title: Reversible oxidation of phosphatase and tensin homolog (PTEN) alters its interactions with signaling and regulatory proteins.
abstract: Phosphatase and tensin homolog (PTEN) is involved in a number of different cellular processes including metabolism, apoptosis, cell proliferation and survival. It is a redox-sensitive dual-specificity protein phosphatase that acts as a tumor suppressor by negatively regulating the PI3K/Akt pathway. While direct evidence of redox regulation of PTEN downstream signaling has been reported, the effect of PTEN redox status on its protein-protein interactions is poorly understood. PTEN-GST in its reduced and a DTT-reversible H2O2-oxidized form was immobilized on a glutathione-sepharose support and incubated with cell lysate to capture interacting proteins. Captured proteins were analyzed by LC-MSMS and comparatively quantified using label-free methods. 97 Potential protein interactors were identified, including a significant number that are novel. The abundance of fourteen interactors was found to vary significantly with the redox status of PTEN. Altered binding to PTEN was confirmed by affinity pull-down and Western blotting for Prdx1, Trx, and Anxa2, while DDB1 was validated as a novel interactor with unaltered binding. These results suggest that the redox status of PTEN causes a functional variation in the PTEN interactome. The resin capture method developed had distinct advantages in that the redox status of PTEN could be directly controlled and measured.
authors.affiliation: School of Life and Health Sciences, Aston Triangle, Aston University, Birmingham B4 7ET, UK.
authors.forename: Ivan
authors.initials: I
authors.lastname: Verrastro
authors.affiliation: School of Life and Health Sciences, Aston Triangle, Aston University, Birmingham B4 7ET, UK.
authors.forename: Karina
authors.initials: K
authors.lastname: Tveen-Jensen
authors.affiliation: Department of Chemistry and Institute of Chemical Biology, Imperial College London, UK.
authors.forename: Rudiger
authors.initials: R
authors.lastname: Woscholski
authors.affiliation: School of Life and Health Sciences, Aston Triangle, Aston University, Birmingham B4 7ET, UK.
authors.forename: Corinne M
authors.initials: CM
authors.lastname: Spickett
authors.affiliation: School of Life and Health Sciences, Aston Triangle, Aston University, Birmingham B4 7ET, UK. Electronic address: a.r.pitt@aston.ac.uk.
authors.forename: Andrew R
authors.initials: AR
authors.lastname: Pitt
mesh_terms: D004268:DNA-Binding Proteins / Q000378:metabolism; D004220:Disulfides / Q000737:chemistry; D005978:Glutathione / Q000378:metabolism; D045325:HCT116 Cells; D006801:Humans; D010084:Oxidation-Reduction; D051059:PTEN Phosphohydrolase / Q000502:physiology*; D054464:Peroxiredoxins / Q000378:metabolism; D040901:Proteomics; D015398:Signal Transduction; D013879:Thioredoxins / Q000378:metabolism
chemical_list: C091435:DDB1 protein, human; D004268:DNA-Binding Proteins; D004220:Disulfides; D013879:Thioredoxins; C515940:PRDX1 protein, human; D054464:Peroxiredoxins; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human; D005978:Glutathione
keywords: Mass spectrometry; Peroxiredoxin-1; Protein oxidation; Protein–protein interactions; Proteomics; Redox biology; Signaling; Thioredoxin-1