Jayantha GUNARATNE1, Mei Xian GOH2, Lee Foon SWA1, Fen Yee LEE2, Emma SANFORD2, Loke Meng WONG2, Kelly A. HOGUE1, Walter P. BLACKSTOCK1, and Koichi OKUMURA2.
1 - Institute of Molecular and Cell Biology, 61 Biopolis Drive, Singapore 138673.
2 - Cancer Science Institute of Singapore, National University of Singapore, 28 Medical Drive, Singapore 117456.
Published in J Biol Chem. 17 March 2011.
The tumor suppressor PTEN (Phosphatase and Tensin homologue) negatively regulates the phosphatidyl-inositol-3-kinase (PI3K) pathway through its lipid phosphatase activity and is one of the most commonly lost tumor suppressors in human cancers. Though the tumor suppressive function involves PTENís lipid phosphatase-dependent and Ėindependent activities, the mechanism leading to the phosphatase-independent function of PTEN is poorly understood. Some PTEN mutants have lipid phosphatase activity but fail to suppress cell growth. Here we use a cancer associated mutant, G20E, to gain insight into the phosphatase-independent function of PTEN by investigating protein-protein interactions using mass spectrometry (MS)-based stable isotope labeling by amino acids in cell culture (SILAC). A strategy named parallel affinity purification (PAP) and SILAC has been developed to prioritize interactors and to compare the interactions between wild-type and G20E PTEN. Clustering of the prioritized interactors acquired by the PAP-SILAC approach shows three distinct clusters: 1) wild-type specific interactors, 2) interactors unique to the G20E mutant and 3) proteins common to wild-type and mutant. These interactors are mainly involved in cell migration and apoptosis pathways. We further demonstrate that the wild-type specific interactor, NUDTL16L1, is required for the regulatory function of wild-type PTEN in cell migration. These finding contribute to a better understanding of the mechanisms of PTENís phosphatase-dependent and -independent functions.