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BIOL 201 > Topic 7: Endomembrane System > Flashcards

Topic 7: Endomembrane System Flashcards

1
Q

What is the endomembrane system?

A

the set of organelles interconnected via vesicles

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2
Q

How are proteins targeted to each of the destinations in the endomembrane system?

A

translation of all proteins starts in cytoplasm

sorting signals that all localization

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3
Q

What are sorting signals?

A

all translation begins in the cytoplasm on free ribosomes

a continuous stretch of amino acid sequence, typically 15-60 amino acids long

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4
Q

What is the ER signal sequence?

A

generally at N terminal, contains block of hydrophobic amino acids

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5
Q

What is the nuclear localization signal sequence?

A

one or two short sequences with positively charged amino acids

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6
Q

What is the mitochondria/chloroplast signal sequence?

A

organelle specific signals at N terminal

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7
Q

What is the ER lumen “retention” signal sequence (KDEL)?

A

this four amino acid sequence causes proteins to be kept in the ER lumen “retention signal”

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8
Q

What is the secretory pathway?

A

proteins translated by bound ribosomes are either destined for the ER or for other compartments within the endomembrane system

one such destination is the plasma membrane for secretion

a good example of this type of transport is neurotransmitter release into the synapse after an action potential

once a protein is in the endomembrane system they never see they cytoplasm again

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9
Q

What are the steps in the secretory pathway?

A

translation by bound ribosomes in ER

from ER, vesicular transport moves cargo and membranes along the secretory pathway

proteins destined for secretion move: ER –> Golgi –> PM

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10
Q

What is the pulse-chase experiment?

A

method for determining location of radioactively labelled compounds

fed living cells radioactive amino acids “the pulse”

fixed the cells for TEM, cut sections, after various timepoints after pulse

dipped sections on grids into photographic emulsion, wait while decay events expose film

develop photographic emulsion

see pots on areas containing radioactivity

do “pulse-chase”, give pulse of radioactivity then a time period (“chase”) without radioactivity

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11
Q

What are Sec mutants?

A

yeast (Saccharomyces cerevisiae) with mutations in key proteins

mutants blocked in different steps of ER-Golgi-PM transport

GFP-secretory protein fusions tracked with confocal “green fluorescent protein” ER resident fusion or thin section TEM

GFP cargo fusions were put into sec mutants = mutant had accumulated of GFP in different compartments

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12
Q

What is the structure and functions of the ER?

A

controls calcium levels

site of lipid synthesis

site of protein translation and folding

cargo synthesized at ER moves to the cis-Golgi

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13
Q

How does cargo travel from organelle to organelle?

A

use vesicles

coat proteins drive the formation of vesicles

cop II, cop I, clathrin

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14
Q

What are Cop II coated vesicles?

A

ER to cis-Golgi, “coatomer”, soluble protein complex that forms coat is recruited from the cytoplasm

in vivo: Cop II molds the vesicle

spontaneously self-assemble into cage-like structures

coat proteins use adapters to associate with membrane and regulate assembly

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15
Q

What is the major cargo of Cop II proteins?

A

vesicles that move from ER –> cis-Golgi use Cop II (anterograde transport)

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16
Q

What are Cop I coated vesicles?

A

Golgi to ER recycling

possibly used in intraGolgi traffic - not known for sure

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17
Q

What is the major cargo of Cop I coated vesicles?

A

proteins that are supposed to stay in ER may accidentally be transported to Golgi (Cop I vesicles recycle back to ER)

ER resident proteins contain KDEL sequence, therefore there must be KDEL receptors in both ER and Golgi to read KDEL sequence, KDEL receptors link via adapter protein to Cop I coats

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18
Q

What are clathrin coated vesicles?

A

PM to endosomes and Trans Golgi to lysosome

clathrin has a distinctive morphology

coatomer coated vesicles (COPII and COPI) are hard to see in thin section/TEM, look like fuzzy coats instead of spiky like clathrin

once a vesicle is formed and uncoated, it must fuse with the correct target membrane

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19
Q

What is the major cargo of clathrin coated vesicles?

A

receptor-mediated endocytosis (RME)

moves cargo from membrane to endosomes (transient membrane band compartment before lysosome)

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20
Q

What are the steps of the formation of clathrin coated vesicles?

A

ligand binds to membrane associated receptors

adaptor proteins cluster receptors on a specific area of membrane

clathrin associates after ligand binding to shape the vesicle

vesicle fission needs other assoc. proteins, eg. dynamin/dynactin (wraps around neck and stretches)

clathrin dissociates once vesicle forms

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21
Q

What is vesicle fusion?

A

targeting of vesicles to specific organelles requires G proteins (Rab)

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22
Q

What are Rab GTPases?

A

docking and tethering vesicles to target membrane

many different kinds of Rab GTPases (on mature vesicle), associate with different vesicles and help direct them to the appropriate destination (not sufficient for specificity)

activated by GEF’s on the donor membrane

23
Q

What are the steps of Rab GTPase docking and tethering?

A

mature vesicle = no coat proteins

SNARE’s allow membrane fusion

part of destination specificity Rabs can exist in GTP or GDP bound forms

tethering protein allows Rab specificity

specificity protein at target membrane

vSNARE/tSNARE combo gives more specificity

proximity of vesicle to target allow vesicle fusion

24
Q

How is membrane fusion is selective?

A

SNARE proteins: transmembrane proteins

specificity and catalyzing vesicle fusion with target membrane

25
Q

How do signal sequences on cargo proteins aide in direction and localization?

