Cell Biology Exam 2 (Intracellular Membrane Traffic)
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
