PDF file, 739K, Fig. S1. SUMOylation of Akt. Fig.S2: Association of Akt with PIAS1 and SENP1. Fig. S3 Identification of SUMOylation site in Akt. Fig. S4 K168 and K183 is important to Akt SUMOylation. Fig. S5 SUMOylation affects the phosphorylation of FOXO proteins. Fig. S6. Mutation of K276 SUMOylation site does not affect its dimerization, association with substrates, and ubiquitination. Fig. S7. SUMOylation has little effect on Akt Subcellular Localization.
ARTICLE ABSTRACTProto-oncogene Akt plays essential roles in cell proliferation and tumorigenesis. Full activation of Akt is regulated by phosphorylation, ubiquitination, and acetylation. Here we report that SUMOylation of Akt is a novel mechanism for its activation. Systematically analyzing the role of lysine residues in Akt activation revealed that K276, which is located in a SUMOylation consensus motif, is essential for Akt activation. Ectopic or endogenous Akt1 could be modified by SUMOylation. RNA interference–mediated silencing of UBC9 reduced Akt SUMOylation, which was promoted by SUMO E3 ligase PIAS1 and reversed by the SUMO-specific protease SENP1. Although multiple sites on Akt could be SUMOylated, K276 was identified as a major SUMO acceptor site. K276R or E278A mutation reduced SUMOylation of Akt but had little effect on its ubiquitination. Strikingly, these mutations also completely abolished Akt kinase activity. In support of these results, we found that expression of PIAS1 and SUMO1 increased Akt activity, whereas expression of SENP1 reduced Akt1 activity. Interestingly, the cancer-derived mutant E17K in Akt1 that occurs in various cancers was more efficiently SUMOylated than wild-type Akt. Moreover, SUMOylation loss dramatically reduced Akt1 E17K–mediated cell proliferation, cell migration, and tumorigenesis. Collectively, our findings establish that Akt SUMOylation provides a novel regulatory mechanism for activating Akt function. Cancer Res; 73(18); 5742–53. ©2013 AACR.