Quantitative measurement of events in the mammalian unfolded protein response.
The accumulation of unfolded or misfolded proteins in the endoplasmic reticulum (ER) triggers the unfolded protein response. IRE1, PERK, ATF6, BiP, EDEM, lipid-linked oligosaccharides (LLOs), and XBP1 directly or indirectly participate in this process. This article provides methods used in our laboratory to quantitatively measure the accumulation of mRNAs encoding BiP and EDEM; splicing of XBP-1; cleavage of ATF6; inhibition of protein synthesis by PERK; and extension of LLOs under control and stress conditions.
Shang
Jie
J
Department of Pharmacology, University of Texas Southwestern Medical Center, 5323 Harry Hines Blvd., Dallas, TX 75390-9041, USA. jie.shang@utsouthwestern.edu
D051702:Activating Transcription Factor 6; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern / Q000379:methods; D018929:Cell Culture Techniques; D004268:DNA-Binding Proteins / Q000096:biosynthesis; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D000091342:Endoplasmic Reticulum Chaperone BiP; D006360:Heat-Shock Proteins / Q000096:biosynthesis; D006801:Humans; D015151:Immunoblotting / Q000379:methods; D008565:Membrane Proteins / Q000096:biosynthesis / Q000235:genetics; D051379:Mice; D018832:Molecular Chaperones / Q000096:biosynthesis; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000096:biosynthesis; D017510:Protein Folding; D000071701:Regulatory Factor X Transcription Factors; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D015398:Signal Transduction; D014157:Transcription Factors / Q000096:biosynthesis; D000071717:X-Box Binding Protein 1
C497111:ATF6 protein, human; D051702:Activating Transcription Factor 6; C497112:Atf6 protein, mouse; D004268:DNA-Binding Proteins; D000091342:Endoplasmic Reticulum Chaperone BiP; D006360:Heat-Shock Proteins; D008565:Membrane Proteins; D018832:Molecular Chaperones; D009687:Nuclear Proteins; D000071701:Regulatory Factor X Transcription Factors; D014157:Transcription Factors; D000071717:X-Box Binding Protein 1; C000605543:XBP1 protein, human; C000605544:Xbp1 protein, mouse