BIOC 212 Lecture Notes - Lecture 12: Phagocytosis, Secretion, Atf4
29 May 2018
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1- Intracellular Trafficking
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UPR Signaling Overview
• Unfolded protein response; stress response at ER membrane
o Reducing conditions
§ ER wants to be oxidizing to promote formation of disulfide bonds
§ Too much reducing agents into the cells triggers UPR
o Prevent glycosylation
o Release calcium from ER --the ER is high in calcium
o Viral infections, etc.
• Need to signal across the ER membrane, from the lumen out to the cytosol
o Then from cytosol to the nucleus to activate transcription
• There are 3 different pathways for signaling
o IRE1a, PERK & ATF6
• UPR is independent of HSR in the cytosol
IRE1 & XBP1
• IRE1 dimerizes/oligomerizes in response to unfolded proteins
o TM protein
o In the lumen, has domain that recognizes unfolded proteins
o In the cytosol, has 2 other parts; a kinase & an endonuclease
• In response to stress, the IRE1 monomer will form a dimer, and then an oligomer
o In the dimer & oligomer state, it becomes active
o The kinase will phosphorylate the other subunit in the dimer
§ Activates the ribonuclease
o The endoribonuclease will cut a specific mRNA that encodes the transcription
factor XBP1
§ Splices out an intron, and a ligase reattaches the mRNA together
§ Introduce frameshift mutation into mRNA since 26 bp intron that is
removed is not a multiple of 3
o New coding sequence is now translated efficiency
§ Before splicing, the protein sequence was not efficiently translated
(XBP1u)
§ New protein called XBP1s (spliced) is soluble, acts as TF in that moves
to nucleus and enhances transcription of chaperones, lipid synthesis
enzymes
§ Since one of the function of the UPR is to enlarge the ER so that can
handle more proteins
• IRE1 dimerizes/oligomerizes in response to unfolded proteins
• Autophosphorylation activates endoRNase activity
• XBP1u is translated at very low levels as membrane-associated protein
• IRE1 splices out 26 base intron, frameshift allows XBP1s to be translated
efficiently as soluble protein
• XBP1s is a transcription factor that upregulates components of ERAD, lipid
synthesis
• Bind to unfolded protein in lumen, causing dimerization and oligomerization which
gives the signal transmission across the membrane
IRE1 Activation
• Direct binding of unfolded protein may cause dimerization
o When have lots of unfolded proteins in the lumen, can get multiple monomers
of IRE1a binding to the same unfolded polypeptide
o Brings them together, form a dimer
• Inactive IRE1 may be bound by BiP (Hsp70) and may prevent dimerization
o Even though IRE1 is not itself unfolded, has an hydrophobic region that forms
a binding site for BiP
§ BiP binding to IRE1 partially covers up the dimerization site
o When have lots of unfolded proteins in the lumen, BiP will prefer to bind
unfolded proteins in the lumen, letting go of IRE1a and allowing it to activate
§ BiP binding to unfolded protein releases IRE1 to form dimers
• Similar to what happens with HSF1 & Hsp90
o HSF1 monomer bound to Hsp90, stress = Hsp90 binds unfolded proteins =
HSF1 trimerizes, activates and move to nucleus to activate transcription of
Hsp90 and other chaperones
• Combination of these two mechanisms is what activates IRE1a
• Oligomers of IRE1 are fully active
o Have clusters of activated IRE1a that are cutting the XBP1 mRNA
RIDD
• Ribonuclease domain of IRE1a works on more than just the XBP1 RNA
o Degrade a certain set of mRNAs in the cytosol
o Regulated IRE1-Dependent mRNA Decay (RIDD)
• IRE1 RNase cuts certain other mRNAs to cause their degradation
o When activate IRE1, activate ribonuclease that will degrade certain mRNAs,
meaning that proteins that are encoded will not be translated
• Decreases translation of proteins entering the ER
o Less proteins entering the ER, such that ER can handle the stress better
• Stops cell growth, until recovery from stress
o Some mRNAs that are degraded promote cell growth
o ER stress affects everything downstream in secretory pathway
Document Summary
Upr signaling overview: unfolded protein response; stress response at er membrane, reducing conditions. Er wants to be oxidizing to promote formation of disulfide bonds. Ire1a, perk & atf6: upr is independent of hsr in the cytosol. Ire1 dimerizes/oligomerizes in response to unfolded proteins: tm protein. In the lumen, has domain that recognizes unfolded proteins. In the cytosol, has 2 other parts; a kinase & an endonuclease. In response to stress, the ire1 monomer will form a dimer, and then an oligomer. In the dimer & oligomer state, it becomes active: the kinase will phosphorylate the other subunit in the dimer. Activates the ribonuclease: the endoribonuclease will cut a specific mrna that encodes the transcription factor xbp1. Splices out an intron, and a ligase reattaches the mrna together. Introduce frameshift mutation into mrna since 26 bp intron that is removed is not a multiple of 3: new coding sequence is now translated efficiency.
