Uncovering changes in the redox protein interactome of PTEN

Corinne M. Spickett, Ivan Verrastro, Andrew R. Pitt

Research output: Contribution to journalConference abstract

Abstract

Phosphatase and tensin homolog (PTEN) is a redox-sensitive, dual-specificity protein phosphatase involved in regulating a number of cellular processes including metabolism, apoptosis, cell proliferation and survival. It acts as a tumor suppressor by negatively regulating the PI3K/Akt pathway. While direct evidence of a redox regulation of PTEN downstream signaling has been reported, the effect of cellular oxidative stress or direct PTEN oxidation on the PTEN interactome is still poorly defined. To investigate this, PTEN-GST fusion protein was prepared in its reduced form and an H2O2-oxidized form that was reversible by DTT treatment, and these were immobilized on a glutathione-sepharose-based support. The immobilized protein was incubated with cell lysate to capture interacting proteins. Captured proteins were eluted from the beads, analyzed by LC-MSMS and comparatively quantified using label-free methods. After subtraction of interactors that were also present in the resin and GST controls, 97 individual protein interactors were identified, including several that are novel. Fourteen interactors that varied significantly with the redox status of PTEN were identified, including thioredoxin and peroxiredoxin-1. Except for one interactor, their binding was higher for oxidized PTEN. Using western blotting, altered binding to PTEN was confirmed for 3 selected interactors (Prdx1, Trx, and Anxa2) and DDB1 was validated as a novel interactor with unaltered binding. Our results suggest that the redox status of PTEN causes a functional variation in the PTEN interactome which is important for the cellular function of PTEN. The resin capture method developed had distinct advantages in that the redox status of PTEN could be directly controlled and measured.
Original languageEnglish
Article numberOP15
Pages (from-to)S9
Number of pages1
JournalFree Radical Biology and Medicine
Volume86
Issue numberSuppl.1
Early online date28 Aug 2015
DOIs
Publication statusPublished - Sep 2015
EventSFRR-E/SNFS Meeting Stuttgart 2015 - University of Hohenheim, Stuttgart, Germany
Duration: 1 Sep 2015 → …

Bibliographical note

SFRR-E/SNFS Conference Abstracts, Stuttgart 2015

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