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. 2020 Feb 24;27(8):2484–2499. doi: 10.1038/s41418-020-0517-0

Fig. 5. MARCH5 and MTCH2 control NOXA-driven MCL1 turnover in diverse cells.

Fig. 5

MARCH5 (a, b), MTCH2 (c, d) or UBE2K (e, f) were deleted from HCT116 cells by CRISPR-Cas9 gene targeting. Indel generation at sgRNA target sites was confirmed by DNA sequencing (Supplementary Table S2). Steady-state MCL1 expression (a, c, e) and the rate of MCL1 turnover (b, d, f) were examined in confirmed knockout clones. Cells were cultured with the protein synthesis inhibitor cycloheximide (50 μg/mL; CHX) for up to 4 h where indicated. MCL1 was markedly stabilized in clones lacking either MARCH5 or MTCH2 (a–d) and to a lesser degree in clones lacking UBE2K (e–f). MARCH5 (g) or MTCH2 (h) were deleted from HELA cells by CRISPR-Cas9 gene targeting. Indel generation at sgRNA target sites was again confirmed by DNA sequencing (Supplementary Table S2). MCL1 degradation caused by HA-NOXA overexpression was abolished in the knockout clones.