线粒体在多倍体鲫鲤核质冲突中的相关作用

The allotetraploid fish population was established first time in the world at the Laboratory of Fish Development of Hunan Normal University, which is fertile for both sexes and owns high hereditary stability. A unique diploidy/triploidy/tetraploidy fish study platform is formed basing on the allotetraploid fish, diploid crucian carp and their triploid hybrid-Xiangyun crucian carp. It is reported that mitochondria plays a major role in the formation of interpopulation hybrids,but the mechanism of cytonuclear conflict in this interpopulation hybrid fish system is unknown. In this proposal, based on the diploidy/triploidy/tetraploidy fish system, we plan to study the role of mitochondrial function in the spermatocyte and oocytes. The research design includes: ultrastructural detection of mitochondria, determination of mtDNA copy number, in vitro measurement of ATP production, measurement of OXPHOS-associated enzyme complex activity and expression of mitochondrial transcription factor A(TFAM) genes in the diploidy/ triploidy/ tetraploidy fish. This study will elucidate the role of mitochondrial function involved in the cytonuclear conflict of meiotic cell in the diploidy/triploidy/tetraploidy fish, which will built a solid foundation for the further study on the mechanism of the formation of interpopulation hybrid fish.

世界首例两性可育的异源四倍体鲫鲤群体在湖南师范大学鱼类发育实验室成功获得。异源四倍体鲫鲤、其二倍体父/母本和三倍体杂交后代组成了一个倍性明确、亲缘关系清晰的多倍体杂交鱼研究体系。最近的研究表明，线粒体在种间杂交物种核质冲突中起着重要作用。而在种间杂交鱼类核质冲突中线粒体相关作用的研究还是一片空白。本项目设想通过利用已成功建立的多倍体鲫鲤杂交鱼体系，采取电子显微镜观察、免疫荧光标记、荧光素酶标和荧光定量PCR分析等实验手段，对比分析不同倍性鲫鲤鱼精/卵母细胞线粒体的形态结构、活性和mtDNA拷贝数，以及mtDNA复制调控因子TFAM的表达特征。揭示多倍体鲫鲤杂交鱼核质冲突过程中线粒体相关作用的机制，为系统研究种间杂交形成多倍体鱼类的分子调控机制打下坚实基础。

研究表明线粒体参与的核质冲突在种间杂交物种形成的过程中起着重要作用，但有关种间杂交鱼类核质冲突中线粒体相关功能作用的研究还很少。本项目在制备了以异源四倍体鲫鲤、其二倍体父/母本和三倍体杂交后代组成的遗传背景明确、亲缘关系清晰的多倍体鱼研究体系的基础上。对不同倍性鱼类线粒体的形态结构、线粒体DNA拷贝数和线粒体活性的进行了检测，并对mtDNA复制调控因子TFAM进行了克隆和表达分析，还从转录组水平分析了异源四倍体鲫鲤及其二倍体父母本的核基因组和线粒体基因组的表达。研究发现异源四倍体鲫鲤之所以能够存活，是因为核基因组和线粒体基因组相互调整适应达到了一个新的平衡；研究发现在杂交鱼后代中线粒体拷贝数与雌雄性生殖配子的形成和质量都相关；在研究中我们还发现，鱼类杂交过程中来自父本的线粒体也会对杂交后代的生存有着重要的影响；此外，根据线粒体研究的前沿进展，本项目还进一步开展了杂交鱼后代父源性线粒体的行为研究。本项目的研究结果揭示了多倍体鲫鲤杂交鱼核质冲突过程中线粒体的部分作用机制，为系统研究种间杂交形成多倍体鱼类的分子机制打下坚实基础。