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R. Kannan, S.G. Parameswaran, C.G. Charlton, S.J. Ryan, D.R. Hinton; Expression, Characterization and Regulation of Methionine Adenosyltransferases (MATs) in Retinal Pigment Epithelial Cells . Invest. Ophthalmol. Vis. Sci. 2006;47(13):2881.
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Methionine adenosyltransferases (MATs) are essential for cellular methylation and for synthesis of polyamines and glutathione. Very little is known regarding expression and regulation of MATs in human RPE cells. In this work, we have characterized isoforms of MATs expressed in RPE cells. We have also studied the effect of C2–ceramide and growth factors in modulating MATs expression.
Human RPE cells from passages 3–4 were maintained overnight in 1% FBS and treated with different doses of C2–ceramide (5µM –30µM) for 24h. In parallel, cultures were incubated overnight with 20ng/ml human recombinant hepatocyte growth factor (HGF)or preincubated with HGF before ceramide treatment. Gene and protein expression were studied by real–time PCR and western blot analysis, respectively. The presence of MAT2A and MAT2B genes were verified after subcloning and sequencing.
We identified and characterized the presence of two isoforms of MATs (MAT2A and MAT2B) in human RPE cells. MAT1A, the liver specific isoform of MAT, could not be detected in RPE. Treatment with C2–ceramide resulted in a dose–response down regulation of MAT2A gene expression. Treatment of RPE with HGF alone increased MAT2A expression significantly (p<0.05 vs controls). The levels of MAT2A remained higher than controls in RPE incubated with HGF before exposure to C2–ceramide.
Two isoforms of MATs namely, MAT2A and MAT2B are expressed in RPE cells and are regulated by ceramide–induced oxidative stress. Treatment with hepatocyte growth factor upregulates MATs thereby decreasing oxidative injury to RPE.
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