Daphnia magna diapause-interrupted embryogenesis has changes in histone modifications at H3K9

As a key component of freshwater ecosystem, Daphnia survives environmental challenges by entering a state of developmental arrest known as diapause. Diapause is not simply a sudden halt in growth and development, but a genetically regulated program involving a sequence of coordinated developmental processes. Histone modifications, as crucial epigenetic mechanisms that regulate gene expression by altering chromatin structure, are hypothesized to play a significant role in diapause regulation. This study focuses on the acetylation (H3K9ac) and tri-methylation (H3K9me3) at the H3K9 site. Given the established roles of H3K9ac in transcriptional activation and H3K9me3 in transcriptional silencing, we hypothesize that these antagonistic modifications play regulatory roles in the diapause developmental program of Daphnia. Via immunocytochemistry, we observed a significant overall decrease in H3K9ac and an increase in H3K9me3 levels in Daphnia magna diapausing cells, suggesting transcriptional silencing of H3K9-related genes. Notably, however, a subset of cells retained high H3K9ac and low H3K9me3 levels, indicating localized transcriptional activity of H3K9 related genes that may be essential for maintaining diapause and facilitating developmental resumption once conditions improve. We also examined the expression of the H3K9ac-related acetyltransferase gene kat2a and the H3K9me3-related methyltransferase gene suv39h1 via quantitative PCR. kat2a was highly expressed during active embryonic development (pre- and post-diapause) and maintained a basal level during diapause, potentially supporting the transcriptional activity of H3K9-related genes in selected cells. On the other hand, suv39h1 transcript levels remained low from ovulation through the entire diapause period, making its contribution to the elevated H3K9me3 levels during diapause unclear. However, the increase in suv39h1 expression toward the end of diapause suggests its potential role in diapause termination and subsequent post-diapause development. Our results illustrate dynamic changes in H3K9 modifications throughout the diapause developmental program in Daphnia, indicating their role in diapause regulation by modulating gene expression via alterations to the chromatin landscape.