Abstract 290: PDZ protein PDZK1 associates with PTEN and inhibits PTEN phosphorylation

Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DC The phosphatase and tensin homolog deleted on chromosome 10 (PTEN) tumour suppressor is a lipid and protein phosphatase that inhibits phosphoinositide 3-kinase (PI3K)-dependent signaling by dephosphorylating phosphatidylinositol 3,4,5-trisphosphate (PtdInsP(3)). The carboxyl terminus (CT) of PTEN and PTEN post-translational modification by phosphorylation were reported to regulate PTEN signaling. However, the modes of regulation of PTEN signaling are not completely understood. The CT of PTEN has a typical PDZ domain-binding motif that interacts with several PDZ domain-containing proteins, which regulates PTEN signaling and function. To screen novel PTEN binding proteins, the PTEN CT was used to overlay against the lysates of rabbit tissue, including kidney, heart and brain. The molecular weight of positive band was similar with that of PDZ-domain containing 1 (PDZK1), a scaffolding protein. We further investigated this PTEN-associated protein by using GST pull-down in combination with mass spectrometry-based proteomics assays. PDZK1 was identified as one of the binding partners of PTEN. PTEN associated with PDZK1 via its PDZ domain 2 and 3, with binding affinity of 20 nM and 25 nM respectively, and the last four amino acids of the CT of PTEN were the key determinants of this interaction. Mutation of any one of the four amino acids to alanine abolished the association of PTEN with PDZK1-PDZ2, PDZK1-PDZ3 or PDZK1 full length, indicating that the PDZK1-PTEN association is stringent. Moreover, full-length PTEN was robustly associated with PDZK1, as determined by co-immunoprecipitation experiments in transiently transfected COS-7 cells, and mutation of the last amino acid valine to alanine also led to the abolishment of the interaction between PTEN full length and PDZK1. Immunofluorescence results showed that PTEN full length and PDZK1 can colocalize at cytoplasm of BHK cells. By interacting with PTEN, PDZK1 could inhibit the phosphorylation of PTEN and Akt activation induced by the stimulation of insulin in a dose-dependent manner, indicating PDZK1 can enhance the PTEN protein activity via their interaction. Taken together, these are the first results that present a interaction between PDZK1 and PTEN. PDZK1 interacts with PTEN to inhibit PTEN phosphorylation, leading to an increase in PTEN activity at inhibiting Akt activation. Our results suggest a regulatory role of PDZ domain binding on PTEN function by controlling its phosphorylation status. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 290.