USP28 Is a Component of the p53-53BP1 Axis
(A) N3 resistance of indicated cell lines assessed as in Figures 1A and 1B.
(B) Quantification of (A). Mean ± SD (n = 3).
(C) N3 resistance of a 53BP1Δ, USP28Δ double-knockout cell line relative to single mutants. Mean ± SD (n = 3, in triplicate).
(D) Lysates prepared from indicated control and N3-treated cell lines were immunoblotted with indicated antibodies.
(E) The transactivation of p53-responsive genes MDM2 and TP53I3 was assessed by qRT-PCR in indicated cell lines. Indicated fold changes were calculated upon normalization against HPRT1 transcripts. Data are representative of two independent experiments; mean ± SD.
(F) Schematic representation of the USP28 protein domain architecture and the domain and point mutants used in this study. UBA, ubiquitin-associated domain; UIM, ubiquitin-interaction motif; UCH, ubiquitin carboxyl-terminal hydrolase domain.
(G) Visual and quantitative analysis of N3 resistance of USP28Δ cell lines stably transduced with indicated USP28 expressing lentiviruses. N3 resistance relative to a control (GST)-transduced USP28Δ cell line was calculated as in (A) and (B). Mean ± SD (n = 3, in triplicate). See also Figure S6.