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. 2010 Feb 15;120(3):744–755. doi: 10.1172/JCI39678

Figure 3. WFS1 enhances ATF6α ubiquitination and degradation.

Figure 3

(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.