Identification of a Genetic Pathway Linking Temperature with Epigenetic Control of Gonad Determination in T. scripta

T. scripta 中温度与性腺决定的表观遗传控制之间的遗传途径的鉴定

• 批准号: 1854642
• 负责人: Blanche Capel
• 金额: $ 90万
• 依托单位: Duke University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-05-01 至 2024-04-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1854642&HistoricalAwards=false
• 关键词: Identification; Genetic; Pathway; Linking; Temperature

Temperature regulates male or female development in many reptiles, including the redeared slider turtle Trachemys scripta elegans, by controlling whether the embryonic gonad develops as a testis or an ovary. However, the mechanism through which temperature influences gonad development has been a puzzle since the discovery of this phenomenon 50 years ago. Previously, the Capel lab identified two genes that show strong expression differences in the turtle gonad at male- versus female-producing temperatures. In this project, the lab will determine how the expression of these genes is controlled by temperature, and how the activity of these genes controls testis versus ovary development. Results from this work will establish the mechanism underlying the effect of temperature on male or female development. The lab is actively involved in using this project as a platform to engage young students and the public in scientific research. This project is also a strong platform to advocate for evidence-based science in setting environmental policies to mitigate the effects of climate change, expected to strongly bias male/female ratios in species where temperature influences gonad development.The Capel lab previously showed that transcription of the epigenetic regulator KDM6B and the RNA-splicing factor, RBM20, are elevated in the turtle gonad at male-producing temperature. KDM6B activates the testis pathway by removing repressive histone marks at DMRT1, a conserved regulator of testis development. In this project, the lab plans to identify the molecular pathway linking temperature with changes in KDM6B and RBM20 transcription and splicing. The lab hypothesizes that a known repressor of KDM6B, pSTAT3, is differentially phosphorylated at high female-producing temperatures through activation of TRP channels, Ca2+ influx, and high activity of CAMK. To test this hypothesis, markers of the male and female pathways will be measured after knockdowns of TRP channels expressed in the gonad, after interference with Ca2+ signaling, or after of pSTAT3 inhibitors. Experiments will (1) determine whether RBM20 is also a target of pSTAT3, or downstream of KDM6B, and (2) test the role of RBM20 using a knock-down approach. Splicoform variants will be analyzed at the two temperatures using the lab's new T. scripta genome assembly. Results will establish the molecular basis for temperature-dependent gonad determination, and may uncover a conserved role for splicing in gonad development. The lab engages students in science research through creation of lesson plans that incorporate cutting edge research into local classrooms, and programs that encourage girls to pursue STEM careers. The lab also brings this science to public attention through podcast interviews, contribution to Duke's ongoing Art of the Scientist exhibition, and Duke's science cafe' series.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

温度通过控制胚胎性腺发育为睾丸还是卵巢来调节许多爬行动物的雄性或雌性发育，包括红龟 Trachemys scripta elegans。然而，自从50年前发现温度影响性腺发育的机制以来，这一现象一直是一个谜。此前，卡佩尔实验室发现了两个基因，它们在雄性和雌性繁殖温度下的龟性腺中表现出强烈的表达差异。在这个项目中，实验室将确定这些基因的表达如何受温度控制，以及这些基因的活性如何控制睾丸和卵巢的发育。这项工作的结果将建立温度对男性或女性发育影响的机制。实验室积极利用该项目作为平台，吸引年轻学生和公众参与科学研究。该项目也是一个强有力的平台，旨在倡导以证据为基础的科学制定环境政策，以减轻气候变化的影响，预计会在温度影响性腺发育的物种中产生强烈的雄性/雌性比例偏差。Capel实验室此前表明，在雄性繁殖温度下，海龟性腺中表观遗传调节因子KDM6B和RNA剪接因子RBM20的转录水平升高。 KDM6B 通过去除 DMRT1（睾丸发育的保守调节因子）上的抑制性组蛋白标记来激活睾丸通路。在该项目中，该实验室计划确定将温度与 KDM6B 和 RBM20 转录和剪接变化联系起来的分子途径。该实验室假设，已知的 KDM6B 阻遏蛋白 pSTAT3 在雌性生产的高温度下通过激活 TRP 通道、Ca2+ 流入和 CAMK 的高活性而被差异磷酸化。为了检验这一假设，在性腺中表达的 TRP 通道被敲低后、在干扰 Ca2+ 信号传导后或在使用 pSTAT3 抑制剂后，将测量雄性和雌性通路的标记。实验将 (1) 确定 RBM20 是否也是 pSTAT3 或 KDM6B 下游的靶标，以及 (2) 使用敲低方法测试 RBM20 的作用。将使用实验室新的 T. scripta 基因组组装在两个温度下分析剪接变体。结果将为温度依赖性性腺测定奠定分子基础，并可能揭示剪接在性腺发育中的保守作用。该实验室通过制定课程计划（将前沿研究纳入当地课堂）以及鼓励女孩追求 STEM 职业的计划，让学生参与科学研究。该实验室还通过播客采访、为杜克大学正在进行的“科学家的艺术”展览以及杜克大学的科学咖啡馆系列做出贡献，让这门科学受到公众的关注。该奖项反映了 NSF 的法定使命，并通过使用基金会的智力价值和更广泛的影响审查标准进行评估，被认为值得支持。

项目成果

Temperature-dependent sex determination is mediated by pSTAT3 repression of Kdm6b

• DOI: 10.1126/science.aaz4165
• 发表时间: 2020-04
• 期刊: Science
• 影响因子: 56.9
• 作者: C. Weber;Yingjie Zhou;Jong Gwan Lee;L. Looger;Guoying Qian;C. Ge;B. Capel

The histone demethylase KDM6B regulates temperature-dependent sex determination in a turtle species

• DOI: 10.1126/science.aap8328
• 发表时间: 2018-05-11
• 期刊: SCIENCE
• 影响因子: 56.9
• 作者: Ge, Chutian;Ye, Jian;Capel, Blanche
• 通讯作者: Capel, Blanche

Sex determination without sex chromosomes

• DOI: 10.1098/rstb.2020.0109
• 发表时间: 2021-07
• 期刊: Philosophical Transactions of the Royal Society B
• 作者: C. Weber;B. Capel