Sensitization of RPE cells by alphaB-crystallin siRNA to SAHA-induced stage 1 apoptosis through abolishing the association of alphaB-crystallin with HDAC1 in SC35 speckles.
To better understand the mechanism underlying the anti-apoptotic activity of alphaB-crystallin in RPE cells.Cells of the human retinal pigment epithelial line ARPE-19 were treated with a histone deacetylase inhibitor (HDACI), suberoylanilide hydroxamic acid (SAHA), with or without alphaB-crystallin siRNA. To examine the mechanism underlying the cell death induced in ARPE-19 ... cells, nuclear staining, flow cytometry, DNA electrophoresis, pulse field gel electrophoresis, Western blot analysis, confocal microscopy, and coimmunoprecipitation assay were undertaken.The present study demonstrated that an HDACI, SAHA, at the usual doses or the silencing of alphaB-crystallin by siRNA alone did not effectively induce apoptosis in ARPE-19 cells. Silencing of alphaB-crystallin likely abolishes the anti-apoptotic activity of alphaB-crystallin. The data indicated that silencing of alphaB-crystallin sensitizes ARPE19 cells to SAHA-induced apoptosis and leads them to stage 1 apoptosis. alphaB-Crystallin associates with HDAC1 on SC35 speckles, and silencing of alphaB-crystallin abolishes this association, resulting in the induction of apoptosis. The data indicated that the association between alphaB-crystallin and HDAC1 on SC35 speckles plays a pivotal role in anti-apoptotic activity.Knockout of alphaB-crystallin may be a promising new approach to enhance therapeutic potency for proliferative vitreoretinopathy without compromising efficacy.
Mesh Terms:
Apoptosis, Blotting, Western, Cell Count, Cells, Cultured, DNA, Down-Regulation, Electrophoresis, Enzyme Inhibitors, Flow Cytometry, Histone Deacetylase 1, Histone Deacetylase Inhibitors, Histone Deacetylases, Humans, Hydroxamic Acids, Immunoprecipitation, Microscopy, Confocal, Pigment Epithelium of Eye, RNA, Small Interfering, Vitreoretinopathy, Proliferative, alpha-Crystallin B Chain
Apoptosis, Blotting, Western, Cell Count, Cells, Cultured, DNA, Down-Regulation, Electrophoresis, Enzyme Inhibitors, Flow Cytometry, Histone Deacetylase 1, Histone Deacetylase Inhibitors, Histone Deacetylases, Humans, Hydroxamic Acids, Immunoprecipitation, Microscopy, Confocal, Pigment Epithelium of Eye, RNA, Small Interfering, Vitreoretinopathy, Proliferative, alpha-Crystallin B Chain
Invest. Ophthalmol. Vis. Sci.
Date: Nov. 01, 2008
PubMed ID: 18552382
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