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z:Cell 300 > Lecture 12: Endocytosis > Flashcards

Lecture 12: Endocytosis Flashcards

1
Q

clathrin

A

from TGN forward and PM backward

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2
Q

COPI

A

golgi stack backward

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3
Q

COPII

A

ER forward

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4
Q

endocytosis critical for maintaining ____

A

cardiovascular health

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5
Q

endocytosis (clathrin-dependent transport) hijacked by ___

A

many viruses for entry in cell

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6
Q

clathrin was the 1st ___ due to ___

A

coat protein discovered due to unique structure and number in cell (a lot)

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7
Q

similarities between COPI and clathrin due to convergent evolution

A

-triskelia (3 structure)
-similar cage shape

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8
Q

COPI, COPII, clathrin are ___

A

similar in size

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9
Q

method used to determine structure of COPI, COPII, clathrin

A

X-ray crystal structures

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10
Q

COPI coats combine ___

A

features of clathrin + COPII

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11
Q

COPI and COPII similarities

A

-similar assembly unit

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12
Q

clathrin are made of ____ chains that assemble into ___

A

heavy and light chains
triskelions

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13
Q

clathrin assembly in cages ___

A

happens spontaneously

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14
Q

clathrin ____ membrane into vesicles

A

deforms

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15
Q

clathrin interacts with _____

A

adaptor protein

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16
Q

uncoating of clathrin vesicles requires ___

A

energy; driven by ATP hydrolysis (Hsp70 and auxlin)

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17
Q

Hsp70 is a chaperone that ____

A

binds to newly syn. proteins and uses ATP (involved in uncoating clathrin vesicles

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18
Q

clathrin interacts with adaptor protein with ___

A

its heavy chain

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19
Q

adaptor proteins that interact with clathrin

A

-AP-1, AP-2, GGA

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20
Q

adaptors link ____ to clathrin

A

cargo

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21
Q

assembly of clathrin cycle

A

-adaptor protein binds to cargo/cargo receptor in cytoplasm
-clathrin binds to adaptor protein : forming bud
-clathrin triskeleton forms as vesicles forms + membrane bending and fision proteins form vesicles
-coated vesicle is uncoated with ATP (Hsp70 +auxlin): removal of clathrin + adaptor proteins so vesicle can bind to target membrane

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22
Q

for clathrin transport< soluble proteins ____

A

require receptors for connection adaptor proteins

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23
Q

4 properties of clathrin adaptors (required + optional)

A
  1. Bind cargo
  2. Bind clathrin
  3. Bind phospholipids (PIP2) (optional)
  4. Bind accesory proteins (optional)
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24
Q

the use of specific adaptor depends on _____ and ___

A

intracellualr site
sorting signal

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25
Q

clathrin transport: TGN to endosome adaptor proteins

A

-GGA
-AP-1

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26
Q

clathrin transport: endosome to TGN adaptor proteins

A

-AP-1

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27
Q

clathrin transport: PM to endosome adaptor proteins

A

-AP-2

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28
Q

endocytosis occurs in _____ rich regions of PM

A

-phosphatidylinositol 4,5-bisphosphate (PIP2)

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29
Q

TGN membrane rich with ___ which acts as velcro

A

PIP

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30
Q

different phosphoinositides bind to ___

A

different adaptors

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31
Q

AP-1 are ____ made ______ subunits

A

heterotetramers made of beta (β),gamma (γ) , mu (µ), lowercase sigma (σ)subunits

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32
Q

subunit size of AP-1 (biggest to smallest)

A

-beta (β),gamma (γ),
-mu (µ),
-lowercase sigma (σ)

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33
Q

-beta (β),gamma (γ) subunits of AP-1 are made of ____

A

ears/appendanges, hinges, trunk

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34
Q

AP-1 has binding sites for ___

A

clathrin + cargo

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35
Q

cargo binding to AP-1 needs which motifs + which subunits bind?

A

LL (B) and YXXO (µ)

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36
Q

retrival of M6PR from endsomes to TGN and sorting of M6PR from TGN to endosomes need which adaptor protein in CCVs?

A

-AP-1

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37
Q

For binding to AP-1, cargo without LL binds to ___

A

µ subunit via YXXO motif

38
Q

GGA stands for

A

(Golgi-localized g-ear-containing ARF-binding
protein)

39
Q

GGA are ____ adaptors that link clathrin to cargo in _____

A

monomeric modular adaptors that link clathrin to cargo in an ARFdependent (GTPase) manner

40
Q

GGA are responsible for sorting M6PRs from ___

A

TGN to endosomes

41
Q

GGA bind to M6PR with motif ___

42
Q

GGA requires ___ to bind to cargo

43
Q

AP-2 functions at ____

44
Q

AP-2 is ___ adaptor protein

A

heterotetramer

45
Q

subunits of AP-2

A

α,β2,µ2, σ2

46
Q

α subunit of AP-2 binds to _____

A

amphiphysin, EPS15 + others

47
Q

β2 subunit of AP-2 binds to _____

A

clathrin (appendages) and di-leuicine (body)

48
Q

µ2 subunit of AP-2 binds to ___

A

YXXO motif

49
Q

amphiphysin, EPS15 + others are ___

A

accessory proteins that regulate assembly

50
Q

AP-2 has ___ -binding domains

A

clathrin,acessory protein,cargo, PIP2

51
Q

Binding of ____ on membrane induces conformational change in AP2 that ___

A

PIP2
that exposes cargo binding sites (ap2 locked to ap2 open)

