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. 2021 Feb 8;8(8):2002874. doi: 10.1002/advs.202002874

Figure 6.

Figure 6

OTUD1 activates the caspase‐dependent apoptotic pathway by recruiting the CUL4A‐DDB1 complex to facilitate ubiquitination‐mediated degradation of MCL1. A) IB to assess MCL1 ubiquitination in KYSE30 cells expressing EV, OTUD1, or OTUD1C320A. B) Quantitative analyses of MCL1 expression in KYSE30 cells expressing EV, OTUD1, or OTUD1C320A and subjected to a CHX pulse‐chase assay. Quantification of MCL1 expression relative to β‐actin expression is shown. The data are the means ± s.e.m.; n  =  3. C) IB to assess MCL1 ubiquitination in KYSE150 cells expressing shC, shD1#1, or shD1#2 shRNAs. D) Quantitative analyses of MCL1 expression in KYSE150 cells expressing shC, shD1#1, or shD1#2 shRNAs and subjected to a CHX pulse‐chase assay. Quantification of MCL1 expression relative to β‐actin expression is shown. The data are the means ± s.e.m.; n  =  3. E–G) IB for the indicated proteins in co‐IP assays. H) Relative viability of AIF‐ablated KYSE30 cells expressing EV, OTUD1, or OTUD1C320A and treated with DDP with or without Z‐VAD (50 × 10−6 m). The data are the means ± s.d.; n = 5. I) Cell death fraction of AIF‐ablated KYSE30 cells expressing EV, OTUD1, or OTUD1C320A and treated with or without Z‐VAD (50 × 10−6 m). The data are the means ± s.e.m.; n  =  3. Two‐tailed t tests, ***p < 0.001. J)IB for the indicated proteins in AIF‐ablated KYSE30 cells expressing EV, OTUD1, or OTUD1C320A or in K) AIF‐ablated KYSE150 cells expressing shC, shD1#1, or shD1#2 shRNAs and treated with or without DDP (5 µg mL−1)/Z‐VAD (50 × 10−6 m).