12.1 The Endoplasmic Reticulum Flashcards
- a network of membrane-enclosed tubules and sacs that extends from the nuclear membrane throughout the cytoplasm
- largest organelle of most eukaryotic cells
endoplasmic reticulum
rough ER is covered with (1), and thus it mainly functions in (2)
- ribosomes
- protein processing
() is not associated with ribosomes and is involved in lipid metabolism
smooth ER
when cells are disrupted, the ER breaks up into small vesicles called ()
microsomes
what are the sacs in the ER called
cisternae
() is the first step in protein trafficking along the secretory pathway
Entrance into the ER
using the (), George Palade and his colleagues were able to show how proteins are secreted
pulse-chase experiment
how were newly synthesized proteins labelled in the pulse-chase experiment
using radioactive amino acids that the pancreatic cell was exposed to
state the secretory pathway
rough ER → Golgi → secretory vesicles → cell exterior
what is the “pulse” stage of the pulse-chase experiment
exposure of cell to radioactive AAs
what is the “chase” stage of the pulse-chase experiment?
incubating the cells in a media with nonradioactive AA
the site of secretory protein synthesis
rough ER
how do proteins travel to the cell surface from the Golgi
they are packaged in secretory vesicles
proteins whose destinations are the cytosol, mitochondria, chloroplasts, or the nuclear interior are synthesized by ()
free ribosomes
compare and contrast free ribosomes from membrane-bound ribosomes
free and membrane bound ribosomes are virtually indistinguishable; a ribosome will bind to the ER membrane only if the protein it is translating needs to be secreted
what are the 2 kinds of translocation
- cotranslational translocation
- posttranslational translocation
compare and contrast cotranslational from posttranslational translocation
- in cotranslational translocation, proteins are translocated into the ER during their translation
- in posttranslational translocation, proteins are translocated into the ER once translation has been completed
an amino acid sequence that the N-terminus of a polypeptide chain that signals the ribosome to be targeted to the ER membrane; it is cleaved during the polypeptide chain’s transfer into the ER lumen
signal sequence
the signal sequence is cleaved by () during translation
signal peptidase
this hypothesis posited that a short amino acid sequence was responsible for targeting ribosomes to the ER membrane when they are translating secretory proteins
signal hypothesis
how did experiments on the signal hypothesis confirm the role of signal sequences?
- (secreted) proteins translated by free ribosomes were slightly larger because the signal sequences weren’t cleaved
- additionally, the addition of a signal sequence to a normally non-secreted protein is sufficient to direct the recombinant protein into the ER
signal sequences are recognized and bound by the ()
signal recognition particle (SRP)
the SRP consists of (1) and (2)
- 6 polypeptides
- a small cytoplasmic RNA (SRP RNA)
the SRP binds to the ribosome and signal sequence, and then targets the entire complex to the rough ER by binding to the () on the ER membrane
SRP receptor
what do you call the protein translocation complex on the ER membrane that the ribosome binds to
translocon
() opens the translocon by moving a plug away from the translocon channel
insertion of the signal sequence
in both yeast and mammalian cells, translocons are complexes are complexes of 3 transmembrane proteins called ()
Sec61 proteins
summary of cotranslational targeting of secretory proteins into the ER
- once signal sequence emerges from (i.e. is translated by) the ribosome, it is recognized and bound by the SRP
- SRP escorts the ribosome-polypeptide-SRP complex to the ER membrane where it binds to the SRP receptor
- SRP is released and ribosome binds to translocon; insertion of signal sequence opens translocon channel
- translation resumes and the signal sequence is cleaved by signal peptidase
- continued translation drives translocation of the growing polypeptide chain across the membrane
- the completed polypeptide chain is released within the ER lumen
proteins destined for incorporation into the plasma membrane or the membranes of other compartments are () instead of being released into the lumen
initially inserted into the ER membrane
Transmembrane proteins are inserted into the ER membrane by ()
⍺-helical membrane-spanning regions
transmembrane proteins are transported through the secretory pathway as () rather than soluble proteins
membrane components
the transmembrane sequences are (), thus allowing them to integrate with the phospholipid bilayer
hydrophobic
the lumen of the ER is topologically equivalent to the
exterior of the cell
Transmembrane regions signal a change in the translocon and lead to the ()
insertion of the polypeptide chain into the ER membrane
summarize how a transmembrane protein with a regular signal sequence is inserted into the ER membrane
- signal sequence allows SRP to target the translating ribosome to the ER membrane
- translation of a membrane-spanning ⍺-helix in the middle of a protein halts translocation
- helices exit the translocon laterally into the ER membrane and further translocation is blocked
- translation proceeds as normal from the cytosolic side of the ER
The orientation of transmembrane proteins is () as they travel along the secretory pathway
established in the ER and maintained
how are transmembrane proteins with no signal sequence targeted to the ER membrane
- SRP recognizes the internal transmembrane sequence present in the polypeptide chain and targets the translating ribosome to the SRP receptor
- there is no cleaving of a signal sequence
orientation of the transmembrane proteins is determined by the ()
amino acids immediately flanking the transmembrane region
