Cell Biology Exam 2 (Intracellular Membrane Traffic)

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what is the progression of protein modification activities in the Golgi (three steps)

1. vesicles leaving the ER form the cis-Golgi 2. the entire cistern matures as it moves distally 3. trans-golgi breaks apart into vesicle for export

ARF does what

ARF protein is responsible for the assembly of both COPI and clathrin coats assembly at Golgi membranes

COPI-coated

Cis-Golgi to ER, cis-Golgi to trans-Golgi, trans Golgi to cell membrane/endosome/lysosome

three types of coated vesicle

Clathrin-coated, COPI coated, COPII coated

what controls the assembly of clathrin coats on endosomes and the COPI and COPII coats on Golgi and ER (also list them)

Coat-recruitment GTPases GEF, GAP, ARF, Sar 1, PIP, Rab(?)

COPII-coated

ER to the cis-Golgi

GAP does what

GAP inactivates the proteins by triggering the hydrolysis of the bound GTP to GDP "GAP - theres a gap in amount of phosphate"

GEF does what

GEF activates proteins by catalyzing the exchange of GDP to GTP "my name Jef = more phosphates"

what do special membrane phospholipids (PIP) do

PIPs regulate coat assembly and Rab protein attachment different PIPs are used to regulate vesicle traffic

Rab effectors are

Rab effectors are downstream mediators of vesicle transport, membrane tethering, and membrane fusion

what can regulate the availability of SNARE proteins

Rab porteins

Rab proteins in GDP bound state

Rab proteins in GDP bound state are inactive and bound to another protein

Rab proteins in GTP bound state

Rab proteins in GTP bound state are active and tightly associate with membrane of an organelle or transport vesicle

Rab proteins do what

Rab proteins play a central part in the specificity of vesicle transport; they bind to Rab effectors

Sar 1 does what

Sar 1 protein is responsible for the assembly of COPII coats at the ER membrane

where are T-snares usually found

T-snare usually found on the target membrane

what do V-snares and T-snares do together

V-snare and T-snare lock together forming a trans-snare complex

where are V-snares usually found

V-snare usually found on the vesicle membranes

destined for immediate delivery to the cell's surface

a constitutive secretory pathway that delivers proteins with no special features to the cell surface

what is the reasoning for autophagy

autophagy can remove large objects - macromolecules, large protein aggregates, and even whole organelles

what do you need to form transport vesicles

coat proteins!

what does dynamin do (two functions)

cytoplasmic protein that pinches off vesicle and also recruits other proteins to bend the patch of the membrane by directly distorting the bilayer structure

after the trans-Golgi, what are the three paths that are protein may take

destined for the lysosome(via endosomes), destined for secretory vesicles, destined for immediate delivery to the cell's surface

what are the pathways in which materials are delivered to lysosomes

endocytosis, phagocytosis, macropincocytosis, autophagy

what is phagocytosis

engulfment

tethering proteins do

extend to link two membranes; on target organelles they bind to Rab proteins

if the target is lysosomes, what is marked with

for the proteins that are glycosylated: mannose-6-phosphate residue on the protein marks it for lysosome

what is another major role of clathrin

forms the outer layer coat of the vesicle

what endocytosis

gains entry into cell without passing through cell membrane

what is macropincocytosis

ingestion of liquid into cell by the budding of small vesicles from cell membrane

what basic functions does the coat do

inner coat gives rise to the vesicle membrane and outer layer coat shapes the vesicle

does entry into vesicles leaving ER happen selectively or default (what is happening molecularly)

it can be either a selective process or happen by default cargo membrane proteins display exit signals on their cytosolic surface that adaptor proteins of the inner COPII coat recognize soluble cargo protein in the ER lumen have exit signals that attach them to transmembrane cargo receptors

what in the heck does a KDEL receptor do

it is a multipass transmembrane protein that binds to the KDEL sequence and packages any protein displaying it into COPI-coated retrograde transport vesicle

what happens in the assembly and disassembly of a clathrin coat (what promotes the assembly and then what happens)

it is initiated by membrane bound receptors; binding of the cargo to the receptors triggers Adaptin, which promotes coat protein assembly; to pinch off the vesicle from the donor membrane, a protein complex called Dynamin is used; following assembly, coat proteins dissociate which leaves transport vesicles

describe the constitutive secretory pathway

it operates in all cells; the TGN to the cell exterior via exocytosis; vesicles form around cargo proteins and are trafficked to the cell membrane; receptors for cargo proteins are not always used

where is there are low affinity for the KDEL sequence

low affinity for KDEL sequence in the ER

how are lysosomes formed

lysosomes are formed by the fusion of transport vesicles budded from the trans Golgi network with endosomes, which contain molecules taken up by endocytosis at the plasma membrane

what do lysosomes contain

lysosomes contain an array of digestive enzymes, most of which are acid hydrolases (active at pH-5);

what triggers macroautophagy (generally, molecules, and vesicle binding protein)

macroautophagy is triggered by cell signaling cascades; the key trigger molecules are mTor (mammalian target of rapamycin), a protein kinase; and vesicle binding protein LC3

what are adaptor proteins (and what do they do)

