E3 ligase EDD1/UBR5 is utilized by the HPV E6 oncogene to destabilize tumor suppressor TIP60

Abstract

Tat-interacting protein of 60 kDa (TIP60) is an essential lysine acetyltransferase implicated in transcription, DNA damage response and apoptosis. TIP60 protein expression is reduced in cancers. In cervical cancers, human papillomavirus (HPV) E6 oncogene targets cellular p53, Bak and some of the PDZ domain-containing proteins for proteasome-mediated degradation through E6AP ligase. Recently, E6 oncogene from high-risk and low-risk categories was also shown to target TIP60. However, the molecular mechanisms and whether destabilization of TIP60 contributes to HPV E6-mediated transformation remain unanswered. Our proteomic analyses revealed EDD1 (E3 identified by differential display), an E3 ligase generally overexpressed in cancers as a novel interacting partner of TIP60. By investigating protein turnover and ubiquitination assays, we show that EDD1 negatively regulates TIP60's stability through the proteasome pathway. Strikingly, HPV E6 uses this function of EDD1 to destabilize TIP60. Colony-formation assays and soft agar assays show that gain of function of TIP60 or depletion of EDD1 in HPV-positive cervical cancer cells significantly inhibits cell growth in vitro. This phenotype is strongly supported by the in-vivo studies where re-activation of TIP60 in cervical cancer cells dramatically reduces tumor formation. In summary, we have discovered a novel ligase through which E6 destabilizes TIP60. Currently, in the absence of an effective therapeutic vaccine for malignant cervical cancers, cervical cancer still remains to be a major disease burden. Hence, our studies implying a distinct tumor suppressor role for TIP60 in cervical cancers show that reactivation of TIP60 could be of therapeutic value.

Angelman syndrome–associated point mutations in the Zn2+-binding N-terminal (AZUL) domain of UBE3A ubiquitin ligase inhibit binding to the proteasome

“…The 26S proteasome is one of the major cellular complexes interacting with UBE3A (21)(22)(23)(24)(25)(26)(27)(28)(29). Although immunoprecipitation and MS cannot distinguish which specific protein subunit within the 26S proteasome directly binds UBE3A, PSMD4 was the only proteasomal subunit identified as a direct UBE3Abinding partner in a yeast two-hybrid screen we recently reported, suggesting that PSMD4 likely bridges the interaction of UBE3A with the 26S proteasome (24).…”

“…Although the proteins of the lid complex are responsible for the recruitment of ubiquitylated proteins, the barrel and its intrinsic proteolytic activities mediate the breakdown of proteins (20). UBE3A associates with the proteasome (21)(22)(23)(24)(25)(26)(27)(28)(29). In that context, the UBE3A-interacting protein PMSD4 (also known as Rpn10/S5a) is part of the regulatory lid complex and known to bind to polyubiquitin chains of ubiquitylated proteins via its ubiquitin-interacting motif (UIM), resulting in their recruitment to the proteasome (30 -32).…”

GRWD1 regulates ribosomal protein L23 levels via the ubiquitin-proteasome system

“…EDD was among the E3 ubiquitin ligases identified by MS analysis of GRWD1-interacting proteins (Tables S1 and S2). EDD belongs to the HECT family of E3 ubiquitin ligases (Rotin and Kumar, 2009) and regulates the stability of various proteins such as topoisomerase II β-binding protein 1 (TopBP1) and Tat-interactive protein 60 (TIP60) (Honda et al, 2002;Subbaiah et al, 2016). The interaction between EDD and GRWD1 was confirmed by co-IP and immunoblotting (Fig.…”

HPV Oncoproteins and the Ubiquitin Proteasome System: A Signature of Malignancy?

“…Thus fluctuations of EDD protein levels might directly affect the viral life cycle, and influence the development of HPV-induced malignancies [94]. In addition, it has also been shown that the association of EDD with HR HPV E6 is important in destabilizing TIP60; a histone acetyltransferase tumor suppressor, thus ultimately contributing to the development of HPV-induced malignancies [95].…”