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Methods for Studying ER Stress and UPR Markers in Human Cells

  • Many experimentally induced or disease-related cellular dysfunctions stress the endoplasmic reticulum, commonly resulting in an accumulation of unfolded proteins in the ER lumen which is sensed by three ER-resident transmembrane proteins, PERK, ATF6, and IRE1. Their activation by such ER stress affects the unfolded protein response, which consists of a shutoff of protein translation and at the same time the switching-on of specific transcription factors that control genes which function to reduce the burden of unfolded proteins to the ER. Here, we describe two sets of methods for monitoring the occurrence of ER stress and UPR signaling in human cells by analyzing markers of activation of all three ER stress sensor proteins. The first set of methods is based on the qualitative and quantitative analysis of UPR-induced transcripts by qPCR. The second set of methods consists of Western blot-based analysis of UPR-induced proteins or protein modifications. Their combined analysis allows assessment of activation of all three ER stress-activated signaling pathways that in combination are characteristic for the UPR.

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Metadaten
Document Type:Part of a Book
Language:English
Parent Title (English):Oslowski (Ed.): Stress responses: Methods and protocols. Methods in Molecular Biology (MIMB), Vol 1292
First Page:3
Last Page:18
ISBN:978-1-4939-2521-6
DOI:https://doi.org/10.1007/978-1-4939-2522-3_1
Pubmed Id:http://www.ncbi.nlm.nih.gov/pubmed?term=25804744
Publisher:Humana Press
Place of publication:New York, NY
Publication year:2015
Tag:ATF4; ATF6; IRE1; PERK; Western blot; XBP1; qPCR; real-time PCR
Departments, institutes and facilities:Fachbereich Angewandte Naturwissenschaften
Institut für funktionale Gen-Analytik (IfGA)
Dewey Decimal Classification (DDC):6 Technik, Medizin, angewandte Wissenschaften / 61 Medizin und Gesundheit / 610 Medizin und Gesundheit
Entry in this database:2015/04/02