Epigenetic regulation of surfactant protein A gene (SP-A) expression in fetal lung reveals a critical role for Suv39h methyltransferases during development and hypoxia

SP-A gene expression is developmentally regulated in fetal lung. Cyclic AMP (cAMP) induction of SP-A expression in human fetal lung type II cells is O(2) dependent and is mediated by increased binding of TTF-1/Nkx2.1 and NF-κB to a critical response element (TBE). This is associated with increased acetylation and decreased methylation of H3K9 at the TBE. Using chromatin immunoprecipitation analysis of fetal lung between 15.5 and 19.0 days of gestation, we observed that the developmental induction of SP-A was associated with increased recruitment of TTF-1, NF-κB, PCAF, and CBP, as well as enhanced acetylation and decreased methylation of histone H3K9 at the TBE. Importantly, expression and TBE binding of the H3K9 methyltransferases, Suv39h1 and Suv39h2, was inversely correlated with the developmental upregulation of SP-A. In human fetal lung epithelial cells, Suv39H1 and Suv39H2 mRNA levels declined with cAMP induction of SP-A. Moreover, hypoxia, which inhibits cAMP stimulation of SP-A, markedly increased Suv39h1 and Suv39h2 binding to the TBE. Finally, short hairpin RNA knockdown of Suv39H1 or Suv39H2 in fetal lung epithelial cells repressed H3K9 methylation and greatly enhanced SP-A expression. Collectively, our findings suggest that Suv39H1 and Suv39H2 are key hypoxia-induced methyltransferases; their decline in fetal lung during late gestation is critical for epigenetic changes resulting in the developmental induction of SP-A.     

Regulation of human pulmonary surfactant protein gene expression by 1alpha,25-dihydroxyvitamin D3

1alpha,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] has been reported to stimulate lung maturity, alveolar type II cell differentiation, and pulmonary surfactant synthesis in rat lung. We hypothesized that 1,25(OH)(2)D(3) stimulates expression of surfactant protein-A (SP-A), SP-B, and SP-C in human fetal lung and type II cells. We found that immunoreactive vitamin D receptor was detectable in fetal lung tissue and type II cells only when incubated with 1,25(OH)(2)D(3). 1,25(OH)(2)D(3) significantly decreased SP-A mRNA in human fetal lung tissue but did not significantly decrease SP-A protein in the tissue. In type II cells, 1,25(OH)(2)D(3) alone had no significant effect on SP-A mRNA or protein levels but reduced SP-A mRNA and protein in a dose-dependent manner when the cells were incubated with cAMP. SP-A mRNA levels in NCI-H441 cells, a nonciliated bronchiolar epithelial (Clara) cell line, were decreased in a dose-dependent manner in the absence or presence of cAMP. 1,25(OH)(2)D(3) had no significant effect on SP-B mRNA levels in lung tissue but increased SP-B mRNA and protein levels in type II cells incubated in the absence or presence of cAMP. Expression of SP-C mRNA was unaffected by 1,25(OH)(2)D(3) in lung tissue incubated +/- cAMP. These results suggest that regulation of surfactant protein gene expression in human lung and type II cells by 1,25(OH)(2)D(3) is not coordinated; 1,25(OH)(2)D(3) decreases SP-A mRNA and protein levels in both fetal lung tissue and type II cells, increases SP-B mRNA and protein levels only in type II cells, and has no effect on SP-C mRNA levels.