Methylation status of CpG islands flanking a cAMP response element motif on the protein phosphatase 2Ac alpha promoter determines CREB binding and activity.
Protein phosphatase 2A (PP2A) is a major serine/threonine protein phosphatase in eukaryotic cells and is involved in many essential aspects of cell function. The catalytic subunit of the enzyme (PP2Ac), a part of the core enzyme, has two isoforms, alpha (PP2Ac alpha) and beta (PP2Ac beta), of which PP2Ac alpha is the major form expressed in vivo. Deregulation of PP2A expression has been linked to several diseases, but the mechanisms that control the expression of this enzyme are still unclear. We conducted experiments to decipher molecular mechanisms involved in the regulation of the PP2Ac alpha promoter in human primary T cells. After preparing serially truncated PP2Ac alpha promoter luciferase constructs, we found that the region stretching around 240 bases upstream from the translation initiation site was of functional significance and included a cAMP response element motif flanked by three GC boxes. Shift assays revealed that CREB/phosphorylated CREB and stable protein 1 could bind to the region. Furthermore, we demonstrated that methylation of deoxycytosine in the CpG islands limited binding of phosphorylated CREB and the activity of the PP2Ac alpha promoter. In contrast, the binding of stable protein 1 to a GC box within the core promoter region was not affected by DNA methylation. Primary T cells treated with 5-azacitidine, a DNA methyltransferase inhibitor, showed increased expression of PP2Ac alpha mRNA. We propose that conditions associated with hypomethylation of CpG islands, such as drug-induced lupus, permit increased PP2Ac expression.
Sunahori
Katsue
K
Division of Rheumatology in Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02115, USA.
Juang
Yuang-Taung
YT
Tsokos
George C
GC
D020816:Amino Acid Motifs / Q000276:immunology; D000818:Animals; D001483:Base Sequence; D050882:CREB-Binding Protein / Q000378:metabolism*; D002478:Cells, Cultured; D018899:CpG Islands / Q000276:immunology; D017362:Cyclic AMP Response Element-Binding Protein / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism*; D019175:DNA Methylation / Q000276:immunology; D004795:Enzyme Stability / Q000276:immunology; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000276:immunology; D011485:Protein Binding / Q000276:immunology; D054648:Protein Phosphatase 2 / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats
D017362:Cyclic AMP Response Element-Binding Protein; D007527:Isoenzymes; D012333:RNA, Messenger; D050882:CREB-Binding Protein; C517189:PPP2CA protein, human; D054648:Protein Phosphatase 2