New study analyzes different mechanisms in the establishment of sex-phenotype dependent methylation in mouse livers

By Hector Galvez and Qinwei Zhuang

AlOgayil, N., Bauermeister, K., Galvez, J.H. et al. Distinct roles of androgen receptor, estrogen receptor alpha, and BCL6 in the establishment of sex-biased DNA methylation in mouse liver. Sci Rep 11, 13766 (2021).

Recently, using mice with different combinations of genetic and phenotypic sex, we were able to identify sex-associated differentially methylated regions (sDMRs) that depended on the sex phenotype (see this related study). However, the mechanisms behind the establishment of those differentially methylated regions are still being studied. In a new paper by Najla AlOgayil from Prof. Anna Naumova’s lab, including contributions from members of our own lab group, we show that androgen receptor (Ar), estrogen receptor alpha (Esr1), and the transcriptional repressor Bcl6 all play a distinct role in the process.

Focusing on a panel of validated sex-phenotype dependent male- and female-biased sDMRs, we tested the developmental dynamics of sex bias in liver methylation and the impacts of mutations in these three genes of interest. Our data show that sex bias in methylation either coincides with, or follows, sex bias in the expression of sDMR-proximal genes, suggesting that sex bias in gene expression may be required for demethylation at certain sDMRs. Briefly, AR and ESR1 influence DNA methylation through their impacts on gene expression, whereas BCL6 not only regulates expression, but also serves as a bridge between expression and demethylation at intragenic regions.

For these analyses, we relied on pyrosequencing methylation assays to compare DNA methylation levels in female and male livers at three different ages, representing fetal (E14.5), prepubertal (4 weeks), and adult (8 weeks) life. Additionally, we analyzed motif enrichment in all sex-phenotype dependent sDMRs using HOMER. Finally, to understand the relationship between sex-biased methylation and sex-biased expression, we studied the expression profiles of sex-biased genes using RNA-seq data from the livers of knockout mice strains for the genes Esr1 and Bcl6, and compared them to control groups using the GenPipes RNA-seq pipeline.

Taken together, the data we published serve as evidence that sex phenotype-dependent autosomal DNA methylation levels in the mouse liver depend on not one, but several factors, including AR, ESR1, and BCL6. However, more research is needed to further elucidate how multiple signaling pathways affecting gene regulation, including steroid sex hormones, can modify methylation, at least in the early ages when methylation patterns are being established.

Published: July 28 2021