Molecular genetics underlying reproductive isolation of species

物种生殖隔离的分子遗传学

• 批准号: RGPIN-2014-06340
• 负责人: Lasko, Paul
• 金额: $ 5.17万
• 依托单位: McGill University
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2016
• 资助国家: 加拿大
• 起止时间: 2016-01-01 至 2017-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=612171
• 关键词: Molecular; genetics; underlying; reproductive; isolation

The research in this proposal will use fruit flies of the Drosophila genus to investigate the molecular mechanisms that ensure that interspecific hybrids cannot reproduce. More specifically, we will investigate the hypothesis that Vasa, a rapidly-evolving protein that is expressed in germ cells during oogenesis, and that is required maternally for specification of primordial germ cells in progeny embryos, is an important player in limiting the fertility of interspecific hybrids. Eukaryotic genomes including those of Drosophila contain many copies of transposable elements (TEs), whose mobility has to be suppressed in order to maintain chromosome integrity. Suppression of TE transposition involves small RNAs called Piwi-interacting RNAs (piRNAs) that target TE-derived mRNAs, and subsequently destabilize them and repress their translation. In interspecific Drosophila hybrid females, TEs are derepressed, raising the possibility that this system has a fundamental role in establishing the reproductive isolation of species. If this is so, then it would be expected that the responsible protein or proteins would evolve very rapidly, so that a protein from one species would be sufficiently different to not function in a different species. Vasa, which has recently been implicated in piRNA processing, has regions that diverge extensively, even between closely related Drosophila species such as D. melanogaster and D. simulans. To examine the relationships between Vasa, piRNA mediated processes, and reproductive isolation, we will first identify which domains of Vas are required for piRNA-mediated processes by analyzing expression of TE-encoded genes, and piRNA expression, in females that only express one of a set of site-specific mutated forms of Vasa. Next, we will examine the extent to which Vasa orthologues from other species can function in D. melanogaster, by introducing these genes transgenically into a vasa mutant background. Finally, based on the results we obtain we will replace various domains of Vasa from non-rescuing species with D. melanogaster counterpart domains to attempt to restore function. This will enable us to map particular domains of Vasa that must be derived from the host species (in this case, melanogaster) for functionality. In the longer term extending beyond the scope of the present proposal we will investigate other piRNA-effector genes in a similar way. We will combine mutational analysis, molecular analysis, and protein structure information produced by others to understand particular molecular associations underlying reproductive isolation. For example, we will test whether a particular critical segment of Vasa interacts with a particular partner protein in a species-specific way. The expected impact of this work will include publications in high-quality international scientific journals, and the trainees funded by the grant (as well as the PI) will present their results at national and international conferences. The trainees will also complete PhD thesis projects that will have involved gaining facility in state-of-the-art molecular genetic and imaging techniques, which will enable them to pursue research careers in either the public or private sectors.

这项建议中的研究将使用果蝇属的果蝇来研究确保种间杂交不能繁殖的分子机制。更具体地说，我们将调查这样一种假设，即Vasa是一种快速进化的蛋白质，在卵子发生期间在生殖细胞中表达，是后代胚胎中原始生殖细胞的母性指定所必需的，是限制种间杂交育性的重要因素。 包括果蝇在内的真核生物基因组中含有许多转座元件(TES)的拷贝，为了保持染色体的完整性，必须抑制TES的迁移率。抑制TE转座涉及到称为Piwi相互作用RNAs(PiRNAs)的小RNA，它以TE衍生的mRNA为靶点，随后破坏它们的稳定并抑制它们的翻译。在种间杂交雌性果蝇中，TES被去抑制，这增加了这一系统在建立物种生殖隔离中具有基础作用的可能性。如果是这样，那么预计负责的一个或多个蛋白质将进化得非常快，因此来自一个物种的蛋白质将有足够的差异，不会在不同的物种中发挥作用。 VASA最近被牵连到piRNA加工过程中，它的一些区域存在着广泛的差异，甚至在近亲关系密切的果蝇物种之间也是如此，比如黑腹果蝇和拟果蝇。 为了研究VASA、piRNA介导的过程和生殖隔离之间的关系，我们首先将通过分析TE编码基因的表达和piRNA在只表达一组VASA位点特异性突变形式之一的雌性动物中的表达，来确定哪些Vas结构域是piRNA介导的过程所必需的。接下来，我们将通过将来自其他物种的Vasa同源基因转基因引入到Vasa突变背景中，来研究这些基因在黑腹毛虫中发挥作用的程度。最后，基于我们得到的结果，我们将用黑腹果蝇的对应结构域替换来自非救援物种的VASA的各个结构域，以尝试恢复功能。这将使我们能够定位VASA的特定结构域，这些结构域必须来自宿主物种(在这种情况下，是黑腹蛇)才能发挥功能。 从长远来看，超出本提案的范围，我们将以类似的方式研究其他piRNA效应基因。我们将结合突变分析、分子分析和其他人产生的蛋白质结构信息来了解生殖隔离背后的特定分子关联。例如，我们将测试VASA的特定关键片段是否以物种特有的方式与特定的伙伴蛋白相互作用。 这项工作的预期影响将包括在高质量的国际科学期刊上发表文章，由赠款资助的受训人员(以及国际和平研究所)将在国内和国际会议上介绍他们的成果。受训人员还将完成博士论文项目，这些项目将涉及获得最先进的分子遗传和成像技术，这将使他们能够在公共或私营部门从事研究事业。