Sex is determined by genetic constitution (GSD) in most animals (as by sex chromosomes in humans), and by environmental temperature (TSD) in others, including reptiles. The proportion of males to females (sex ratio) greatly influences populations. Male-biased populations reproduce slowly because females are limited, while female-biased populations may have lower genetic variability because few males father all offspring. Yet we know little about how naturally-fluctuating temperatures of nests affect sex ratios of TSD reptiles. This gap obscures the effect of TSD on sex ratio evolution, the growth potential of populations, the rate of change of genetic variability, and how species adapt to the environment. This project will help elucidate the effect that naturally-fluctuating temperature has on the expression of genes involved in gonadal formation in TSD turtles, and how males and females are produced in nature. Explicit hypotheses derived from previous gene expression and ecological research will be tested to help reveal how TSD species may respond to short- and long-term climate change.
Intellectual Merit: This project will advance our understanding of the effects of environmental change on the regulation of sexual development and sex ratio production at the gene-expression level in TSD species. The application of molecular techniques under ecologically-relevant conditions will shed light on how and why sex is determined by nest temperature in so many reptiles, thus facilitating the study of sex ratio evolution under TSD and its implications for the conservation of endangered TSD species facing global warming.
Broader Impacts: This project will (a) integrate undergraduate discovery-based learning through a novel service-learning activity to recruit biology undergraduates into the field and lab modules of this project; and (b) broaden the participation of underrepresented groups in biology by providing direct training opportunities for students and their mentors; and by engaging women and minorities through outreach activities.
In most animals, sex is determined by genotypic mechanisms (GSD). However, in many reptiles sex is determined by the environmental temperature (TSD). Sex ratio is a key life history component. Yet we know little about how natural temperature fluctuations experienced by nests in the wild affect sex ratios of TSD reptiles. This gap hinders our understanding of sex ratio evolution in TSD species and consequently, on how TSD affects the growth potential of populations, their effective population size, and their rates of genetic and phenotypic evolution. Results from this project shed light on the effect that naturally fluctuating temperature has on the expression of genes involved in gonadal formation in TSD turtles, and how sex ratios are produced in nature. In particular, by comparing gene activity at constant, simple fluctuations, and natural-nest fluctuation profiles, we found that temperature variation alters the timing and co-expression of genes involved in gonadal formation in a complex manner in the wild. Our findings underscore the potential role of Sf1 and Wt1 as critical genes in TSD-specific sexual development; indicate that the role of Aromatase expression in female development is not absolute but context-dependent; uncovered the first evidence of the involvement of Dax1 in male development in a TSD vertebrate, and suggest that still-unknown modules of genes might exist that respond independently under different environmental conditions to produce males and females in nature. Also, contrasting gene expression among TSD and GSD species revealed evidence that the regulation of the network of genes underlying reptilian TSD has accrued significant evolutionary changes among lineages, that GSD gene expression is less responsive to environmental perturbation as is its sexual development, and point to Sf1 as a potential sex-determining gene in GSD turtles. Explicit hypothesis derived from previous gene expression and ecological research were tested to help reveal how TSD species may respond to short- and long-term climate change. While greater amplitude of temperature variation under simple fluctuations resulted in sex reversals and suggested that the feminizing effect of higher global average temperature might be countered by increased thermal variance, later experiments using natural nest profiles showed that sex reversals only occurred around nest profiles that normally would produce males. Thus, our data suggest that greater thermal variance predicted to accompany higher average temperature under climate change could accelerate the feminization of populations and consequently increase their extinction risk. Intellectual Merit — This project has advanced our understanding of the effects of environmental change on the regulation of sex differentiation and sex ratio production at the gene-expression level in TSD species. The application of molecular techniques within ecologically-relevant conditions shed light on the ecological and evolutionary significance of TSD in oviparous animals, thus facilitating the study of sex ratio evolution under TSD and its implications for the conservation of endangered TSD species facing global warming. The complex responses that were observed indicate that both increasing the mean incubation temperature and the thermal variance affect the regulation of the gene network underlying sex determination in nature, and highlight the urgency to decipher the global effect of thermal fluctuations on genome-wide expression to decipher the complex molecular regulation underpinning sexual development if we are to fully understand the real-world perils faced by TSD systems as climate changes. Broader Impact — This project provided training opportunities and integrated undergraduate discovery-based learning for undergraduate students and K-12 teachers through the Iowa Turtle Army (ITA) program developed by the PI. During the tenure of this grant, 27 undergraduate students and 3 K-12 teachers participated into the field and lab modules in the PI’s lab, and learned field and experimental biology, developmental and molecular biology, herpetology and conservation. This project also provided training and development opportunities to 2 graduate and 2 postdoctoral students in research and mentoring. Additionally, this project helped broaden the participation of underrepresented groups in biology by providing direct training opportunities for underrepresented students (22 female and 4 minority) and their mentors (1 minority); and by engaging women and minorities through outreach activities such as the Program for Women in Science and Engineering, the George Washington Carver Summer Program, and the Undergraduate Diversity Program of the Society for the Study of Evolution and the Society for Systematic Biology, among others. K-12 teachers developed curricular material during their summer internships with the ITA, including posters and lesson plans. It is intended that this material be shared for their use by peer teachers and institutions, and can be downloaded form the ITA’s website created as part of this project.
