Oxidative stress induces protein phosphatase 2A-dependent dephosphorylation of the pocket proteins pRb, p107, and p130

被引:98
作者
Cicchillitti, L
Fasanaro, P
Biglioli, P
Capogrossi, MC
Martelli, F
机构
[1] IRCCS, Ist Dermopat Immacolata, Lab Patol Vasc, I-00167 Rome, Italy
[2] IRCCS, Ist Cardiol Fdn I Monzino, Lab Biol Vasc & Terapia Genica, Milan, Italy
[3] IRCCS, Ist Cardiol Fdn I Monzino, Dipartimento Chirurg Cardiovasc, Milan, Italy
关键词
D O I
10.1074/jbc.M300511200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Oxidative stress induces cell death and growth arrest. In this study, the regulation and the functional role of the retinoblastoma family proteins pRb, p107, and p130 in the cellular response to oxidative stress were investigated. Treatment of endothelial cells with H2O2 induced rapid hypophosphorylation of the retinoblastoma family proteins. This event did not require p53 or p21(Waf1/Cip1/Sdi1) and was not associated with cyclin/cyclin-dependent kinase down-modulation. Four lines of evidence indicate that H2O2-induced hypophosphorylation of pRb, p107, and p130 was because of the activity of protein phosphatase 2A (PP2A). First, cell treatment with two phosphatase inhibitors, okadaic acid and calyculin A, prevented the hypophosphorylation of the retinoblastoma family proteins, at concentrations that specifically inhibit PP2A. Second, SV40 small t, which binds and inhibits PP2A, when overexpressed prevented H2O2-induced dephosphorylation of the retinoblastoma family proteins, whereas a SV40 small t mutant unable to bind PP2A was totally inert. Third, PP2A core enzyme physically interacted with pRb and p107, both in H2O2-treated and untreated cells. Fourth, a PP2A phosphatase activity was co-immunoprecipitated with pRb, and the activity of pRb-associated PP2A was positively modulated by cell treatment with H2O2. Because DNA damaging agents inhibit DNA synthesis in a pRb-dependent manner, it was determined whether the PP2A-mediated dephosphorylation of the retinoblastoma family proteins played a role in this S-phase response. Indeed, it was found that inhibition of PP2A by SV40 small t overexpression prevented DNA synthesis inhibition induced by H2O2.
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页码:19509 / 19517
页数:9
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