Variation in developmental conditions can affect a variety of embryonic processes and shape a number of phenotypic characteristics that can affect offspring throughout their lives. This is particularly true of oviparous species where development typically occurs outside of the female, and studies have shown that traits such as survival and behavior can be altered by both temperature and exposure to steroid hormones during development. In species with temperature-dependent sex determination (TSD), the fate of gonadal development can be affected by temperature and by maternal estrogens present in the egg at oviposition and there is evidence that these factors can affect gene expression patterns. Here, we explore how thermal fluctuations and exposure to an estrogen metabolite, estrone sulfate, affect the expression of several genes known to be involved in sexual differentiation; Kdm6b, Dmrt1, Sox9, FoxL2, and Cyp19A1. We found that most of the genes responded to both temperature and estrone sulfate exposure, but that the responses to these factors was not identical in that estrone sulfate effects occur downstream of temperature effects. Our findings demonstrate that conjugated hormones such as estrone sulfate are capable of influencing temperature dependent pathways to potentially alter how embryos respond to temperature and highlight the importance of studying the interaction of maternal hormone and temperature effects.

Methods have been described in the body of the manuscript.

Dryad 2022_Study1_Dmrt1 R code has two data files:

Dryad 2022_Study1_Dmrt1.xlsx and Dryad 2022_Study1_Dmrt1_Baseline.xlsx

Dryad 2022_Study1_Hatchling R code has two data files:

Dryad 2022_Study1_Hatchling_Dmrt1.xlsx and Dryad 2022_Study1_Hatchling_FoxL2.xlsx

All other R code has a single xlsx file with a corresponding name.