Figure 7: CYLD deubiquitinates p53 and induces its stabilization.
From: The tumour suppressor CYLD regulates the p53 DNA damage response
HEK-293 T (a,d) and HCT116 (b) cells were transfected with the indicated plasmids and were subsequently treated with MG-132/CpT. p53 was immunoprecipitated followed by immunoblot with anti-p53(DO-1), anti-ubiquitin and anti-CYLD antibodies as indicated. Expression levels of p53 and CYLD proteins are shown in immunoprecipitates and WCE. (c) HCT116 were transfected with HA-CYLD or an empty vector and treated with CpT for the indicated time points followed by immunoblot with anti-p53, HA and tubulin antibodies. (e,f) HEK-293 T cells were transfected with the indicated constructs and treated with MG-132/CpT. p53 was immunoprecipitated and its levels and type of ubiquitination were analysed by immunoblot using the indicated antibodies. (g) Ubiquitinated Flag-p53 (Flag-p53(Ub)n) was immunoprecipitated from MG-132/CpT-treated HEK-293 T cells and subjected to an in vitro deubiquitination assay using the indicated DUBs. Total and K48-linked ubiquitin chains on p53 are shown by immunoblot using specific antibodies.
