During this reporting period we investigated the role of the epigenetic regulators EZH1 and EZH2 in liver homeostasis, and generated mice that carried mutant Ezh1 and Ezh2 alleles in hepatocytes. Only the combined loss of EZH1 and EZH2 caused a depletion of global trimethylation on Lys 27 of histone H3 (H3K27me3) marks and the specific loss of these marks on over ∼1900 genes. These mice exhibited progressive liver abnormalities manifested by the development of regenerative nodules and concomitant periportal fibrosis, inflammatory infiltration, and activation of A6-positive hepatic progenitor cells at 8 months of age. In response to chronic treatment with carbon tetrachloride, all experimental mice, but none of the controls, showed increased hepatic degeneration associated with liver dysfunction and reduced ability to proliferate. After partial hepatectomy, mutant mice displayed increased liver injury and a blunted regenerative response. Genome-wide analyses identified approximately 50 genes that had lost H3K27me3 marks, and their expression was significantly increased. These genes were involved in the regulation of cell survival, fibrosis, and proliferation. H3K27me3 levels and liver physiology were unaffected in mice lacking either EZH1 globally or EZH2 specifically in hepatocytes. This work demonstrates a critical role and redundancy of EZH1 and EZH2 in maintaining hepatic homeostasis and regeneration. During this reporting period we have also continued to explore the role of the histone methyltransferases EZH1 and EZH2 in the biology of mammary epithelium. Establishment and differentiation of mammary alveoli during pregnancy are controlled by prolactin through the transcription factors STAT5A and STAT5B (STAT5), which regulate temporal activation of mammary signature genes. Our study addressed the question whether the methyltransferase and transcriptional co-activator EZH2 controls the differentiation clock of mammary epithelium. Ablation of Ezh2 from mammary stem cells resulted in precocious differentiation of alveolar epithelium during pregnancy and the activation of mammary-specific STAT5 target genes. This coincided with enhanced occupancy of these loci by STAT5, EZH1 and RNA Pol II. Limited activation of differentiation-specific genes was observed in mammary epithelium lacking both EZH2 and STAT5, suggesting a modulating but not mandatory role for STAT5. Loss of EZH2 did not result in overt changes in genome-wide and gene-specific H3K27me3 profiles, suggesting compensation through enhanced EZH1 recruitment. Differentiated mammary epithelia did not form in the combined absence of EZH1 and EZH2. Transplantation experiments failed to demonstrate a role for EZH2 in the activity of mammary stem and progenitor cells. In summary, while EZH1 and EZH2 serve redundant functions in the establishment of H3K27me3 marks and the formation of mammary alveoli, the presence of EZH2 is required to control progressive differentiation of milk secreting epithelium during pregnancy.
