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ANAT 262 > Endocytosis IV > Flashcards

Endocytosis IV Flashcards

1
Q

What are rabs

A

Small gtpases
On when bound to gtp
Off when bound to gdp
(2 domains exposed, effectors bind switch 1 and 2)

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2
Q

When rabs bind effectors

A

When in gtp state

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3
Q

Describe effector proteins

A

Typical effector proteins include tethers, snares, mt motors and lipid modifying enzymes

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4
Q

What do rab proteins usually have

A

Usually have one or more fatty acids - 2 typically at c terminus that allows insertion into membranes when in gtp state
After hydrolzyed = special proteins pluck it off membrane
Arf and sar

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5
Q

Describe gap

A

Brings onto membrane - causes hydrolysis and leaves membrane

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6
Q

Name groups of ras superfamily

A

5 main families
Ras
Rho - actin remodeling
Rab
Arf=cop1
Sar1=cop2, effector sec23/24
Ran = nuclear trafficking

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7
Q

Describe ras superfamily porteins

A

All member cycle between active gtp- and inactive gdp bound forms = molecular switches
Rab fam = largest group
Active, gtp bound rab proteins bind to effectors

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8
Q

Describe p115 and rab 1

A

Rab1 gtp bound = binds p115 tethering protein p115 = effector of rab1
Hydrolysis helps control things in refined way

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9
Q

Describe how many rabs in humans and name where most rabs found

A

60 rabs found in humans
Most rabs found on specific organelles
Rab5,7
Diff isoforms and diff specific locations
Rab 1 = on Vtcs, cop2 coated vescles ad cis golgi
Rab6 found on tgn
Rab5 = early endsoome
Rab7 = late endosome

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10
Q

Rab cycle - how are rabs brought to membrane

A

By gefs = gtp exhancge factors
Diff for each rab ~40 of them

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11
Q

Rab cycle - rab binding

A

Rabs in gtp form - and on membrane can bind effectors,can bind many, diff rabs can also bind multiple effectors
Some membranes have more than 1 rab

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12
Q

Rab cycle - rab gap

A

Many
Inactive rabs by causing gtp hydrolysis
= causes loss of binding to effectors
Hydrolysis = turns off rab

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13
Q

Rab cycle - rab gdi

A

Removes rab dip from membranes and sequesters it in cytoplasm - until gef wants it to go to membrane again
Only 3 in humans
Extracts rab from membrane, rab gdi has hydrophobic fatty acids -2 of them in hydrophobic pocket = not exposed to cytoplasm

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14
Q

Are gefs and gaps the same for each rab

A

Different for each rab
Protein gef for one could be gap for other rab

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15
Q

Describe rab effectors

A

Proteins that bind to rabs only when rabs in gtp state
Typically one rab has many efffectos that Carry out diff functions on same organelle- sorta coordinated, will deal with function/communication

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16
Q

Describe SOME rab effector protein functions

A

Tethering proteins -eea1 for rab5, p115 rab1
Regulators of motor proteins
Regulators of lipid metabolism

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17
Q

Describe rab gtpases - regulators of endocytosis

A

Lipid anchored, peripheral membrane proteins
Active rab proteins binds to effectors to coordinate events on vesicle - sorting, unloading, tethering, fusion, locomotion
Can serve as endosoem identity markers

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18
Q

Describe rab localization

A

> 60 rabs In huamns
Often unique rabs coresspond to diff compartments/organelles

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19
Q

Where rab 5

A

Early ednsomes/phagosomes/autophagosomes

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20
Q

Where rab 7

A

Late endosomes/phgaosomes, autophagosomes, lysosomes

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21
Q

Where rab 4,11,9

A

Rab 4 and rab11 = recycling endosoems
Rab 9 = retromer pathways
Many diff roles

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22
Q

Exp = overexpress rab 5

A

= get large early endosomes
Horseradish peroxisome
Enzyme catalyze reaction
Involving diaminobenzidine = seen in ‘em

