裂殖酵母核糖体异质性的生物学功能和分子机制的研究

It is commonly thought that all ribosome particles in cells have the same biochemical composition, and carry out one activity of protein translation. However, such paradigm is challenged by the alternative hypothesis of ribosome heterogeneity, which postulates that ribosomes may have variable compositions and may differentially translate specific mRNAs. The biochemical nature of ribosome heterogeneity and the mechanisms underlying the differential functions of heterogeneous ribosomes remain largely unknown..Our preliminary investigation in the fission yeast have indicated that ribosome composition may vary depending on which of the rps8 paralogs (Rps801p or Rps802p) is incorporated into the 80S ribosome particle, which in turn, may influence whether Rpl42p is incorporated or not. .Here, we propose to test directly the existence of ribosome heterogeneity in fission yeast mediated by Rps8/Rpl42. We will further explore its functional significance by testing the possible anti-stress growth advantages conveyed by ribosome heterogeneity. Lastly, we seek to identify the target mRNAs that are differentially translated by heterogenous ribosomes..The success of these proposed work will for the first time establish the biochemical basis for ribosome heterogeneity and provide a comprehensive set of supporting evidence integrating results deriving from biochemical, genetic and high-throughput investigations.

传统上，人们认为细胞内所有的核糖体其生化组成都是固定的，功能也是单一的。但是随着在多种物种中陆续发现了核糖体异质性，这种观点正被逐渐改变。核糖体异质性学说认为，核糖体在组成和功能上都是可变的。然而，现在还不清楚这些异质核糖体的生化基础及其功能差异的分子机制。.我们在裂殖酵母中已经发现核糖体小亚基蛋白S8旁系同源蛋白Rps801p和Rps802p造成大亚基蛋白L42蛋白含量的差异，提出基于S8/L42核糖体异质性的工作模型。本项目将以此为研究对象，在裂殖酵母核糖体中直接验证这一工作模型，并且阐明S8/L42异质核糖体的生物学功能，最终探索异质核糖体差异性翻译的靶基因。.本项目的成功将第一次从生化水平上阐明核糖体异质性的组成并且为核糖体异质性提供生化、遗传等多学科综合的实验证据支持，为核糖体病变的解决和药物靶点的选择提供指导。

传统上，人们认为细胞内所有核糖体其生化组成固定，功能单一。这种观点越来越受到挑战，特别是在多种物种中陆续发现了“核糖体异质性”的模型。该模型提出功能性核糖体颗粒在其生化组成上可能存在差异，从而导致其生物学功能上的差异。然而，异质核糖体的生化基础及其功能差异的分子机制目前尚不清楚。通过对模式生物裂殖酵母S.pombe的研究，我们提供了广泛的遗传学证据，结果表明编码相同核糖体蛋白组分的旁系同源蛋白基因，其序列上可能仅有细微的差异，但是它们中的许多基因在功能上是不同的。以rps8旁系同源基因rps801和rps802为例，我们发现位于E位点附近的核糖体大亚基Rpl42p的水平在rps801和rps802基因缺失菌株中存在差异。进一步地研究表明，rps8旁系同源基因敲除菌株的表型差异归因于不同的Rpl42p水平，而不是Rps801p和Rps802p之间序列的细微差异。此外，我们还发现，其他三组40S核糖体蛋白旁系同源基因敲除也是通过在完整核糖体中Rpl42p水平的差异，与rps8旁系同源基因敲除一样，表现出类似的表型差异。综上所述，我们的工作发现了一种以Rpl42p水平差异作为常见形式的核糖体异质性，提示核糖体E位点可能因多种刺激而表现出多样化的功能。