ERα/ERß-directed CBS transcription mediates E2ß-stimulated hUAEC H2S production.

ABSTRACT

Elevated endogenous estrogens stimulate human uterine artery endothelial cell (hUAEC) hydrogen sulfide (H2S) production by selectively upregulating the expression of H2S synthesizing enzyme cystathionine ß-synthase (CBS), but the underlying mechanisms are underdetermined. We hypothesized that CBS transcription mediates estrogen-stimulated pregnancy-dependent hUAEC H2S production. Estradiol-17ß (E2ß) stimulated CBS but not cystathionine γ-lyase (CSE) expression in pregnant human uterine artery ex vivo, which was attenuated by the estrogen receptor (ER) antagonist ICI 182,780. E2ß stimulated CBS mRNA/protein and H2S production in primary hUAEC from nonpregnant and pregnant women, but with greater responses in pregnant state; all were blocked by ICI 182,780. Human CBS promoter contains multiple estrogen-responsive elements (EREs), including one ERE preferentially binding ERα (αERE) and three EREs preferentially binding ERß (ßERE), and one full ERE (α/ßERE) and one half ERE (½α/ßERE) binding both ERα and ERß. Luciferase assays using reporter genes driven by human CBS promoter with a series of 5'-deletions identified the α/ßEREs binding both ERα and ERß (α/ßERE and ½α/ßERE) to be important for baseline and E2ß-stimulated CBS promoter activation. E2ß stimulated ERα/ERß heterodimerization by recruiting ERα to α/ßEREs and ßERE, and ERß to ßERE, α/ßEREs, and αERE. ERα or ERß agonist alone trans-activated CBS promoter, stimulated CBS mRNA/protein and H2S production to levels comparable to that of E2ß-stimulated, while ERα or ERß antagonist alone abrogated E2ß-stimulated responses. E2ß did not change human CSE promoter activity and CSE mRNA/protein in hUAEC. Altogether, estrogen-stimulated pregnancy-dependent hUAEC H2S production occurs by selectively upregulating CBS expression via ERα/ERß-directed gene transcription.