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The annual killifish, Austrofundulus limnaeus, may enter embryonic diapause at three distinct points of development, termed diapause I, II, and III. Previous studies suggest a role for steroid hormones in the regulation of diapause in annual killifish. This study concerns the hormonal and genomic components involved in the developmental decision to enter or escape diapause II from both a maternal and embryonic perspective. Steroid hormone levels were measured in tissues isolated from adult female fish that were producing either high or low proportions of escape embryos. Levels of steroid hormones were also measured in new fertilized embryos that were known to be on either an escape or diapausing developmental trajectory. In addition, cDNA microarray gene expression analysis was used to identify gene sequences that may be associated with the regulation of entry into diapause in this species. Decreases in maternal estrogen levels associated with aging are correlated with decreasing escape embryo production, but there is no direct association between measured steroid hormone levels and escape embryo production. However, maternal production of escape embryos is correlated with increased ratios of 17 β-estradiol to testosterone in ovary tissue, and cDNA microarray gene expression analysis indicates differentially regulated sequences associated with escape embryo production in maternal tissues. Both of these independent measures suggest hormonal involvement in the regulation of diapause. Embryonic levels of steroid hormones in newly fertilized embryos are not correlated with entry or escape from diapause II, although ii incubation in exogenous cortisol and 17 β-estradiol causes an increase in the proportion of escape embryos. Gene expression analysis again suggests hormonal involvement. Interestingly, genes involved in epigenetic control of gene expression though chromatin condensation are differentially regulated in both maternal tissues producing escape embryos, and in embryos on the different developmental trajectories. These data suggest that hormonal control of gene expression through alterations of chromatin condensation may regulate the decision to enter or escape diapause II.

Genomic and Hormonal Components of Altered Developmental Pathways in the Annual Killifish, Austrofundulus limnaeus