拟南芥tRNA甲基转移酶亚基AtTRM61调控早期胚胎与胚乳发育的分子机制

RNA modification is an emerging research area. The transfer RNA (tRNA) is an important component of RNA modification. In yeast and human cells, the m1A58 methylation modification of the initiated tRNA, which is carried out by AtTRM6/61, has been proved that it’s essential for cell viability. However, its functions in individual development are still unclear, especially in plant reproductive development. The previous results in this research showed that in trm61 mutants, there were severe defects in early embryo and endosperm development, and AtTRM61 can bind to AtTRM6 and the protein kinase AtMPK4 directly. These results suggested that the AtTRM6/61 complex is essential for seed development, and that AtMPK4 may work together with AtTRM61 in the regulation of initiated tRNA modification. Here, we will study the expression pattern of AtTRM61, and according to construct a partially complemental mutant using the tissue-specific promoters, we will further study the biological functions of AtTRM61 in the regulation of embryo and endosperm development. Moreover, by yeast library screening and genetic interaction assays, we will establish the molecular network of AtTRM61 regulated expression of initiated tRNA and the protein synthesis. This would shed light on the study of the functions of tRNA modification in plant reproductive development.

RNA修饰是一个新兴的热门领域，转运RNA（tRNA）修饰是其中重要的组成部分。在酵母与人类中，tRNA甲基转移酶TRM6/61参与的起始tRNA m1A58甲基化修饰，对细胞活力的维持具有重要作用，但其在个体发育中的功能，尤其是植物生殖发育中的功能并不清楚。预实验结果显示：拟南芥trm61突变体早期胚胎与胚乳败育，且AtTRM61可同时结合AtTRM6与蛋白激酶AtMPK4，暗示AtTRM6/61为种子发育必需，且AtMPK4可能参与AtTRM61对起始tRNA修饰的调节。本项目将进一步研究AtTRM61的表达，利用胚胎与胚乳特异表达启动子获得部分回复的纯合突变体，阐明AtTRM61调控胚胎与胚乳发育的生物学功能；同时，通过酵母文库筛选与遗传互作分析等手段，探索AtTRM61调控起始tRNA表达与蛋白质翻译的分子网络机制。这将为解析tRNA修饰在植物生殖发育过程中的功能提供重要理论基础。

RNA修饰是一个新兴的热门研究领域，转运RNA（tRNA）修饰是其中重要的组成部分。在tRNA甲基转移酶TRM6/61的作用下，tRNA发生m1A58修饰，该修饰在大多数真核生物中都十分保守。尽管如此，在生物学功能方面，该复合物仅在细胞水平进行了探究，而在个体发育中的功能，尤其是植物生殖发育中的功能并不清楚。本研究以TRM6/61的T-DNA插入突变体为基础，观察了其功能缺失对拟南芥生殖发育的影响，发现该复合体参与了种子发育和配子发育的调节，并且都在早期发育阶段就表现出显著的缺陷。本研究还从基因时空表达模式的角度探究了其发挥功能的基础。而为了探究其发挥作用的机理，本研究通过分子生物学和遗传学等手段，利用特异性启动子驱动的蛋白表达，进一步在个体水平上验证了AtTRM6/61复合体通过影响起始tRNA的表达水平进而影响生殖发育的过程。在探究其中更复杂的分子调控网络过程中，研究发现了包括MPK4、L18e以及UAP56B等在内的若干蛋白均能与TRM61发生直接的互作，这种相互作用可能在调节基因转录、蛋白质翻译方面起到重要作用。本研究一方面揭示了AtTRM6/61复合体在植物生殖发育调节过程中的重要作用，另一方面也从机制层面解析了其通过甲基转移酶活性调节起始tRNA的甲基化修饰进而影响其转录表达的功能，并且在更为复杂的调节网络机制研究中提供了一定的方向，为解析该复合物在生命体中的重要作用提供了更加丰富的理论基础。