rab proteins

Question 1

what are rab proteins and what do they do?

Answer 1

• GTPases
• regulate specific trafficking steps/sorting
• distinct subcellular localisations
• are required for fusion
• cycle on and off membrane (as are small GTPases)

Question 2

what family are rab proteins a member of?

Answer 2

ras superfamily

Question 3

what are the 2 different forms of rab protein?

Answer 3

GTP form - ACTIVE at membrane

GDP form - inactive - in cytosol?

Question 4

what allows rab to cycle on and off of the membrane?

Answer 4

lipid grouup - can anchor rab into membrane

Question 5

what is GDI?

Answer 5

GDP dissociation inhibitor

• chaperones protect the lipid group from soluble cytoplasm
• has some role in targetting correct rab to correct membrane

Question 6

how is rab recuited to the membrane?

Answer 6

by GEF

• guanine exchange factor
• replace GDP with GTP - so rab is active at the membrane

Question 7

what does GAP stand for and what does this do?

Answer 7

• GTPase activating protiens

- removed GTP from rabs (to GDP) - allows dissociation from the membrane + recycling to occur

Question 8

can rabs recruit different effectors?

Answer 8

yes

effector e.g V-SNARE

Question 9

where does rab5 localise to?

Answer 9

• localises to endocytic pathway - early endosome

- has roles in endosomal fusion

Question 10

what happens in a rabGTP mutant?

Answer 10

• rab cannot dissociate from the membrane

Question 11

what happens in rab GDP mutants?

Answer 11

all of the GEFs are occupied by GDP mutant rab so endogneous protein cannot bind to GEF

Question 12

what happens when rab5Q79L is overexpressed?

Answer 12

• constitutionally active form of rab5GTP

- means that rab5 cannot dissociate from the membrane- formation of large endosomes and lots of docking sites

Question 13

how can rab proteins define membrane domains?

Answer 13

enrichment of one rab over another in certain areas of the endosome
- this is a way of segregating different receptors

Question 14

give the roles that Rab5 has?

Answer 14

Rab5 regulates multiple events in the early endocytic pathway

• quality control role - makes sure vesicles fuse with right membrane
• movwement of endosomes
• sorting cargo
• • uncoating clathrin
• tethering
• fusion

Question 15

How does rab5 have so many different roles?

Answer 15

• it is able to bind to different effector molecules

e. g. different tethering proteins

Question 16

what does GST stand for in GST pull down?

Answer 16

-glutathione - S - transferase

Question 17

How does a GST pull down work?

Answer 17

• attach GST to a protein
• beads capture GST and as GST is also bound to proteins of interest this pulls the protein down too
• beads coated in GSH which binds to GST
• proteins pulled done run on gel

Question 18

what rabs label the late endosome?

Answer 18

rab7 and rab9

Question 19

what does rab conversion facilitate?

Answer 19

rab conversion (e.g. rab 5 to rab 7) allows movement of materials along the endocytic pathway, e.g. from early to late endosome.

Question 20

How is rab conversion from rab 5 to rab 7 possible?

Answer 20

Rab5GAP and Rab7GEF are in a complex/close proximity to each other

-therefore rab5 can be inactivated and rab 7 can become activated

Question 21

what is EEA1?

Answer 21

• it is an early endosomal marker

- tethering protein (rab effector) which allows vesicle to come into close proximity of its target membrane

Question 22

what is rabaptin?

Answer 22

rab effector which can bind to rabX5 (GEF)

Question 23

what does sorting into an MVB usually result in?

Answer 23

degradation