Adaptive Protein Evolution of the Drosophila Germline Stem Cell Gene Bag of Marbles

项目摘要 动物含有少量的生殖干细胞（GSC），这些细胞对于制造精子或卵子至关重要。 决定一个分裂的干细胞更新自己或分化，并调节早期GSC分裂 是发展中最关键的过程。对果蝇的研究，黑腹果蝇， 确定了许多参与GSC维持和分化的细胞信号传导的关键基因。我们 已经表明，大理石基因袋（bam），GSC分化的关键“开关”，是适应性进化的 在D. melanogaster、黑腹果蝇D. simulans、D.日本料理，但不是在六 其他相关物种。在我们目前的资助期间，我们已经产生了新的bam无效突变， D. melanogaster、黑腹果蝇D. simulans，拟谷盗D. yakuba，尖吻隐翅虫D. teissieri和D. ananassae测试 功能守恒。这使我们能够证明，令人惊讶的是，与D。黑腹动物， bam对雌性D.也不是雄性GSC分化所必需的 in D. teissieri和D. ananassae。我们已经证明，感染不同菌株的细胞内 内共生体Wolbachia pipientis在挽救bam的生育缺陷特征方面的程度各不相同 亚型这项建议的重点是确定所观察到的功能和进化后果， bam的序列多样性，并区分bam在GSC功能中的作用 已被选定为新的功能，与假设之间的相互作用bam和W。皮皮恩蒂斯河 是推动巴姆变革的进化力量这包括生成和分析新的bam D. melanogaster和D. simulans，并测试这些新的hypomorphs与intra 和种间沃尔巴克氏体菌株。我们将对W. pipientis。我们 正在开发一个计算模型，以产生实验可检验的序列预测， 在W. pipientis和GSC基因。最后我们将 在功能上评估D. melanogaster和D. simulans的雌、雄配子体发育和育性对bam的需求消失。 teissieri与近缘种D.日本料理。我们学到的东西将大大有助于我们的 了解GSC在不同物种发育过程中的调节方式， 发育功能可以进化，以及它们如何由于突变和与 生殖系寄生虫可导致不育、生殖系癌症和生殖隔离。

PROJECT SUMMARY Animals contain a small number of germline stem cells (GSCs) that are essential for making sperm or eggs. The decision for a dividing stem cell to renew itself or to differentiate, and the regulation of early GSC divisions are some of the most critical processes in development. Studies in the fruit fly, Drosophila melanogaster, have identified many of the key genes involved in the cellular signaling for GSC maintenance and differentiation. We have shown that the gene bag of marbles (bam), the key “switch” for GSC differentiation, is adaptively evolving with a dramatic excess of amino acid substitutions in D. melanogaster, D. simulans, D. yakuba but not in six other related species. During our current funding period we have generated new bam null mutations in sequenced genomes of D. melanogaster, D. simulans, D. yakuba, D. teissieri, and D. ananassae to test for conservation of function. These have allowed us to demonstrate that surprisingly unlike in D. melanogaster, bam is not required for female GSC differentiation in D. teissieri nor is it required for male GSC differentiation in D. teissieri and D. ananassae. We have shown that infection with different strains of the intracellular endosymbiont Wolbachia pipientis vary in their degree of rescue of the fertility defects characteristic of the bam hypomorph. This proposal focuses on identifying the functional and evolutionary consequences of observed sequence diversification at bam and discriminating between the hypothesis that bam’s role in GSC function has been selected for new functions, vs the hypothesis that interactions between bam and W. pipientis has been the evolutionary force driving changes at bam. This includes generating and analyzing novel bam hypomorphs in D. melanogaster and D. simulans, and testing interactions of these new hypomorphs with intra and interspecific Wolbachia strains. We will carry out RNA-seq studies of the bam rescue by W. pipientis. We are developing a computational model to generate experimentally testable predictions of sequence diversification under several mechanisms of interaction between W. pipientis and GSC genes. Finally, we will functionally evaluate evolutionary diversification of the bam protein sequence between D. melanogaster and D. simulans, and the loss of the requirement for bam for female and male gametogenesis and fertility in D. teissieri compared to its close relative D. yakuba. What we learn will contribute significantly to our understanding of how GSCs are regulated during development in diverse species, how such critical developmental functions can evolve, and how their mis-regulation due to mutation and interaction with germline parasites can lead to infertility, germline cancers, and reproductive isolation.