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Accumulation of the SET protein in HEK293T cells and mild oxidative stress: cell survival or death signaling

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Abstract

SET protein (I2PP2A) is an inhibitor of PP2A, which regulates the phosphorylated Akt (protein kinase B) levels. We assessed the effects of SET overexpression in HEK293T cells, both in the presence and the absence of mild oxidative stress induced by 50 μM tert-butyl hydroperoxide. Immunoblotting assays demonstrated that SET accumulated in HEK293T cells and increased the levels of phosphorylated Akt and PTEN; in addition, SET decreased glutathione antioxidant defense of cell and increased expression of genes encoding antioxidant defense proteins. Immunofluorescence analysis demonstrated that accumulated SET was equally distributed in cytoplasm and nucleus; however, in cells that had been exposed to oxidative stress, SET was found in large aggregates in the cytoplasm. SET accumulation in HEK293T cells correlated with inhibition of basal apoptosis as evidenced by a decrease in annexin V staining and activity of caspases; under mild oxidative stress, SET accumulation correlated with caspase-independent cell death, as evidenced by increased PI and annexin V/PI double staining. The results suggest that accumulated SET could act via Akt/PTEN either as cell survival signal or as oxidative stress sensor for cell death.

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Acknowledgments

Research supported by FAPESP (Fellowship 2005/03380-2, Young research project 2006/06334-4, Grant 2009/52228-0), CNPq, CAPES and the Intramural Research Program of NIH, National Institute of Dental and Craniofacial Research. The authors thank Fabiana R. Morais for technical support with the FACS analysis.

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Authors and Affiliations

  1. Departamento de Análises Clínicas, Toxicológicas e Bromatológicas, Faculdade de Ciências Farmacêuticas de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brazil

    Andréia M. Leopoldino, Cristiana B. Garcia, Luciana O. Almeida & Sérgio A. Uyemura

  2. Division of Oral Pathology/Medicine/Radiology, Department of Periodontics and Oral Medicine, University of Michigan School of Dentistry, Ann Arbor, MI, USA

    Cristiane H. Squarize

  3. Departamento de Física e Química, Faculdade de Ciências Farmacêuticas de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brazil

    Cezar R. Pestana, Ana C. M. Polizello & Carlos Curti

  4. Departamento de Biologia Molecular, Faculdade de Medicina de São José do Rio Preto, São José do Rio Preto, SP, Brazil

    Eloiza H. Tajara

  5. Oral and Pharyngeal Cancer Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD, USA

    J. Silvio Gutkind

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  1. Andréia M. Leopoldino
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Leopoldino, A.M., Squarize, C.H., Garcia, C.B. et al. Accumulation of the SET protein in HEK293T cells and mild oxidative stress: cell survival or death signaling. Mol Cell Biochem 363, 65–74 (2012). https://doi.org/10.1007/s11010-011-1158-x

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