Methylation of the <i>Gpat2</i> promoter regulates transient expression during mouse spermatogenesis

Abstract

Spermatogenesis is a highly regulated process that involves both mitotic and meiotic divisions, as well as cellular differentiation to yield mature spermatozoa from undifferentiated germinal stem cells. Although <i>Gpat2</i> was originally annotated as a glycerol-3-phospate acyltransferase by sequence homology to <i>Gpat1</i>, GPAT2 is highly expressed in testis but not in lipogenic tissues and is not up-regulated during adipocyte differentiation. New data show that GPAT2 is required for the synthesis of piRNAs, a group of small RNAs that protect the germ cell genome from retrotransposable elements. In order to understand the relationship between GPAT2 and its role in the testis, we focused on <i>Gpat2</i> expression during the first wave of mouse spermatogenesis. <i>Gpat2</i> expression was analyzed by qPCR, <i>in situ</i> hybridization, immunohistochemistry and Western blot. <i>Gpat2</i> mRNA content and protein expression were maximal at 15 dpp and restricted to pachytene spermatocytes. To achieve this transient expression, both epigenetic mechanisms and <i>trans</i>-acting factors are involved. <i>In vitro</i> assays showed that <i>Gpat2</i> expression correlates with DNA demethylation and histone acetylation and that it is up-regulated by retinoic acid. Epigenetic regulation by DNA methylation was confirmed <i>in vivo</i> in germ cells by bisulfite sequencing of the <i>Gpat2</i> promoter. Consistent with the initiation of meiosis at 11 dpp, methylation decreased dramatically. Thus, <i>Gpat2</i> is expressed at a specific stage of spermatogenesis, consistent with piRNA synthesis and meiosis I prophase, and its on-off expression pattern responds predominantly to epigenetic modifications.