A

sequence tags

there is information on cargo that facilitate localization

e.g. M6P: mannose-6-phosphate tag (sorting signal for lysosome)

26
Q

What are SNARE proteins?

A

bring membranes close together, displace water and promote membrane fusion

helical domains: v and t SNARE form twisted complexes that pull membranes together

27
Q

What are v-SNARE proteins?

A

associated with vesicle membranes

large cytoplasmic domain

many different varieties

28
Q

What are t-SNARE proteins?

A

on target membranes

specific to v-SNARES

29
Q

What is the structure of the Golgi?

A

Golgi apparatus refers to all the stacks in a cell

each flattened membrane disk is called a cisterna, plural cisternae (cis/medial/trans cisternae)

each cisterna has a unique complements of proteins associated with it

facing the Golgi lumen are many membrane-bound enzymes that make and modify polysaccharides

each stack has a cis (forming) face and trans (leaving) face with a network of membranes at each face

30
Q

What are the two directions of movement of material through the Golgi?

A

anterograde: forward movement, cis to medial to trans

retrograde: backwards movement, believed to be for recycling of escaped residents

31
Q

What are the two models of how materials move through Golgi?

A

vesicle trafficing model

cisternal progression model

32
Q

What is the vesicle transport model?

A

receive vesicles from ER to the cis Golgi

moves to the medial Golgi using vesicles

uses vesicles to move

observed vesicles interacting with Golgi and cisternal proteins

33
Q

What is the evidence for the vesicle transport model?

A

there are cisternae specific enzymes

vesicles are observed associating with Golgi

34
Q

What is the cisternal progression model?

A

each cisterna forms from the fusion of vesicles, then “matures”, along with its cargo as it moves through the stack cis to trans

whole of medial Golgi matures into trans Golgi

cargo stays inside, no vesicles

Golgi is “moving” in a way

molecules may be too big to fit in vesicles

35
Q

What is evidence for the cisternal progression model?

A

large molecules (too big to fit in vesicles associated with Golgi) pass through Golgi (eg. procollagen)

36
Q

How do both models contribute to movement of materials through the Golgi?

A

anterograde: use cisternal maturation, ER –> Golgi

retrograde: go back because KDEL was read wrong, use vesicles, vesicles from ER form new cis Golgi

37
Q

What is the function of the Golgi?

A

glycosylation, packaging and sorting of glycoproteins

38
Q

What is glycosylation?

A

covalent addition of sugars

function: cell-cell recognition, protection from dehydration and form recognition by pathogens

glycosylation starts in ER and completes stepwise through Golgi cisternae

therefore glycosylation is only found on extracellular surface or facing lumen of endomembrane organelles

39
Q

How does glycosylation add oligosaccharides to proteins?

A

it is rare to have glycoproteins in cytoplasm

oligosaccharide processing occurs in ER and Golgi

first step: sugar tree is added in ER as the oligosaccharide is transferred off a glycolipid

40
Q

What is constitutive secretion?

A

unregulated secretion, “housekeeping”

steady stream of vesicles to PM that every cell has all the time for housekeeping (modify, turn over PM proteins, growth and secretion of some proteins)

41
Q

What is regulate exocytosis?

A

contents made (hormones, mucus, etc.) and bud from TGN then vesicles stored full of material close of PM

when signal comes, exocytosis occurs

secretory granules dock at the cell surface and release their contents when they are stimulated to do so (due to rise in intracellular calcium concentration)

42
Q

What is the lysosomal pathway?

A

lysosome contain hydrolytic enzymes

proteins destined for lysosome contain M6P tag (glycosylation)

M6P receptors in TGN (collect and concentrate M6P containing cargo)

late endosome = low pH, causes activation of M6P cargo/receptor (receptors recycled back to TGN)

late endosome matures into lysosome where endocytosed cargo is degraded by M6P-containing hydrolytic enzyme

43
Q

What is the endocytosis pathway?

A

endocytosis is the process where vesicles bud off the plasma membrane and internalize membrane and extracellular fluid contents

pinocytosis and receptor-mediated endocytosis

44
Q

What is pinocytosis?

A

cell drinking

also known as bulk-phase endocytosis

brings about uptake of extracellular fluids without recognition by surface of plasma membrane

45
Q

What is receptor-mediated endocytosis?

A

brings about uptake of specific macromolecules (ligands) following their binding to receptors on plasma membrane

a ligand is any molecule that can bind a receptor

46
Q

What are endosomes?

A

network of membranous tubules

47
Q

What are early endosomes?

A

associated with periphery of cell

neutral pH

M6P (-)

48
Q

What are late endosomes?

A

are located more interior of the cell

lower pH

M6P (+)

49
Q

What are coated pits?

A

sites on membrane where receptors for receptor-mediated endocytosis are concentrated

pits contain clathrin and adaptin that associated with intracellular domain of receptor

coat protein cluster receptors and forms vesicle

50
Q

What are the steps of receptor mediated endocytosis?

A

receptor on cell surface binds to molecules to be taken up (ligand)

fuse with endosome; acidic interior causes dissociation of ligand from receptor

ligand degraded following transfer to lysosome

receptor can be recycyled

51
Q

What organelles do materials pass through in the secretory pathway?

A

ER –> Golgi –> PM

52
Q

What organelles do materials pass through in the lysosomal pathway?

A

ER –> Golgi –> lysosome

53
Q

What organelles do materials pass through in the endocytosis pathway?

A

PM –> endosome –> lysosome

BIOL 201 (13 decks)

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