Transcriptional Regulation of Mammary Alveolar Proliferation and Differentiation during Early Pregnancy

Proliferation and differentiation of the mammary gland during pregnancy is regulated by a wide variety of factors. Using gene expression data, we have predicted that the E2F5 transcription factor has a role in the mammary gland during pregnancy. Using CUT&RUN for E2F5 in combination with gene expression data revealed that there were a number of E2F5 target genes associated with gene expression in early pregnancy, suggesting a critical role. This prediction was then functionally tested through analysis of mice with an ablation of E2F5 in the mouse mammary epithelium using a Floxed E2F5 in combination with MMTV-Cre. This revealed a striking delay in alveolar proliferation and differentiation at the early stages of pregnancy. The significance of this effect was lost in both a second pregnancy and by the mid-point of pregnancy in single parous females. Analysis of E2F5 targets revealed overlap but a slight divergence from the consensus E2F sequence and that E2F5 was bound to the promoter of many genes involved in cellular proliferation, including known regulators of pregnancy associated alveolar expansion such as Stat6. However, progesterone stimulation of cells in a reporter assay revealed that it counteracts the repression of Stat6 by E2F5, leading to expression of Stat6. Together these data reveal the complexity of the transcriptional regulation of alveolar proliferation in early pregnancy.