polypeptide chains are inserted into the translocon with (+) charged residues on the
cytosolic side
a series of transmembrane sequences with altering orientations results in proteins that
span the membrane multiple times
how are transmembrane proteins with transmembrane sequences at the C-terminus brought to the ER membrane
- transmembrane domains of these proteins are recognized by TRC40
(distinct targeting factor) once translation is complete - TRC40 then escorts the proteins to the ER membrane, where it is inserted via the GET1-GET2 receptor
for secretory proteins, most protein processing occurs either during (1) or (2)
- translocation across the ER membrane
- within the ER lumen
the primary role of the lumenal ER proteins is to
assist in the folding and assembly of newly translocated polypeptides
polypeptides fold into their 3D conformations within the ER, assisted by ()
molecular chaperones that facilitate folding
the Hsp70 chaperone, (), is thought to bind to the unfolded polypeptide chain as it crosses the membrane and then mediate protein folding within the ER
BiP
the formation of disulfide bonds in the oxidizing environment of the ER is facilitated by the enzyme () located in the ER lumen
protein disulfide isomerase (PDI)
the process of glycosylating proteins on specific asparagine residues
N-linked glycosylation
give an overview of N-linked glycosylation
- an oligosaccharide (consisting of 14 sugars) is added to an acceptor asparagine residue on the growing polypeptide chain
- 3 glucose residues are removed while the protein is still within the ER
- the oligosaccharide is then transferred as a unit to acceptor asparagine
() synthesizes the oligosaccharide that is added to the growing polypeptide chain in N-linked glycosylation
dolichol
in N-linked glycosylation, the oligosaccharide is added to acceptor asparagine in the consensus sequence ()
Asn-(X)-Ser/Thr
glycosylation helps:
- prevent protein aggregration
- promote protein folding and sorting
glycolipids that anchor proteins to the plasma membrane and contain phosphatidylinositol
glycosylphosphatidylinositol (GPI) anchors
GPI anchors are assembled in the ()
ER membrane
how are GPI anchors added to a protein
- they are added immediately after completion of protein synthesis
- the C-terminal transmembrane domain is cleaved and exchanged for the GPI anchor
- these proteins are attached to the membrane only by their associated glycolipid
in (), misfolded proteins are identified, returned from the ER to the cytosol
ER-associated degradation (ERAD)
in ER-associated degradation (ERAD), misfolded proteins are degraded by the ()
ubiquitin-proteosome system
the () is activated when there is an excess of unfolded proteins accumulated in the ER
unfolded protein response pathway (UPR pathway)
in summary, activation of the UPR pathway leads to (1) and (2)
- expansion of the ER and production (transcription) of additional chaperones (and enzymes) to meet the need for increased protein folding
- transient reduction in the amount of newly synthesized proteins entering the ER
sustained activity of the UPR pathway leads to ()
cell death
specifically, the UPR results from the activation (by unfolded proteins) of the ff. signalling molecules (stress sensors) in the ER membrane
- IRE1
- ATF6
- PERK
what does IRE1 do to stimulate UPR
results in cleavage and activation of the mRNA encoding a transcription factor (XBP1), which stimulates the transcription of UPR target genes
what does ATF6 do to stimulate UPR
cleaved to release ATF6 transcription factor
what does PERK do to stimulate UPR
a protein kinase that phosphorylates the translation factor elF2, which inhibits general translation (reduces amount of protein entering ER); it also results in the preferential transcription factor ATF4 (contributes to induction of UPR target genes)
Membrane lipids are synthesized in the ()
smooth ER
membrane lipids are synthesized in association with () rather than in the aqueous environment of the cytosol
already existing cellular membranes
the membranes of eukaryotic cells are composed of 3 main types of lipids:
- phospholipids
- glycolipids
- cholesterol
basic structural components of the membrane; mostly derived from glycerol
phospholipids
overview of ER membrane lipid synthesis
- in the cytosolic side of the ER, membrane-bound enzymes transfer fatty acids from coenzyme A (fatty acyl CoA complexes) to glycerol-3-phosphate → results in phosphatidic acid
- the resulting phosphatidic acid is inserted into the membrane
- enzymes (phosphatase) in the cytosolic side of ER convert phosphatidic acid to diacylglycerol
- enzymes then catalyze the addition of different polar head groups to diacylglycerol; adding phosphocholine results in phosphatidylcholine
to ensure even growth of both the cytosolic and lumenal sides of the membrane, () catalyze the rapid translocation fo phospholipids across the ER membrane
flippases
The smooth ER is the site of (1) and (2)
- synthesis of steroid hormones
- degradation of many toxic compounds
the smooth ER also serves as the major site of synthesis of (1) and (2)
- cholesterol
- ceramide
overview of protein/lipid export from ER
- vesicles bud off from a specialized region of the ER called the ER exit side (ERES)
- vesicles then fuse to form the vesicles and tubules of the ER-Golgi intermediate compartment (ERGIC)
- from the ERGIC, the proteins are then carried to the Golgi
lumenal proteins targeted for the Golgi are bound by () that are selectively packaged into vesicles
transmembrane proteins
resident ER proteins destined to remain in ER lumen are marked by () located at their C-terminus; a recycling receptor in the ERGIC or Golgi recognizes these sequences and transports these proteins back to the ER
(Lys-Asp-Glu-Leu) KDEL retrieval sequences
KDEL retrieval sequences are recognized by
a recycling receptor in the ERGIC or Golgi
