The E6 Oncoproteins of High-Risk Papillomaviruses Bind to a Novel Putative GAP Protein, E6TP1, and Target It for Degradation

Abstract

The high-risk human papillomaviruses (HPVs) are associated with carcinomas of the cervix and other genital tumors. Previous studies have identified two viral oncoproteins, E6 and E7, which are expressed in the majority of HPV-associated carcinomas. The ability of high-risk HPV E6 protein to immortalize human mammary epithelial cells (MECs) has provided a single-gene model to study the mechanisms of E6-induced oncogenic transformation. In this system, the E6 protein targets the p53 tumor suppressor protein for degradation, and mutational analyses have shown that E6-induced degradation of p53 protein is required for MEC immortalization. However, the inability of most dominant-negative p53 mutants to induce efficient immortalization of MECs suggests the existence of additional targets of the HPV E6 oncoprotein. Using the yeast two-hybrid system, we have isolated a novel E6-binding protein. This polypeptide, designated E6TP1 (E6-targeted protein 1), exhibits high homology to GTPase-activating proteins for Rap, including SPA-1, tuberin, and Rap1GAP. The mRNA for E6TP1 is widely expressed in tissues and in vitro-cultured cell lines. The gene for E6TP1 localizes to chromosome 14q23.2-14q24.3 within a locus that has been shown to undergo loss of heterozygosity in malignant meningiomas. Importantly, E6TP1 is targeted for degradation by the high-risk but not the low-risk HPV E6 proteins both in vitro and in vivo. Furthermore, the immortalization-competent but not the immortalization-incompetent HPV16 E6 mutants target the E6TP1 protein for degradation. Our results identify a novel target for the E6 oncoprotein and provide a potential link between HPV E6 oncogenesis and alteration of a small G protein signaling pathway.

Regulation of the G2/M transition by p53

“…Expression of a truncated form of p53 that forms inactive tetramers with endogenous wildtype p53 shortened the G2 delay in IMR-90 ®broblasts, but not as e ectively as did HPV-E6 . HPV-E6 can bind to the ERC-55 calcium binding protein, the human homologue of the Drosophila tumor suppressor discs-large, and a putative GTPase-activating protein E6TP1 (Chen et al, 1995;Gao et al, 1999;Imai et al, 1997;Kiyono et al, 1997). The di erent observations made with HPV-E6 and the truncated form of p53 suggests that other targets of HPV-E6 may contribute to the G2 checkpoint.…”

Identification of a novel telomerase repressor that interacts with the human papillomavirus type-16 E6/E6-AP complex

“…In a screen of both fetal brain and HeLa cell cDNA libraries, several known E6/ E6-AP interactors were identified, including p53, HHR23A (human homolog of rad23), and a homolog of the E6TP1 gene (Unigene Hs. 406879; Scheffner et al 1993;Gao et al 1999;Kumar et al 1999). In addition, a new E6/E6-AP target protein identified in the screen was NFX1 (nuclear factor binds to the X1 box), a transcriptional repressor of MHC class II genes (Song et al 1994).…”

The E6AP Ubiquitin Ligase Is Required for Transactivation of the hTERT Promoter by the Human Papillomavirus E6 Oncoprotein

“…Although the best known function of HPV E6 is the targeting and ubiquitin-dependent degradation of the p53 tumor suppressor protein (5-7), E6 also binds to additional cellular proteins and has functions that are independent of p53 degradation (8). The growing list of E6 target proteins includes E6-BP (9), paxillin (10), hDIg (the human homologue of the Drosophila disc large tumor suppressor protein) (11,12), Mcm7 (minichromosome maintenance protein 7) (13,14), IRF-3 (IFN regulatory factor 3) (15), Myc (16,17), Bak (Bcl-2-homologous antagonist/killer) (18,19), E6TP-1 (E6-targeting protein 1) (20,21), CREB-binding protein/p300 (22,23), Tyk2 (protein-tyrosine kinase 2) (24), hScrib (the human homologue of the Drosophila Scribble (Vartul) tumor suppressor protein) (25), PKN (a novel protein kinase with a catalytic domain homologous to that of protein kinase C) (26), MUPP1 (multi-PDZ domain protein 1) (27), MAGI-1 (membrane-associated guanylate kinase protein) (28), Gps2 (G-protein pathway suppressor 2) (29), ADA3 (30,31), and tuberin (32).…”