Aldosterone reprograms promoter methylation to regulate αENaC transcription in the collecting duct.
Aldosterone increases tubular Na(+) absorption largely by increasing α-epithelial Na(+) channel (αENaC) transcription in collecting duct principal cells. How aldosterone reprograms basal αENaC transcription to high-level activity in the collecting duct is incompletely understood. Promoter methylation, a covalent but reversible epigenetic process, has been implicated in the control of gene expression in health and disease. We investigated the role of promoter methylation/demethylation in the epigenetic control of basal and aldosterone-stimulated αENaC transcription in mIMCD3 collecting duct cells. Bisulfite treatment and sequencing analysis after treatment of the cells with the DNA methyltransferase (DNMT) inhibitor 5-aza-2'-deoxycytidine (5-Aza-CdR) identified clusters of methylated cytosines in a CpG island near the transcription start site of the αENaC promoter. 5-Aza-CdR treatment or small interfering RNA-mediated knockdown of DNMT3b or methyl-CpG-binding domain protein (MBD)-4 derepressed basal αENaC transcription, indicating that promoter methylation suppresses basal αENaC transcription. Aldosterone triggered a time-dependent decrease in 5mC and DNMT3b and a concurrent enrichment in 5-hydroxymethylcytosine (5hmC) and ten-eleven translocation (Tet)2 at the αENaC promoter, consistent with active demethylation. 5-Aza-CdR mimicked aldosterone by enhancing Sp1 binding to the αENaC promoter. We conclude that DNMT3b- and MBD4-dependent methylation of the αENaC promoter limits basal αENaC transcription, in part by limiting Sp1 binding and trans-activation. Aldosterone stimulates the dispersal of DNMT3b and recruitment of Tet2 to demethylate the αENaC promoter to induce αENaC transcription. These results disclose a novel epigenetic mechanism for the control of basal and aldosterone-induced αENaC transcription that adds to previously described epigenetic controls exerted by histone modifications.
Div. of Renal Diseases and Hypertension, The Univ. of Texas Medical School at Houston, 6431 Fannin, MSB 5.124, Houston, TX 77030. Bruce.C.Kone@uth.tmc.edu.
Zhiyuan
Z
Yu
Qun
Q
Kong
Bruce C
BC
Kone
D000450:Aldosterone / Q000502:physiology*; D000818:Animals; D001374:Azacitidine / Q000031:analogs & derivatives; D002460:Cell Line; D002478:Cells, Cultured; D003596:Cytosine / Q000378:metabolism; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000235:genetics / Q000378:metabolism*; D019175:DNA Methylation / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D000077209:Decitabine; D049308:Dioxygenases; D004706:Endodeoxyribonucleases / Q000235:genetics / Q000378:metabolism; D053503:Epithelial Sodium Channels / Q000378:metabolism*; D007685:Kidney Tubules, Collecting / Q000378:metabolism*; D051379:Mice; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D000097584:DNA Methyltransferase 3B
D000450:Aldosterone; D001374:Azacitidine; D003596:Cytosine; D004248:DNA (Cytosine-5-)-Methyltransferases; D004268:DNA-Binding Proteins; D000077209:Decitabine; D049308:Dioxygenases; D004706:Endodeoxyribonucleases; D053503:Epithelial Sodium Channels; D011518:Proto-Oncogene Proteins; D016329:Sp1 Transcription Factor; D000097584:DNA Methyltransferase 3B; C551914:Tet2 protein, mouse; C492249:Mbd4 protein, mouse
chromatin; epigenetic; epithelial Na+ channel; methylcytosine; methyltransferase; transcription factor