Wolfram syndrome 1 gene negatively regulates ER stress signaling in rodent and human cells
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
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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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 3

WFS1 enhances ATF6α ubiquitination and degradation.

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WFS1 enhances ATF6α ubiquitination and degradation. (A) IB analysis meas...
(A) IB analysis measuring ATF6α, WFS1, and actin levels in MIN6 cells stably expressing shGFP (control) or shWFS1 treated with 40 μM cycloheximide (CX) for 0, 2, and 4 hours (n = 3). (B) IB analysis measuring ATF6α, WFS1, and actin levels in INS1 832/13 cells expressing GFP (control) or WFS1 treated with 40 μM cycloheximide for 0, 2, and 6 hours (n = 3). (C) ATF6α was subjected to IP using an anti-ATF6α antibody from an INS1 832/13 cells inducibly expressing shWFS1 (treated for 48 hours with 2 μM doxycycline) and treated with MG132 (20 μM) for 3 hours. IPs were then subjected to IB with anti-ubiquitin and anti-ATF6α antibodies, and input lysates were blotted with anti-ATF6α, anti-WFS1, and anti-actin antibodies (n = 3). (D) ATF6α was subjected to IP using an anti-ATF6α antibody, from INS1 832/13 cells overexpressing GFP (control) or WFS1, then treated with MG132 (0.1 μM) overnight. IPs were subjected to IB with anti-ubiquitin and anti-ATF6α antibodies. Input lysates were subjected to IB with anti-ATF6α, anti-WFS1, and anti-actin antibodies (n = 3). (E) Wfs1–/– and WT littermate mouse pancreata were analyzed by immunohistochemistry using anti-ATF6α and anti-insulin antibodies. Scale bars: 50 μm.