DIVERSITY IN THE NUMBERS,TYPES AND SEQUENCES OF TRNA'S

蛋白质合成依赖于一个复杂的细胞机器，包括转移RNA（tRNA），氨酰-tRNA合成酶，起始，延伸和释放因子，以及核糖体RNA（rRNA）和核糖体蛋白。 由于翻译是一个必不可少的细胞过程，强烈的稳定选择预计会影响其组成部分。 然而，来自基因组测序项目的结果表明，尽管翻译的一般过程在整个进化过程中是保守的，但生物体以许多不同的方式保持准确和有效的蛋白质合成。特别是，生物体内部和之间在数量和类型以及其tRNA的序列和结构特征方面存在意想不到的差异。 因此，过渡器官的弹性显然比通常认为的要大。 所提出的实验是出于这些观察，并旨在回答三个相关的问题。是什么机制导致了同功受体之间的序列变异？ 是什么机制使生物体在进化过程中失去“必需的”tRNA基因？ 细胞可以耐受的tRNA功能受损的程度是多少？拟议中的研究使用生物化学和遗传学方法来回答这些问题。 它将检查（i）反密码子在tRNA功能和进化中的作用;（ii）补偿tRNA基因丢失的机制;以及（iii）细胞活力和生长所必需的tRNA功能水平。 这些结果将阐明tRNA中决定其在蛋白质合成过程中氨基酸接受和捐赠功能的元素。此外，他们将阐明影响这些活动的tRNA的翻译装置的属性。因此，这些结果将与由于细胞中的环境而从单个基因型表达不同表型的程度的一般问题有关。 理解这一现象对于解释DNA序列信息和发展已经提出的通过操纵翻译机制来控制细菌感染的一些策略是至关重要的。

Protein synthesis depends on a complex cellular machine comprised of transfer RNA (tRNA), aminoacyl-tRNA synthetases, initiation, elongation, and release factors, as well as ribosomal RNA (rRNA) and ribosomal proteins. Because translation is an essential cellular process, strong stabilizing selection is expected to influence its components. However, results from the genome sequencing projects show that although the general process of translation is conserved throughout evolution, organisms maintain accurate and efficient protein synthesis in many different ways. In particular, there are unexpected differences within and among organisms in the numbers and types as well as in the sequence and structural features of their tRNAs. Thus there is apparently more resiliency in the transitional apparatus than is typically thought. The proposed experiments are motivated by these observations and are designed to answer three related questions. What are the mechanisms that account for sequence variation among isoacceptors? What are the mechanisms that allow organisms to lose "essential" tRNA genes during evolution? What is the extent of impairment in tRNA function that can be tolerated by the cell? The proposed research uses biochemical and genetic approaches to answer these questions. It will examine (i) the role of the anticodon in tRNA function and evolution; (ii) the mechanisms that compensate for tRNA gene loss; and (iii) the levels of tRNA function that are necessary for cell viability and growth. The results will elucidate elements in tRNAs that dictate their amino acid accepting and donating functions during protein synthesis. In addition, they will elucidate the properties of the translational apparatus that influence these activities of tRNAs. Thus the results will be pertinent to the general problem of the extent to which different phenotypes are expressed from a single genotype due to circumstances in the cell. An understanding of this phenomenon is critical to the interpretation of DNA sequence information and to the development of some strategies that have been proposed for controlling bacterial infections by manipulating the translation machinery.