23
Q

Exp = mutant rab5 cannot bind gap

A

= early endosome very small

24
Q

Exp = cannot hydrolze gtp

A

Huge early endosome
—- concludes = rab5 involved in fusion early endosome

25
Describe rab 5
Master regulator of early endosomes Activated rab5 receipts effectors for tethering and fusion between early endosomes Pi3kinase: p13p production - defines early endosomal membrane
26
Describe phosphoinositides
Minor lipid - found in small quantities,many proteins bind to it Various positions phosphorlated - head group attached to fatty acids Can be phosphorulated to make 7 diff phosoinositide species Sub cellular locations strictly controlled - used as trafficking signals
27
phosphoinositides = which on wher e
Early endosome - pi3p Late endosome - pi(3.5)p2 Important for targeting proteins to early and late endosomes
28
Effectors that bind rab5 and pi3p
Eea1 and rabenosyn 5 require binding to pi3p and rab5 simultaneously
29
What is eea1
Early endosome antigen 1 Filamentous, tethering factor Assembly into home dimers
30
What does eea1 bind to
Binds eea1 on another endosome and then fuses with snares Bind to p13p - fyve domain, rab 5 (2 rab binding domains)
31
Describe process - eea1 in early endosome Fusion
Membranes bind Long shape = membranes brought close together Rab5 recruits exhncage factor for rab7 to early endosome Recruits gap rab5 = causes rab5 to be inactivated and leave early endosome
32
What does rabenosyn 5 do
Rabenosyn-5 believed to mediate cargo to enter either recycling pathways Interact with snares Links to recycling pathways
33
What does rabenosyn 5 bind
Binds to 4 elements =rab5, pi3p, rab4, ehd1
34
Describe rabenosyn 5 and rab4
Fast recycling
35
Describe rabenosyn 5 and ehd1
Slow recycling —> delivery to recycling endosomes Involved in transferrin recycling
36
Describe eea1 and rabenosyn 5 in snare recruitment
Both eea1 and rabenosyn 5 recruit snare components to rab5 active domain Ee1 directly interact with snares syntaxin6 and syntaxin13 Rabenosyn5 indirectly interacts with syntaxins Snares on 2 diff membranes associate —> forms complex —> leads to fusion of membranes
37
What is rab7
Master regulator of late endosomes
38
What does rab7 require
3 proteins for minus end mt transport - tethering proteins
39
Describe rilp - rab7
Recruits and connects late endosome to dynastic dynein machinery
40
Describe orp1l - rab7
Cholesterol sensory Loads dynactin receptor betaIII spectrum Dynactin recruits dyenin motor Biii spectrin binds to late endsoome - cholesterol high = recruits spectrin - recruits dynactin - recruits dyenin - moves towards - end - mtoc
41
Describe rab7 for plus end directed movement
Fyco1 = also recruited, rab effector, competes with rilp binding
42
Describe fyco1 - rab7
Connects late endosome/autophagosome to kinesin for + end directed movement May occur in low cholesterol levels
43
Describe why fyco1/rilp beneficial
Late endosome stays in periphery of cell Helps - cholesterol taken from ldl particle - if cell has increased cholesterol = more ready to fuse with lysosome in centre of cell If low cholesterol = stay at edge
44
Describe rab7 and vesicular tethering
Rab7 effector = hops (homotypic fusion and vacuole protein sorting) complex for homotypic (between late endosomes) and heterotypic (between late endosoem and lysosome) - fusion Also helps recruit retromer coat
45
Describe rab7 and vesicular tethering = subunits
* HOPS complex composed of 6 subunits * VPS33 binds to SNAREs (vacuolar Vam3, Nyv1) * Complex binds to Rab7 on either ends (via VPS39, VPS41)
46
Describe degradative cargo delivery = from early to late end shoes = name models
Pre existing compartment model Maturation model
47
Describe degradative cargo delivery = from early to late end shoes = model 1
Vesicles come off early endosome and take things to late endosome
48
Describe degradative cargo delivery = from early to late end shoes = model 2
Early endsoome turned into late endosome (maturing endosome)
49
Describe maturation from early to late endosome
Change in sub cellular localization of endosome - motors Decrease in acidity - ph, late endosomes more acidic Change in lipid composition - pi3p to pi3,5p2 Closing off access to recycling branch - retain vacuolar portion only and start recycling to Golgi Receive acidic hydrolysis for lysosomal function from tgn Change in fusion machinery - tethers and snares
50
Describe endodosme maturation - gen
Rab5 = controls all needed for early endosome Rab 7 = controls all needed for late endosome So if replace rab5 with rab 7 = early —> late endosome
51
Name and describe the 4 steps of rab conversion
1. Activated rab5 gtp recruits rab7 gef (exhncage factor, recruits rab7 to membrane) 2. GEF Catalyzes exchange from gdp —> gtp on rab7 3. activated rab7 recruits tbc2 (rab5 gap) to catalyze hydrolysis of gtp of rab5 (also removes rab5 gef) 4. Gdp bound rab5 leaves endosomal membrane
52
Describe rab5 specifically - during endosome maturation
Rab5 recruits complex that brings rab7 to membrane Same complex also causes rabex5 (rab5 gef) to leave membrane = leads to loss of rab5 Also rab7 recruits rab5 gap tbc2 = so rab5 regulates its down removal and the conversion of the endosoem from an early to late endosome
53
Where is rab7 found - delivery to lysosomes
On both late endosomes and lysosomes Along with proper snare proteins to fuse So late endosomes and lysosomes can be tethered together with hops complex
54
Describe fusion of endosome with lysosome
Fusion of a late endosome with a lysosome produces a hybrid organelle = characteristics of both = endolyososome As material digested = endolyosome becomes lysosome like again - looks uniform -dense

ANAT 262 (24 decks)

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