Assay of the redox state of the tumor suppressor PTEN by mobility shift

Han, Seong-Jeong; Ahn, Younghee; Park, Iha; Zhang, Ying; Kim, In-Young; Kim, Hyun Woo; Ku, Chang-Sub; Chay, Kee-Oh; Yang, Sung Yeul; Ahn, Bong Whan; Jang, Dong Il; Lee, Seung-Rock

doi:10.1016/j.ymeth.2015.01.007

Cited by 19 publications

(15 citation statements)

References 24 publications

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“…Protease inhibitor (Complete ULTRA Tablets) was purchased from Roche Diagnostics GmbH (Indianapolis, IN, USA). PTEN antibody was prepared as previously described [50]. Anti-Trx1 and anti-rabbit IgG horseradish peroxidase-conjugated antibodies were purchased from Ab Frontier (Daejeon, Korea).…”

Section: Methodsmentioning

confidence: 99%

Redox Regulation of the Tumor Suppressor PTEN by Hydrogen Peroxide and Tert-Butyl Hydroperoxide

Zhang

Han

Park

et al. 2017

IJMS

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Organic peroxides and hydroperoxides are skin tumor promoters. Free radical derivatives from these compounds are presumed to be the prominent mediators of tumor promotion. However, the molecular targets of these species are unknown. Phosphatase and tensin homologs deleted on chromosome 10 (PTEN) are tumor suppressors that play important roles in cell growth, proliferation, and cell survival by negative regulation of phosphoinositol-3-kinase/protein kinase B signaling. PTEN is reversibly oxidized in various cells by exogenous and endogenous hydrogen peroxide. Oxidized PTEN is converted back to the reduced form by cellular reducing agents, predominantly by the thioredoxin (Trx) system. Here, the role of tert-butyl hydroperoxide (t-BHP) in redox regulation of PTEN was analyzed by using cell-based and in vitro assays. Exposure to t-BHP led to oxidation of recombinant PTEN. In contrast to H2O2, PTEN oxidation by t-BHP was irreversible in HeLa cells. However, oxidized PTEN was reduced by exogenous Trx system. Taken together, these results indicate that t-BHP induces PTEN oxidation and inhibits Trx system, which results in irreversible PTEN oxidation in HeLa cells. Collectively, these results suggest a novel mechanism of t-BHP in the promotion of tumorigenesis.

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Section: Methodsmentioning

confidence: 99%

Redox Regulation of the Tumor Suppressor PTEN by Hydrogen Peroxide and Tert-Butyl Hydroperoxide

Zhang

Han

Park

et al. 2017

IJMS

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“…In general, several protein phosphatases contain redox-sensitive cysteines in their catalytic center, and some KEAP1 inhibitors may modify and inactivate these phosphatases, hence disturbing signaling networks. One of these phosphatases is PTEN (Lee et al, 2002;Kitagishi and Matsuda, 2013;Han et al, 2015). The catalytic C124 residue of PTEN can be modified through adduct formation with strong electrophiles such as CDDO-Im (Pitha-Rowe et al, 2009) and tertbutylhydroquinone (Rojo et al, 2014b).…”

Section: A Electrophilic Nuclear Factor (Erythroid-derived 2)-like 2mentioning

confidence: 99%

Transcription Factor NRF2 as a Therapeutic Target for Chronic Diseases: A Systems Medicine Approach

et al. 2018

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“…Dulbecco's modified Eagle's medium (DMEM), fetal bovine serum (FBS), Lipofectamine 2000 transfection reagent, and anti-actin antibody were purchased from Thermo Fisher Scientific (Waltham, MA, USA). The PTEN and Trx antibodies were prepared as described previously [39, 40]. Anti-rabbit IgG horseradish peroxidase-conjugated antibody was purchased from Ab Frontier (Daejeon, Korea).…”

Section: Methodsmentioning

confidence: 99%

“…Oxidative modifications of PTEN specifically involved in the formation of intramolecular disulfide bonds were readily identified by nonreducing sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblot analysis, as described previously [39]. After incubation, cells were washed three times with ice-cold PBS and lysed with NP-40 lysis buffer (20 mM Tris-HCl (pH 7.5), 150 mM NaCl, 5% glycerol, 0.1% NP-40, 1 mM phenylmethylsulfonylfluoride, and protease inhibitor cocktail) containing 10 mM NEM.…”

Section: Methodsmentioning

confidence: 99%

Peroxiredoxin III Protects Tumor Suppressor PTEN from Oxidation by 15-Hydroperoxy-eicosatetraenoic Acid

Zhang

Park

Han³

et al. 2019

Oxidative Medicine and Cellular Longevity

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Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) is a lipid and protein phosphatase that coordinates various cellular processes. Its activity is regulated by the reversible oxidation of an active-site cysteine residue by H2O2 and thioredoxin. However, the potential role of lipid peroxides in the redox regulation of PTEN remains obscure. To evaluate this, 15-hydroperoxy-eicosatetraenoic acid (15s-HpETE), a lipid peroxide, was employed to investigate its effect on PTEN using molecular and cellular-based assays. Exposure to 15s-HpETE resulted in the oxidation of recombinant PTEN. Reversible oxidation of PTEN was also observed in mouse embryonic fibroblast (MEF) cells treated with a 15s-HpETE and Lipofectamine mixture. The oxidative dimerization of thioredoxin was found simultaneously. In addition, the absence of peroxiredoxin III aggravated 15s-HpETE-induced PTEN oxidation in MEF cells. Our study provides novel insight into the mechanism linking lipid peroxidation to the etiology of tumorigenesis.

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Assay of the redox state of the tumor suppressor PTEN by mobility shift

Cited by 19 publications

References 24 publications

Redox Regulation of the Tumor Suppressor PTEN by Hydrogen Peroxide and Tert-Butyl Hydroperoxide

Redox Regulation of the Tumor Suppressor PTEN by Hydrogen Peroxide and Tert-Butyl Hydroperoxide

Transcription Factor NRF2 as a Therapeutic Target for Chronic Diseases: A Systems Medicine Approach

Peroxiredoxin III Protects Tumor Suppressor PTEN from Oxidation by 15-Hydroperoxy-eicosatetraenoic Acid

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