title: BiP availability distinguishes states of homeostasis and stress in the endoplasmic reticulum of living cells.
abstract: Accumulation of misfolded secretory proteins causes cellular stress and induces the endoplasmic reticulum (ER) stress pathway, the unfolded protein response (UPR). Although the UPR has been extensively studied, little is known about the molecular changes that distinguish the homeostatic and stressed ER. The increase in levels of misfolded proteins and formation of complexes with chaperones during ER stress are predicted to further crowd the already crowded ER lumen. Surprisingly, using live cell fluorescence microscopy and an inert ER reporter, we find the crowdedness of stressed ER, treated acutely with tunicamycin or DTT, either is comparable to homeostasis or significantly decreases in multiple cell types. In contrast, photobleaching experiments revealed a GFP-tagged variant of the ER chaperone BiP rapidly undergoes a reversible quantitative decrease in diffusion as misfolded proteins accumulate. BiP mobility is sensitive to exceptionally low levels of misfolded protein stressors and can detect intermediate states of BiP availability. Decreased BiP availability temporally correlates with UPR markers, but restoration of BiP availability correlates less well. Thus, BiP availability represents a novel and powerful tool for reporting global secretory protein misfolding levels and investigating the molecular events of ER stress in single cells, independent of traditional UPR markers.
authors.affiliation: Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, NY 10461, USA.
authors.forename: Chun Wei
authors.initials: CW
authors.lastname: Lai
authors.forename: Deborah E
authors.initials: DE
authors.lastname: Aronson
authors.forename: Erik Lee
authors.initials: EL
authors.lastname: Snapp
mesh_terms: D000818:Animals; D002460:Cell Line; D002470:Cell Survival; D006224:Cricetinae; D004721:Endoplasmic Reticulum / Q000378:metabolism* / Q000473:pathology*; D000091342:Endoplasmic Reticulum Chaperone BiP; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000378:metabolism*; D006706:Homeostasis; D006801:Humans; D017510:Protein Folding; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013312:Stress, Physiological; D014162:Transfection; D014783:Viscosity
chemical_list: D000091342:Endoplasmic Reticulum Chaperone BiP; D006360:Heat-Shock Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins
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