Eukaryotic tRNA Maturation

描述（由申请人提供）： 核糖核酸酶P（RNase P）是在所有生物体中发现的仅有的两种含有催化活性RNA亚基的酶之一。该酶的主要已知功能是切割前体tRNA转录物以产生成熟的5'末端，但已知其在体内也具有几种其他底物。在真核细胞核中，这种酶已经进化成一种复杂得多的核糖核蛋白，其蛋白质含量是细菌酶的20倍。这种酶的进化过程是特别有趣的，因为真核RNA亚基已经失去了作为核酶的能力，在蛋白质的“辅因子”的情况下，即使结构和关键残基在RNA亚基中是保守的。本实验室以前的研究已经建立了酿酒酵母核RNase P的亚基组成，并表明它是细胞核中早期前tRNA加工的空间有序途径的一部分。RNase P亚基的生物化学和遗传学研究揭示了它们之间以及与其他细胞组分之间的广泛相互作用。我们的研究结果表明，相对于细菌和细胞器的核RNase P的亚基复杂性的大幅增加，可能需要正确的定位和时间的RNase P切割在高度有序的核环境，以及潜在的RNA底物之间的歧视。本提案中描述的实验调查了为什么真核生物RNase P全酶的复杂性增加是正确功能所必需的。

DESCRIPTION (provided by applicant): Ribonuclease P (RNase P) is one of only two enzymes containing catalytically active RNA subunits that are found in all living organisms. The primary known function of the enzyme is to cleave precursor tRNA transcripts to give mature 5' termini, but it is known to also have several other substrates in vivo. In eukaryotic nuclei the enzyme has evolved to be a far more complex ribonucleoprotein, having twenty times more protein than the bacterial enzyme. The evolutionary progression of this enzyme is particularly interesting, since the eukaryotic RNA subunit has lost the ability to act as a ribozyme in the absence of the protein "cofactors", even though the structure and key residues are conserved in RNA subunit. Previous studies in this lab have established the subunit composition of the Saccharomyces cerevisiae nuclear RNase P and have shown that it is part of a spatially ordered pathway for early pre-tRNA processing in the nucleus. Biochemical and genetic investigations of the RNase P subunits have revealed extensive interactions with each other and with other cellular components. Our results suggest that the large increase in subunit complexity of the nuclear RNase P relative to bacteria and organelles might be required for correct positioning and timing of RNase P cleavage in the highly ordered nuclear environment, as well as for discriminating among potential RNA substrates. Experiments described in this proposal investigate why the increased complexity of the eukaryotic RNase P holoenzyme is necessary for proper function.