major coat component in clathrin-coated vesicles, form a discrete inner layer of the coat, positioned between clathrin cage and the membrane binds the clathrin coat to the membrane and traps various transmembrane proteins, including transmembrane receptors that capture soluble cargo molecules inside the vesicle-so called cargo receptors

how are vesicles targeted for lysosomes (first set of steps)

mannose 6 phosphate is added to the precursors of lysomal enzymes. the m6p tagged hydrolases then segregate from all other types of proteins in the TGN because adaptor proteins in the clathrin bind the M6P receptors. the clathrin coated vesicles bud off from the m6p receptors, and the empty receptors are retreived in retromer coated vesicles to the TGN for further rounds of transport

what are lysosomes

membrane enclosed organelles filled with soluble hydrolyctic enzymes that digest molecules

what family are coat-recruitment GTPases a part of

monomeric GTPases

two different type of effectors

motor proteins, tethering proteins

what relation does endocytosis have with lysosomes

much of what lysosomes digest come from endocytosis

what does a transport vesicle do once it is tethered to its target membrane

once a transport vesicle has been tethered to its target membrane, it unloads its cargo by membrane fusion

motor proteins do

propel vesicles along actin filaments or microtubules to their target membrane

how are proteins modified during this stage

proteins are modified in successive stages as they move from cistern to cistern across the attack

destined for secretory vesicles

proteins with signals directing them to secretory vesicles are concentrated in such vesicles part of a regulated secretory pathway that is present only in sepecialized secretory cells

destined for lysosome (via endosome) path

proteins with the mannose 6 phosphate marker are diverted to lysosome in clathrin coated transport vesicles

if the target is secretory vesicle, what is it marked with

recognition sequences that are often tagged with sugar residues

what do resident ER membrane proteins contain

resident ER membrane proteins contain signals that bind directly to COPI coats and are thus packaged into COPI-coated transport vesicles for retrograde delivery to the ER

describe the regulated secretory pathway

selected proteins in the TGN are diverted into secretory vesicles, where the proteins are concentrated and stored until an extracellular signal stimulates their secretion; actively transported by the cytosol

what is autophagy

self eating material; (?). mechanisms used by cells to clear non-functional cellular components or protein clusters/aggregates

why are vesicles necesse est

since proteins are contained within membrane bound organelles, the only way to get them to other compartments is via membrane bound vesicles

what must the KDEL receptor cycle between

the KDEL receptor must cycle between the ER and Golgi apparatus, and its affinity for KDEL sequence must differ in these two compartments

what needs to happen to the SNARE before it can function again (what happens molecularly as well)

the SNARE complex must be pried apart before they can function again NSF uses ATP hydrolysis to unravel the intimate interactions between the domains of the SNARE proteins

what must the transport vesicle incorporate

the appropriate SNARE and Rab proteins

what does the formation of the autophagasome do

the autophagasome fuses to lysosomes that help degrade particules

what happens once the ER functions been fulfilled (where are they moved)

the cells utilize the oligosaccharides for new functions (glycosylation in the Golgi) they move through the CGN, medical cisternae, and TGN

how are vesicles targeted for lysosomes (second set of steps)

the material to be ingested is progressively enclosed by a small portion of the plasma membrane - endocytic vesicle. the endocytic vesicle fuses with early endosomes, where internalized cargo is sorted: some cargo molecules are returned to the plasma membrane by recycling endosomes, and others are designated for degradation by inclusion in late endosomes. membrane proteins destined for degradation are internalized in intralumenal vesicles. the development of late endosomes no longer send vesicles to the plasma membrane, and they fuse with one another and endolysosomes and lysosome to degrade their contents

what is the function of the retrieval pathway

the retrieval pathway returns escaped proteins back to the ER depending on ER retrieval signals

where is there high affinity for the KDEL sequence

there is high affinity for the KDEL sequence in vesicular tubule clusters and Golgi, so as to capture escaped soluble ER resident proteins that are present there at low concentration

how do these lysosomes keep a low pH

these lysosomal membranes have ATP-consuming proton pumps

where are proteins that have entered the ER and are destined for the Golgi apparatus or beyond first packaged

these proteins are first packaged into COPII-coated transport vesicles

what do proteins that are delivered to zones in the Golgi have

these proteins have receptors on the lumenal side and adaptor proteins and coat proteins on the cytosolic side

what must soluble ER resident proteins bind to

these proteins must bind to a specialize receptor protein called a KDEL receptor

what happens as soon as vesicular tubular cluster are formed (what do these things they form do)

they begin to bud off transport vesicles of their own these are vesicles are COPI coated and they function in the retreival pathway which means they carry back ER resident proteins that have escaped, as well as cargo receptors and SNAREs that participate in ER budding and vesicle fusion reactions

what do lysosomes degrade

they degrade material that is endocytosed as well as digest cellular components that must be degraded

what is the function of vesicular tubular clusters

they function as transport containers that bring material from the ER to the Golgi Apparatus

what do clathrin and COPI coated vesicles do after they pinch off

they shed their coat

clathrin-coated directions

trans-golgi to lysoome, cell membrane to early endosome/trans-golgi

how do transport vesicles work

transport vesicles bud off from one compartment and fuse with another; they carry material as cargo from the lumen and the membrane of the donor compartment to the lumen and membrane of the target compartment

what are formed when ER derived vesicles fuse with one another

vesicular tubular clusters


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