Supplementary material from "Brief exposure to warm temperatures reduces intron retention in Kdm6b in a species with temperature-dependent sex determination"

Animals with temperature-dependent sex determination (TSD) respond to thermal cues during early embryonic development to trigger gonadal differentiation. TSD has primarily been studied using constant temperature incubations, where embryos are exposed to constant male- or female-producing temperatures, and these studies have identified genes that display sex-specific expression in response to incubation temperature. Kdm6b, a histone demethylase, has received specific attention as it is among the initial genes to respond to incubation temperature and is necessary for testis development. Interestingly, Kdm6b retains an intron when eggs are incubated at a constant male-producing temperature, but the role of thermal variability in this developmental process is relatively understudied. Species with TSD regularly experience thermal cues that fluctuate between male- and female-producing temperatures throughout development but it is unclear how Kdm6b responds to such variable temperatures. In this study, we investigate temperature-sensitive splicing in Kdm6b by exposing embryos to male- and female-promoting thermal conditions. We show a rapid decrease in levels of the intron retaining transcript of Kdm6b upon exposure to female-producing conditions. These results demonstrate that, under ecologically relevant conditions, temperature-sensitive splicing can differentially regulate genes critical to TSD.