Epigenetic regulation of histone modifications and Gata6 gene expression induced by maternal diet in mouse embryoid bodies in a model of developmental programming

Background: Dietary interventions during pregnancy alter offspring fitness. We have shown mouse maternal low protein diet fed exclusively for the preimplantation period (Emb-LPD) before return to normal protein diet (NPD) for the rest of gestation, is sufficient to cause adult offspring cardiovascular and metabolic disease. Moreover, Emb-LPD blastocysts sense altered nutrition within the uterus and activate compensatory cellular responses including stimulated endocytosis within extra-embryonic trophectoderm and primitive endoderm (PE) lineages to protect fetal growth rate. However, these responses associate with later disease. Here, we investigate epigenetic mechanisms underlying nutritional programming of PE that may contribute to its altered phenotype, stabilised during subsequent development. We use embryonic stem (ES) cell lines established previously from Emb-LPD and NPD blastocysts that were differentiated into embryoid bodies (EBs) with outer PE-like layer. Results: Emb-LPD EBs grow to a larger size than NPD EBs and express reduced Gata6 transcription factor (regulator of PE differentiation) at mRNA and protein levels, similar to Emb-LPD PE derivative visceral yolk sac tissue in vivo in later gestation. We analysed histone modifications at the Gata6 promoter in Emb-LPD EBs using chromatin immunoprecipitation assay. We found significant reduction in histone H3 and H4 acetylation and RNA polymerase II binding compared with NPD EBs, all markers of reduced transcription. Other histone modifications, H3K4Me2, H3K9Me3 and H3K27Me3, were unaltered. A similar but generally non-significant histone modification pattern was found on the Gata4 promoter. Consistent with these changes, histone deacetylase Hdac-1, but not Hdac-3, gene expression was upregulated in Emb-LPD EBs. Conclusions: First, these data demonstrate ES cells and EBs retain and propagate nutritional programming adaptations in vitro, suitable for molecular analysis of mechanisms, reducing animal use. Second, they reveal maternal diet induces persistent changes in histone modifications to regulate Gata6 expression and PE growth and differentiation that may affect lifetime health.