Wolfram syndrome 1 gene negatively regulates ER stress signaling in rodent and human cells
Sonya G. Fonseca, … , M. Alan Permutt, Fumihiko Urano
Sonya G. Fonseca, … , M. Alan Permutt, Fumihiko Urano
Published March 1, 2010; First published February 15, 2010
Citation Information: J Clin Invest. 2010;120(3):744-755. https://doi.org/10.1172/JCI39678.
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Research Article Metabolism

Wolfram syndrome 1 gene negatively regulates ER stress signaling in rodent and human cells

Abstract

Wolfram syndrome is an autosomal-recessive disorder characterized by insulin-dependent diabetes mellitus, caused by nonautoimmune loss of β cells, and neurological dysfunctions. We have previously shown that mutations in the Wolfram syndrome 1 (WFS1) gene cause Wolfram syndrome and that WFS1 has a protective function against ER stress. However, it remained to be determined how WFS1 mitigates ER stress. Here we have shown in rodent and human cell lines that WFS1 negatively regulates a key transcription factor involved in ER stress signaling, activating transcription factor 6α (ATF6α), through the ubiquitin-proteasome pathway. WFS1 suppressed expression of ATF6α target genes and repressed ATF6α-mediated activation of the ER stress response element (ERSE) promoter. Moreover, WFS1 stabilized the E3 ubiquitin ligase HRD1, brought ATF6α to the proteasome, and enhanced its ubiquitination and proteasome-mediated degradation, leading to suppression of ER stress signaling. Consistent with these data, β cells from WFS1-deficient mice and lymphocytes from patients with Wolfram syndrome exhibited dysregulated ER stress signaling through upregulation of ATF6α and downregulation of HRD1. These results reveal a role for WFS1 in the negative regulation of ER stress signaling and in the pathogenesis of diseases involving chronic, unresolvable ER stress, such as pancreatic β cell death in diabetes.

Authors

Sonya G. Fonseca, Shinsuke Ishigaki, Christine M. Oslowski, Simin Lu, Kathryn L. Lipson, Rajarshi Ghosh, Emiko Hayashi, Hisamitsu Ishihara, Yoshitomo Oka, M. Alan Permutt, Fumihiko Urano

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Figure 5

WFS1 interacts with and stabilizes the E3 ligase HRD1.

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WFS1 interacts with and stabilizes the E3 ligase HRD1. (A) Hrd1 was subj...
(A) Hrd1 was subjected to IP from INS1 832/13 cells, and IPs were subjected to IB analysis using anti-WFS1 and anti-Hrd1 antibodies (n = 3). (B) Total lysates from INS1 832/13 cells inducibly expressing shWFS1 (treated with 2 μM doxycycline for 48 hours) were analyzed by IB using anti-WFS1, anti-Hrd1, and anti-actin antibodies (n = 3). (C) IB analysis measuring HRD1 levels in MIN6 cells stably expressing shGFP (control) or shWFS1 treated with 40 μM cycloheximide for 0, 0.5, 1, and 2 hours (n = 3). (D) Wfs1–/– and WT littermate mouse pancreata were analyzed by immunohistochemistry using anti-HRD1 and anti-insulin antibodies (n = 3). Scale bars: 100 μm. (E) COS7 cells were transfected with pcDNA3, HRD1–c-Myc, HRD1–c-Myc and WT WFS1, or HRD1–c-Myc and WFS1 mutants (P724L, G695V, and ins483fs/ter544) expression plasmids. Expression levels of HRD1–c-Myc, WFS1, and actin were measured by IB using anti–c-Myc, anti-WFS1, and anti-actin antibodies, respectively. WT and mutant WFS1 are denoted by single and double asterisks, respectively. (F) COS7 cells were transfected with pcDNA3, HRD1–c-Myc, HRD1–c-Myc and WT WFS1-Flag, HRD1–c-Myc and WFS P724L-Flag, and HRD1–c-Myc and WFS1 G695V-Flag expression plasmids. The lysates were subjected to IP with anti-Flag antibody and IB with anti–c-Myc antibody to study the interaction between HRD1 and WFS1.