Doctoral Dissertation Research: Cold adaptation and gene expression

在过去的约45万年里，人属的成员多次从赤道的非洲迁徙到更季节性和更寒冷的气候。非颤抖的产热作用--棕色脂肪产生热量的过程--可能是我们物种成功适应这些环境并帮助促进这些地理扩张的关键部分。UCP1基因的产物，一种被称为热原的蛋白质，推动了这一过程。在过去的大约30万年里，非人灵长类动物也从非洲扩张到了更寒冷的纬度。此前，研究人员在一些非人类灵长类动物中发现了这种基因调控的选择信号。在这项博士论文研究项目中，研究人员通过使用创新的方法和先前收集的样本来验证和扩展这些发现。来自STEM学科代表性不足群体的学生通过这个项目接受培训和专业发展。使用一种新针对特定物种验证的既定方案，研究人员将存档的野生非人灵长类成纤维细胞系化学转化为化学诱导的棕色脂肪(Ciba)细胞。转化结果通过RT-qPCR定量UCP1(在棕色脂肪中唯一表达)的RNA表达，油红O染色和免疫荧光染色观察脂滴的形成和培养转化的完整性来验证。然后将Ciba暴露于去甲肾上腺素以刺激细胞系的冷反应，并使用批量RNA测序比较有无冷反应的UCP1基因表达。结果被用来检验在四个基因座上UCP1表达的派生等位基因剂量效应的预测，使得以前发现在较冷的气候中出现频率较高的每个派生等位基因将与UCP1表达的显著增加相关。分析考虑了潜在的协变量，如性别、BMI和体重。研究人员的验证先前的结果表明，NST系统在非人类灵长类动物中经历了积极的选择，以应对较冷的气候，并提供了我们对这一过程的理解。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

In the past ~450,000 years, members of the genus Homo migrated multiple times from equatorial Africa to more seasonal and cold climates. Non-shivering thermogenesis – a process by which brown fat generates heat – was likely a key part of the success of our genus in adapting to these environments and helping to facilitate these geographic expansions. The product of the gene UCP1, a protein called thermogenin, drives this process. Nonhuman primates have also expanded from Africa into colder latitudes over the past ~300,000 years. Previously, the investigators found signals of selection in the regulation of this gene in some nonhuman primates. In this doctoral dissertation research project, the investigators validate and expand those findings by using innovative methods and previously collected samples. Students from groups underrepresented in STEM disciplines receive training and professional development through this project.Using an established protocol newly validated for specific species, the investigators chemically convert archived wild nonhuman primate fibroblast cell lines into chemically induced brown adipose (ciBA) cells. Outcomes of the conversion are verified by quantifying RNA expression of UCP1 (which is uniquely expressed in brown adipose) via RT-qPCR, with Oil Red O staining and immunofluorescence staining to observe lipid droplet formation and the completeness of culture conversion. The ciBA are then exposed to norepinephrine to stimulate a cold response in the cell lines, and UCP1 gene expression with and without the cold response are compared using bulk RNA sequencing. Results are used to test the prediction of a derived allele dosage effect in UCP1 expression across four loci, such that each derived allele, previously found to have appeared at higher frequencies in colder climates, will be associated with a significant increase in UCP1 expression. Analyses consider potential covariates like sex, BMI, and body weight. Validation of the investigators previous results suggest that the NST system has undergone positive selection in response to colder climates in nonhuman primates and inform our understanding of this process in our own genus.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.