Section 7C: Vesicle tethering and fusion

Question 1

What does vesicle targeting to a compartment involve?

Answer 1

1. Tethering
2. Rab GTPase

Question 2

What is Tethering?

Answer 2

• binding loosely to the correct target protein
• does not mean material can go easily, they’re just held in proximity to where they need to eventually fuse and deliver their content; but those contents aren’t delivered yet

Question 3

Which proteins carry out the process of tethering?

Answer 3

Rab GTPases

Question 4

What are the two steps for membrane fusion?

Answer 4

1. tethering
2. docking/fusion

Question 5

Rab GTPase-mediated tethering to target organelles of transport vesicles (steps)

Answer 5

• a specific Rab can be activated either on vesicle or target membrane
• GTP-bound Rab binds to Rab “effector proteins”
• Rab effector proteins carry out the Rab regulated function
• Rab effectors are myriad: enzymes, motor proteins, tethers, etc

Question 6

Every vesicle tends to have a ______ Rab protein

Answer 6

unique

Question 7

What are the unique Rab proteins responsible for?

Answer 7

Each one of them are responsible for taking a vesicle to a unique target compartment

Question 8

Rab 5

Answer 8

Early endosomes (endocytosis - degradation and recycling)

Question 9

Rab 7 and Rab 9

Answer 9

Lysosome (endocytosis - degradation)

Question 10

Rab 2

Answer 10

Golgi —> ER (retreival)

Question 11

Rab 1

Answer 11

ER —> Golgi (synthesis)

Question 12

What does targeting and fusing vesicles to the right organelle involve?

Answer 12

1. Rab GTPases
2. SNARE proteins

Question 13

What are the two different types of SNAREs

Answer 13

1. Vesicular SNARE (v-SNARE): on vesicle
2. Target SNARE (t-SNARE): on target
   - these two are good at binding to each other

Question 14

What causes the lipid bilayer to fuse?

Answer 14

When v-SNARE and t-SNARE bind to each other very tightly they form a trans-SNARE complex

Question 15

Why does at least one t-SNARE protein have to be a transmembrane or peripheral membrane protein?

Answer 15

• 2 proteins (v-SNARE and t-SNARE) come together, forces the membranes together because those two proteins are anchored in the lipid bilayer
• if they were not anchored, they could just get ripped off the membrane

Question 16

What happens if SNARE proteins don’t work?

Answer 16

Neurotransmitters won’t be released

Question 17

v-SNAREs

Answer 17

transmembrane, single chain

Question 18

t-SNAREs

Answer 18

2 or 3 chains, at least 1 transmembrane, others may be peripheral membrane proteins

Question 19

How does botox work?

Answer 19

It degrades certain types of SNARE proteins that are involved in neurotransmitter release; causing muscles to relax (stops muscle contraction)

Question 20

Sequential steps of SNARE-mediated membrane fusion

Answer 20

1. Trans-SNARE complex formation causes water exclusion: energetically unfavourable
2. Membranes are within 1.5 nm of each other: sufficient to cause lipids to cross layers
3. Stalk formation: one layer of the membrane undergoes fusion (outer leaflets fuse, the inner leaflet of the lipid bilayers do not mix yet)
4. Hemifusion: 2nd, inner layer is juxtaposed, permitting lipid molecules to move across the inner layer of the membrane (inner leaflets now mix)
5. Full fusion occurs, two membrane compartments become one

Question 21

Are SNARE proteins going from individual v-SNARE and t-SNARE proteins into a SNARE complex favourable or unfavourable?

Answer 21

Favourable

Question 22

How does a cell invest energy to reset for the next vesicle that has to feed?

Answer 22

It separates the v-SNARE from the t-SNARE after fusion has taken place; away to recycle the proteins

Question 23

What is NSF?

Answer 23

an enzyme that hydrolyzes ATP or uses ATP hydrolysis to separate v-SNAREs from t-SNAREs (the SNARE complex)

Question 24

If a cell no longer has ATP will it be able to sustain vesicle fusion?

Answer 24

No, because it will run out of the “free” v-SNAREs and t-SNAREs and won’t be able to regenerate them

Question 25

Why are both the SNAREs in the same compartment after full fusion?

Answer 25

Because the vesicle became a part of the target compartment; the target compartment absorbed the vesicle as well as all its contents