related to group of proteins called Ras proteins
What happens when GTP is hydrolysed to GDP?
The protein coat is lost
COP1 vesicles can fuse very easily with ER
How was COP vesicles discovered?
Fed mammalian cells with non-hydrolysable GTP
What binds GTP?
ARf1 and SAR1
What does ARf1 and SAR1 do?
Recruit all the proteins and form vesicles
GTP cannot be hydrolysed
Protein coat is not lost
Don’t get fusion with any other membrane
What was observed when cell lines were fed with non-hydrolysable GTP?
Massive build up of vesicles all throughout the cell
Trans Golgi Network
Sorting of things, anything that leaves the Golgi and goes through lysosomes
What occurs at Trans Golgi Network?
Things going to the plasma membrane to export proteins
Things going to form lysosomes
Constitutive export and regulated export
What do Lysosomes Contain?
Nearly 50 hydrolytic enzymes
Specialised for breaking different kinds of binds
Destroying different types of Proteins and lipids
What are the hydrolytic enzymes never allowed to touch?
Cytosol
When are the hydrolytic enzymes active ?
At very acidic PH (acidic hydrolases)
Where are the hydrolytic enzymes initially synthesised?
ER and Golgi (PH is neutral)
What does Golgi add?
Further processing to the Mannose chains to produce the unique signature for lysosomes
Where are lysosomal proteins synthesised and delivered to?
Synthesised: ER
Delivery: lysosome
What happens in the Golgi complex (cis-cisternae)?
Lysosomal enzymes are recognised by enzymes that catalyse the two step addition of phosphate group to mannose sugars of N-linked carbohydrate chains
What are the enzymes that catalyse the two step addition of phosphate group to Mannose sugars?
N-Acetylglucosamine
Phosphodiester glycosidase
What does N-Acetylglucosamine phosphotransferase do?
Transfers 2 N-acetylglucosamine from a nucleotide sugar donor phosphate to Mannose residues of N-linked oligosaccahrides
What is UDP glucose used for?
Form a phosphodiester bond with one of Mannose residues
What does phosphodiester glycosidase enzyme cleave?
2 glucosamine
Mannose 6-phosphate residues as part of the oligosaccharide chain
What is the signature for lysosomes?
Mannose 6-phosphate motifs
Mannose 6-phosphate receptors
Interact specifically with lysosomal enzymes on luminal side of the vesicle
Interact specifically with adaptors on the cytosolic surface of vesicle
What is the result of M6PR on TGN membrane?
Lysosome enzyme on TGN become concentrated into clathrin coated vesicles
What do clathrin coat form?
Vesicles which bud off after formation of clathrin cage
Migrate the vesicle to the cytosol until they fuse with a late endosomes
Why are Mannose 6-phosphate receptor present in the plasma membrane?
Capture lysosomal enzymes that are secreted into extracellular space
Return the enzymes to a pathway that directs them to a lysosome
The vesicles released from Golgi Contain?
Proton pump that use the energy of ATP to transport hydrogen ions from cytoplasm to the interior
What happens as the vesicle travels in the cytoplasm?
PH drops
What is the PH of late endosomes?
Lower than that of Golgi
What does change in PH used by?
Cell to recycle the receptors of lysosomal enzymes
What is the binding affinity of the receptors for Mannose 6-phosphate m?
Very PH sensitive in the mild acidic range
What happens at PH 6.5?
Endosomes release lysosomal enzymes
Destabilise the structure of Mannose phosphate receptor
Disrupts the interaction between lysosomes and mannose phosphate receptor
What does the proton pump in the membrane of lysosomes keep the PH around?
5
What are the lysosomal enzymes bound onto?
Mannose 6 phosphate receptor
What is the name of the Adaptor?
Golgi associated gamma adaptin (GGA)
What is the microtubule directly stretched between?
Organelles
What is found within microtubules?
Big bulges of vesicles
The smaller one travelling up the thin horizontal
What do vesicles move along?
Microtubule
What is Hand over slide (Hamburg)?
Beta tubulin on the microtubule
ATP hydrolysis and one foot goes over and next foot slides along
The red foot moved over and stored kinetic energy and the blue foot slides across
Happens at molecular level
Hand over hand
Kinesin molecules walk down the microtubule inside the cell transporting its cargo
The adaptor is attached to gigantic vesicle
How does HIV get to the nucleus?
On Dynein and kinesin
Motor proteins
What does Fibrous Tethering protein mediate?
Initial contact between transport vesicles and it’s target membrane
Fibrous tethering proteins
Form molecular bridge between two Membranes over a large distance (50-200nm)
What is tethering?
Early stage of vesicle fusion that requires specificity between vesicle and target membrane
What are RAB proteins on vesicle and target membrane involved in?
Recruiting specific tethering proteins that mediate initial contact between two Membranes
What does SNARES mediate?
Vesicle fusion
What do SNARE proteins have?
More than 35 membrane proteins
What does SNARE motif consist of?
60-70 amino acids which can form a complex with another SNARE motif
What is the most studied SNARE proteins?
Mediating rocking of synaptic vesicles with presynaptic membrane during the release of a NT
What are the two types of SNARES?
V-snares
T-snares
What is V-snare?
Incorporated into vesicle during budding (synaptobrevin)
What is T snare?
Located on the target membrane surface (SNAP-25 and syntaxin)
What happens in synapses and epithelial cells?
Regulated exocytosis contents being released into the environment
What is Kidd and Release model?
The vesicle bind very briefly
Opening up of the pore and the content is released and the vesicle can move away
Why do all cells endocytose constantly?
To replace things in the plasma membrane
Refresh lipids
Change structure
What is receptor mediated endocytosis?
The receptor bind cargo proteins
Outside: start of a indent
High density: concentrated yolk protein being accumulated in the life of the start of the formation of vesicle
Greater density on the inside of cell on other side of membrane - assembly of the protein coat to produce new vesicles
Vesicle pinching: dense coat on inside and concentrated cargo on the outside of cell
What is Latrice made up from?
Protein called clathrin
What is clathrin made up of?
Triskelion
What does Triskelion Contain?
3 heavy chain
2 light chain
What is the lumen of endosomes acidified by?
H+-ATPase
Where is early endosomes located?
Near cell periphery
What do early endosomes do?
Sort Materials
Send bound ligands to the late endosomes
Where are late endosomes located?
Near the nucleus
What are late endosomes known as?
Multivesicular bodies (MVBs)
What are the properties of signalling receptors?
Bind to extracellular ligand that carry messages
Marked for endocytosis by ubiquitination
Sorted into membrane that form internal vesicles in late endosomes
Late endosomes fuse with Lysosomes where receptors are degraded
What are the properties of house keeping receptors ?
Responsible for uptake of materials
Dissociation from bound ligands triggered by low PH in endosomes
Recycled to the plasma membrane
