The Unfolded Protein Response: Methods and Protocols

Part I Determination of unfolded protein levels

1 Measuring cysteine exposure in unfolded proteins with tetraphenylethene maleimide and its analogues

Shouxiang Zhang and Yuning Hong

Heinrich Kroukamp, Kai Peng, Ian T. Paulsen, and Riaan den Haan

3 Indirect methods to measure unfolded proteins in living cells using fluorescent proteins

Elisabet Navarro-Tapia, and Roberto Pérez-Torrado

4 A platform technology for monitoring the Unfolded Protein Response

Bhagyashree Bachhav, Carlos A. Origel Marmolejo, Yafet Arefeayne, and Laura Segatori

5 The use of fluorescent protein fusions to monitor the unfolded protein response and protein foldase-substrate interactions in plant protoplasts

Rina Carrillo, Elizabeth Feldeverd, and David A. Christopher

Part II Methods to study UPR signal transmission

6 High-throughput analysis of protein turnover with tandem fluorescent protein timers

Jia Jun Fung, Karla Blöcher-Juárez, and Anton Khmelinskii

7 Detection of HAC1 mRNA Splicing by RT-PCR in Saccharomyces cerevisiae

Jagadeesh Kumar Uppala and Madhusudan Dey

Part III Analysing the outcomes of the UPR pathway activation

8 Determination of protein translation during the activation of the UPR

Weihan Li and Robert H. Singer

9 Determination of the stability and intracellular (intra-nuclear) targeting and recruitment of pre-HAC1 mRNA in the Saccharomyces cerevisiae during the activation of UPR

Sunirmal Paira and Biswadip Das

10 Assays to study IRE1 activation and signaling

Paloma Moraga, Raul Aravena, Hery Urra, and Claudio Hetz

11 Analysis of XBP1 contribution to hyperosmolarity-induced lipid synthesis.

Cecilia I. Casali, Leandro Parra, Luciana C. Erjavec, and María del Carmen Fernández Tome

12 Integrating an ER stress reporter for monitoring genome wide UPR-ER in budding yeast

Shuvadeep Maity, Asmita Ghosh, and Kausik Chakraborty

Part IV UPR studies in mammalian models

13 Analyze mouse knockout models of UPR pathway elements

Chao Zheng, Cheng Wang, Qiang Jie, and Liu Yang

14 In vitro stimulation of IRE1a/XBP1-deficient B cells with LPS

Huabin Zhu, Chen Jiang, Randal J. Kaufman, Honglin Li, and Nagendra Singh

15 Detection of PERK signaling in the central nervous system

Zhixin Lei, Sarrabeth Stone, and Wensheng Lin

Part V UPR in alternative models

16 Methods to study the mitochondrial unfolded protein response (UPRmt) in Caenorhabditis elegans

17 Drosophila Unfolded Protein Response (UPR) assays in vitro and in vivo

Hidetaka Kato, Deepika Vasudevan, and Hyung Don Ryoo

18 Protein preparation for proteomic analysis of the unfolded protein response in Arabidopsis thaliana

Yunting Pu and Federica Brandizzi

Part VI UPR and disease

19 Structure-based drug discovery of IRE1 modulators

Diana Pelizzari-Raymundo, Leif A. Eriksson, Eric Chevet, and Xavier Guillory

20 Chemical screening to detect ER stress regulators

Mehrnoosh Ghafouri, Chester B. Gauss, and Andrew M. Fribley