Dot1l deficiency leads to increased intercalated cells and upregulation of V-ATPase B1 in mice

Exp Cell Res. 2016 Jun 10;344(2):167-75. doi: 10.1016/j.yexcr.2015.09.014. Epub 2015 Sep 25.

Abstract

The collecting duct in the mammalian kidney consists of principal cells (PCs) and intercalated cells (ICs), which regulate electrolyte/fluid and acid/base balance, respectively. The epigenetic regulators of PC and IC differentiation remain obscure. We previously used Aqp2 and V-ATPase B1B2 to label PCs and ICs, respectively. We found that mice with histone H3 K79 methyltransferase Dot1l disrupted in Aqp2-expressing cells (Dot1l(AC)) vs. Dot1l(f/f) possessed ~20% more ICs coupled with a similar decrease in PCs. Here, we performed multiple double immunofluorescence staining using various PC and IC markers and confirmed that this finding. Both α-IC and β-IC populations were significantly expanded in Dot1l(AC) vs. Dot1l(f/f). These changes are associated with significantly upregulated V-ATPase B1 and B2, but not Aqp2, AE1, and Pendrin. Chromatin immunoprecipitation assay unveiled a significant reduction of Dot1l and H3K79 di-methylation bound at the Atp6v1b1 5' flanking region. Overexpression of Dot1a significantly downregulated a stably-transfected luciferase reporter driven by the Atp6v1b1 promoter in IMCD3 cells. This downregulation was impaired, but not completely abolished when a methyltransferase-dead mutant was overexpressed. Taken together, our data suggest that Dot1l is a new epigenetic regulator of PC and IC differentiation and Atp6v1b1 is a new transcriptional target of Dot1l.

Keywords: Conditional knockout; Histone methyltransferase Dot1l; Intercalated cells; Principal cells; V-ATPase.

MeSH terms

  • Animals
  • Anion Exchange Protein 1, Erythrocyte / metabolism
  • Aquaporin 2 / metabolism
  • Aquaporin 3 / metabolism
  • Down-Regulation
  • Fluorescent Antibody Technique
  • Histone-Lysine N-Methyltransferase
  • Histones / metabolism
  • Kidney / cytology*
  • Lysine / metabolism
  • Methylation
  • Methyltransferases / deficiency*
  • Methyltransferases / genetics
  • Methyltransferases / metabolism
  • Mice
  • Promoter Regions, Genetic / genetics
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Up-Regulation*
  • Vacuolar Proton-Translocating ATPases / genetics
  • Vacuolar Proton-Translocating ATPases / metabolism*

Substances

  • Anion Exchange Protein 1, Erythrocyte
  • Aquaporin 2
  • Histones
  • RNA, Messenger
  • Aquaporin 3
  • Dot1l protein, mouse
  • Methyltransferases
  • Histone-Lysine N-Methyltransferase
  • Atp6v1b1 protein, mouse
  • Vacuolar Proton-Translocating ATPases
  • Lysine