Factors Influencing Genetic Transcription Initiation And Termination

RNA形成部分的研究旨在理解和利用细胞DNA复制过程，HIV复制与这些细胞事件的关系，以及如何将这些信息用于治疗目的。我们正在研究DNA复制或转录过程中形成的RNA/DNA杂交体的形成和分解。核糖核酸酶H是参与去除RNA/DNA杂合体的RNA的重要酶，并且在该病毒的RNA基因组转化为DNA期间，与细胞中的DNA复制和HIV复制密切相关。细胞的RNA酶H和HIV共享利用类似蛋白质结构切割RNA的共同酶机制。改变特定疾病相关基因水平的药物正在开发中，以利用细胞内的RNase H。调节RNases H的表达可以增强药物的疗效。在这些努力中采用分子遗传学、生物化学和小鼠动物模型。 在转录过程中，RNA/DNA杂合体可以形成，并且通常被RNase H切割。RNase H通过与RNA的多次接触和与DNA链的相互作用识别并特异性切割杂交体的RNA，使用与Wei Yang博士（NIDDK）合作确定的双金属离子机制。 我们现在知道，真核生物的RNA酶H比细菌的酶更复杂，并且小鼠RNA酶H1（可能还有其他真核生物的RNA酶H1）的酶活性在与RNA/DNA杂交体结合时形成复合物，使得该酶在降解某些类型的RNA/DNA杂交体时更有效。我们与NCI-Frederick的研究小组合作，一直在研究RNase H的抑制剂，这些抑制剂可能被证明在HIV-AIDS治疗中有用。从我们早期的工作表明，小鼠RNase H1对发育至关重要，我们知道，重要的是要有专门针对艾滋病毒-艾滋病RNase H而不是细胞酶的药物。潜在药物的筛选现在包括使用我们实验室分离的人类RNase H1。 AGS是一种综合征，其原因已被指定为人类RNA酶H2的三个亚基中的任何一个的突变。 我们对酿酒酵母同源酶的研究首次描述了真核生物RNA酶H2的三个亚基组成。 在S.酿酒酵母不会导致任何主要的表型，除非突变体与其他几种基因缺失中的任何一种结合，这种情况称为合成致死。我们目前正在研究人类和沙门氏菌的生化特性。研究人员正在研究RNase H2在DNA复制、修复和重组中的作用。

Research in the Section on Formation of RNA is directed toward understanding and utilization of processes involved in cellular DNA replication, the relationship of HIV replication to these cellular events, and how to use this information for therapeutic purposes. We are examining the formation and resolution of RNA/DNA hybrids formed during DNA replication or transcription. Ribonucleases H are important enzymes participating in removal of the RNA of the RNA/DNA hybrids and are intimately related to DNA replication in cells and in HIV replication, during the conversion of the RNA genome of this virus to DNA. RNases H of cells and HIV share common enzymatic mechanisms of cleavage of RNA utilizing similar protein architectures. Drugs to alter levels of specific disease-related genes are being developed to take advantage of RNases H within the cell. Regulated expression of RNases H could enhance the efficacy of the drugs. Molecular genetic, biochemical, and mouse animal models are employed in these efforts. During transcription, RNA/DNA hybrids can form and are usually cleaved by RNase H. RNase H recognizes and specifically cleaves RNA of hybrids by several contacts to the RNA and interactions with the DNA strand using a two metal ion mechanism as determined in collaboration with Dr. Wei Yang (NIDDK). We now know that the eukaryotic RNases H are more complex than the bacterial enzymes, and that the enzymatic activity of mouse RNase H1 (and presumably other eukaryotic RNases H1) form a complex when binding to an RNA/DNA hybrid that enables the enzyme to be more efficient when degrading certain types of RNA/DNA hybrids. In collaboration with the groups at NCI- Frederick we have been examining inhibitors of RNases H that could prove useful in HIV-AIDS therapy. From our earlier work showing that the mouse RNase H1 is critical for development, we know that it is important to have drugs that specifically target the HIV-AIDS RNase H not the cellular enzyme. Screening of potential drugs now includes use of the human RNase H1 isolated in our laboratory. AGS is a syndrome whose cause has been assigned to mutations in any of the three subunits of human RNase H2. Our work on the homologous enzyme from Saccharomyces cerevisiae was the first describing the three subunit composition of eukaryotic RNases H2. Deletion of any genes encoding the three subunits of RNase H2 in S. cerevisiae does not result in any major phenotype unless the mutant is combined with any of several other gene deletions, a condition known as synthetic lethality. We are currently examining the biochemical properties of human and S. cerevisiae RNase H2, and, in particular, are examining the role of RNase H2 in DNA replication, repair and recombination.