Fig. 4: DCAF15 destabilizes cohesin-bound PDS5A and CDCA5.

a Lysates from HEK293T cells stably expressing HA-tagged DCAF15 with the indicated siRNA and/or FLAG-tagged cDNAs, treated with 5 μM MLN4924 for 6 h, were subjected to FLAG-immunoprecipitation and Western blot analysis for the indicated proteins. Immunoblots are representative of two independent experiments. b Lysates from Cas9+ HEL cells infected with lentiviruses encoding ROSA26- or DCAF15-targeting sgRNAs were subjected to immunoprecipitation with the indicated amounts of α-SMC1A antibody and Western blot analysis for the indicated proteins. Immunoblots are representative of three independent experiments. c Same as in (b), except that cells were treated with 5 µg/mL cycloheximide (CHX) for indicated times. Immunoblots are representative of two independent experiments. d Same as in (b), except that cells were treated with 10 µM MG132 (proteasome inhibitor, MG), 5 µM MLN4924 (MLN), or 3 µM indisulam (Ind) for indicated times. Immunoblots are representative of three independent experiments. e Lysates from HEK293T cells transiently transfected with FLAG-DCAF15, HA-SMC1A, HA-PDS5A, and HA-CDCA5 were subjected to FLAG-immunoprecipitation. Following, FLAG-immunoprecipitants were subjected to in vitro ubiquitination assay and Western blot analysis for the indicated proteins. Immunoblots are representative of two independent experiments. f Model of CRL4-DCAF15 binding the head domain of SMC1A and promoting the ubiquitination of PDS5A when bound to the SMC1A-SMC3-RAD21 tripartite ring. Source data are provided as a Source Data file.