Up-regulation of Tumor Susceptibility Gene 101 Protein in Ovarian Carcinomas Revealed by Proteomics Analyses

doi:10.1074/mcp.M600305-MCP200

Up-regulation of Tumor Susceptibility Gene 101 Protein in Ovarian Carcinomas Revealed by Proteomics Analyses*

From the ^‡Department of Pharmacology and Toxicology, Sealy Center for Cancer Cell Biology, School of Medicine, The University of Texas Medical Branch, Galveston, Texas 77555 and ^§Department of Pathology, The University of Texas M. D. Anderson Cancer Center, Houston, Texas 77030

‖To whom correspondence should be addressed: Dept. of Pharmacology and Toxicology, The University of Texas Medical Branch, 301 University Blvd., Galveston, TX 77555-1031. Tel.: 409-772-9656; Fax: 409-772-9642; E-mail: xcheng{at}utmb.edu

Abstract

Small GTPase RAS plays a critical role in cellular signaling and oncogenic transformation. Proteomics analysis of genetically defined human ovarian cancer models identified the tumor susceptibility gene 101 (TSG101) as a downstream target of RAS oncogene. Mechanistic studies revealed a novel post-translational regulation of TSG101 through the RAS/RAF/MEK/MAPK signaling pathway and downstream molecules p14^ARF/HDM2. Immunoanalysis using ovarian cancer samples and microtissue array revealed elevated TSG101 levels in human ovarian carcinomas. Silencing of TSG101 by short interfering RNA in ovarian cancer cells led to growth inhibition and cell death. Concurrent with the apparent growth-inhibitory effect, the levels of the CBP/p300-interacting transactivator with ED-rich tail 2 (CITED2) and hypoxia-inducible factor 1α (HIF-1α), as well as its cellular activity, were markedly reduced after TSG101 knockdown. These results demonstrate that TSG101 is important for CITED2- and HIF-1α-mediated cellular regulation in ovarian carcinomas.

Footnotes

Published, MCP Papers in Press, November 16, 2006, DOI 10.1074/mcp.M600305-MCP200
↵¹ The abbreviations used are: MEK, mitogen-activated protein kinase/extracellular signal-regulated kinase kinase; 2-DE, two-dimensional electrophoresis; CITED2, CBP/p300-interacting transactivator with ED-rich tail 2; GFP, green fluorescent protein; HMD2, human homolog of MDM2; HIF, hypoxia-inducible factor; MDM2, mouse double minute 2; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide; TSG101, tumor susceptibility gene 101; siRNA, short interfering RNA; MAPK, mitogen-activated protein kinase; CBP, cAMP-response element-binding protein (CREB)-binding protein; ERK, extracellular signal-regulated kinase; PI3K, phosphatidylinositol 3-kinase; Tricine, N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine; HRE, HIF-1α response element.
↵* This work was supported in part by National Institute of Health Grant GM066170 and institutional startup funds (to X. C.). Help from the core facilities was supported by NIEHS, National Institutes of Health Center Grant ES06676. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¶ Supported by American Cancer Society Research Scholar Grant RSG-04-028-01-CCE.
- Received August 10, 2006.
- Revision received October 30, 2006.