Identification of Annexin A1 as a Novel Substrate for HAP-Mediated Ubiquitylation

E6-associated protein (E6AP) is a cellular ubiquitin protein ligase that mediates ubiquitylation and degradation of p53 in con. junction with the high-risk human papitlomavirus E6 proteins. However, the physiological functions of E6AP are poorly understood. To identify a novel biological function of E6AP, we screened for binding partners of E6AP using CST pull-down and mass spectrometry. Here we identified annexin A1, a member of the annexin superfamily, as an E6AP-binding protein. Ectopic expression of E6AP enhanced the degradation of annexin A1 in vivo. RNAi-mediated downregulation of endogenous E6AP increased the levels of endogenous annexin A1 protein. E6AP interacted with annexin At and induced its ubiquitylation in a Ca2+-dependent manner. CST pull-down assay revealed that the annexin repeat domain III of annexin A1 is important for the E6AP binding. Taken together, our data suggest that annexin A1 is a novel substrate for E6AP-mediated ubiquitylation. Our findings raise the possibility that E6AP may play a role in controlling the diverse functions of annexin A1 through the ubiquitin-proteasome pathway. J. Cell. Biochem. 106: 1123-1135, 2009. (c) 2009 Wiley-Liss, Inc.