title: Quantitative measurement of events in the mammalian unfolded protein response.
abstract: When the homeostasis of endoplasmic reticulum (ER) is disturbed by the accumulation of unfolded or misfolded proteins, a series of signaling responses collectively called the unfolded protein response (UPR) is triggered. UPR transducers IRE1, PERK, ATF6, and UPR-responsive genes such as GRP78/BiP, ERAD genes such as EDEM, and synthesis of the protein N-linked glycosylation donor lipid-linked oligosaccharides (LLOs) are mobilized. This chapter provides methods used in our laboratory to quantitatively measure the accumulation of mRNAs encoding BiP and EDEM, splicing of XBP1, cleavage of ATF6, inhibition of protein synthesis by PERK, and extension of LLOs under control and stress conditions.
authors.affiliation: Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas, Texas, USA.
authors.forename: Jie
authors.initials: J
authors.lastname: Shang
mesh_terms: D051702:Activating Transcription Factor 6 / Q000235:genetics / Q000378:metabolism; D000818:Animals; D015152:Blotting, Northern / Q000379:methods; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000235:genetics / Q000378:metabolism*; D000091342:Endoplasmic Reticulum Chaperone BiP; D006360:Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting / Q000379:methods; D008070:Lipopolysaccharides / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D000071701:Regulatory Factor X Transcription Factors; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D056811:Unfolded Protein Response; D000071717:X-Box Binding Protein 1
chemical_list: D051702:Activating Transcription Factor 6; D004268:DNA-Binding Proteins; C486654:EDEM1 protein, human; D000091342:Endoplasmic Reticulum Chaperone BiP; C000717247:HSPA5 protein, human; D006360:Heat-Shock Proteins; D008070:Lipopolysaccharides; D008565:Membrane Proteins; D000071701:Regulatory Factor X Transcription Factors; D014157:Transcription Factors; D000071717:X-Box Binding Protein 1; C000605543:XBP1 protein, human; C023023:lipid-linked oligosaccharides