52
Q

assembly of clathrin coats at PM

A

-recruitment of nucleating proteins (FCHO1/2) that create membrane curves at domains rich with PIP2
-Intersectin and EPS15 link FCHO protein to AP2 that links to clathrin

53
Q

FCHO1/2 are ___ proteins that induce ____ at ____

A

nucleating
membrane curvature
PM domains rich with PIP2

54
Q

FCHO1, 2 are _____ proteins that
bind to and bend membranes (— charged)

A

BAR domain-containing (++++ charged)

55
Q

FCHO1, 2 then bind scaffold proteins _____that link it to ____

A

-Eps15 and Intersectin
-AP2 complexes

56
Q

CCVs creation at PM

A

-recruitment of FCHO, EPS15, intersection >membrane curvature
-AP2 recruited and bind to cargo
-clahtrin recuited by AP2
-GTPase activity by Dynamin for vesicle scission
-auxilin incorporated into triskelion structure and recruit Hsp 70 (ATP hydrolysis)- uncoating

57
Q

ATP hydrolysis by ___ (with auxilin) drives uncoating of CCVS

58
Q

auxilin incorporated into ___

A

structure of clathrin triskelia

59
Q

dynasore (def.)

A

small molecule that blocks dynamin function

60
Q

endocytosis motifs are similar to ____

A

lysosomal sorting motifs

61
Q

endocytosis motifs

A

-tyrosine based (NPXY & YXXO)
-dileucine (LL), phsophoserine

62
Q

clearing LDL from blood serum with endocyotsis

A

-LDL receptor + ligand binding promotes recruitment of receptor coated pits
-vesicles uncoated and forms endosomes
-lysosome forms and LDL is broken down
-at neutral pH, endosome releases receptor to bind to another LDL particle

63
Q

Genetic mutations in LDL receptor gene (1/500) _____can lead to increased risk ____

A

lacking targeting domain (no LL, tyrosine motif)
of heart attack due to atherosclerosis

64
Q

true or false: all ligands are degraded after endocytosis

65
Q

example or ligand that isn’t degraded after endocytosis

A

Ferrotransferrin (ligand + iron) binds to transferrin receptor –> CCV –> uncoating –> low PH release iron from ligand but ligand still bound to receptor) –> ligand (apotransferrin) retained and recycled to surface, ligand released at neutral pH

66
Q

scission of coated vesicles driven by ___

A

GTP hydrolysis

67
Q

Dynamin is a ___

68
Q

mutation in or near GTPase domain of dynamin lead to ____

A

blocking of scission of CCVs from PM

69
Q

dynamin works at ___

70
Q

dynamin has mutliple functional domains

A

-GTPase domain (4 domains together)
-PHD domain
-PRD domain
-GED domain

71
Q

GTPase domain of dynamin does what?

A

-hydrolyzes GTP

72
Q

PHD domain of dynamin does what?

A

binds to PIP2 (inositiol phospholipids)/acidic phospholipids in PM

73
Q

PRD domain of dynamin does what?

A

binds to proteins with SH3 domains like amphiphysin

74
Q

GED domain of dynamin does what?

A

has GAP function activates GTPase
increase rate of GTP hydrolysis

75
Q

mutation in GTPase domain of dynamin does what?

A

decrease endocytosis

76
Q

mutation in PHD domain of dynamin does what?

A

decrease endocytosis

77
Q

mutation in GED domain of dynamin does what?

A

increase endocytosis

78
Q

mutation in PRD domain of dynamin does what?

A

blocks endocytosis

79
Q

GTPase activity of dynamin in different forms

A

-very active (2-100 GTP)- rings, collars
-increase 100-1000x at necks of coated vesicles, spiral form

80
Q

GTP hydrolysis results in pitch change of dynamin spirals and promotes ____

81
Q

amphiphysin is ____

A

a link between CC PIT and dynamin through AP2

82
Q

FCHO, AP2, Clathrin, Amphiphysin, dynamin, EPS15, Intersectin interactions

A

FCHO to Intersectin /EPS15 to AP2 to clathrin
AP2 to Amphiphysin to dynamin

83
Q

domain of Amphiphysin that links to AP-2

83
Q

domain of Amphiphysin that links to dynamin

A

SH3 domain to PRD domain of dynamin

84
Q

regulatory view of dynamin

A

Oligomerization and increased GTPase activity inhibits
endocytosis –> GTP bound dynamin controls effectors that mediate vesicle formation

85
Q

mechanochemical view of dynamin

A

GTP hydrolysis and conformational change in dynamin are required for endocytosis

86
Q

Unifying model for dynamin mechanism

A

originally in regulatory form (tetrametric) (slow GTP hydrolysis) then mechanochemical (spiral) form (fast GTP hydrolysis)

87
Q

Switch between functional states of dynamin regulated by ____

A

SH3 domain containing binding partners (ex. amphiphysin) that sense critical events in CCV maturation

88
Q

______ of dynamin controls rate-limiting step in endocytosis (CCVs formation)

A

slow form of GTPase (tetramer)

89
Q

mutation in GTPase domain of dynamin keeps____

A

dynamin in rate-limit step longer increasing endocytosis

z:Cell 300 (17 decks)

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