Figure 5.

ZNF313 is a novel E3 ubiquitin ligase for p21^WAF1. (a) Elevation of p21^WAF1 protein stability in ZNF313-depleted HCT116 cells. (b) Validation of the E3 ligase activity of ZNF313. A self-ubiquitination activity of ZNF313 was tested using wild-type, RING-MT and ubiquitin-interacting motif (UIM) deletion mutants. (c) Decreased ubiquitination of p21^WAF1 by ZNF313 depletion. (d) Loss of p21^WAF1-destabilizing activity of ZNF313 by RING mutation. (e) Comparison of p21^WAF1-destabilizing activity of WT- and RING-mutated ZNF313. (f) Loss of senescence-inhibiting activity of ZNF313 by RING mutation. CHX, cycloheximide; HA, hemagglutinin; IB, immunoblotting; RING, Really Interesting New Gene; SA, senescence-associated; Ub, ubiquitin; UIM, ubiquitin-interacting motif; WT, wild type