The Control of Gene Expression by Eukaryotic RNase III

描述（由申请人提供）：转录后调控已成为控制定量和定性基因表达的有效方法。RNase III家族双链RNA内切酶在多种基因表达途径中起着至关重要的作用：它们参与核糖体RNA前体的加工，参与剪接和rRNA代谢的小RNA的加工，以及小RNA的加工。它们还参与RNA干扰过程中小干扰RNA的产生。我们的长期目标是：(i)详尽地鉴定由真核生物RNase III家族内切酶成员控制的基因表达途径，（ii）了解这些酶结合和切割dsRNA的机制，（III）了解这些酶的活性如何整合到细胞代谢中。利用酿酒酵母Rntlp酶作为真核生物RNase III酶的模型和功能基因组学，我们将研究由该酶控制的新的基因表达途径。Rntlp在监视和调节编码铁摄取蛋白的mrna和参与蛋氨酸回收途径的蛋白质中的功能将被研究。将开发新的基因组策略来鉴定真核生物RNases III调节的其他基因表达途径。利用位点定向诱变和RNA底物的位点特异性修饰，研究rnt1p对双链RNA的识别和切割位点选择机制。RNase III活性在细胞环境中的整合将被研究。特别是，我们将破译RNase III被共转录募集到其底物的机制，以及RNase III在低铁条件下下调的机制。这些研究将有助于我们了解RNase III在基因表达控制中的作用以及真核细胞代谢对缺铁的适应机制。

DESCRIPTION (provided by applicant): Post-transcriptional regulation has emerged as a potent way to control quantitatively and qualitatively gene expression. The RNase III family of double-stranded RNA endonucleases plays crucial functions in multiple gene expression pathways: they participate in the processing of precursors of ribosomal RNA, of small RNAs involved in splicing and rRNA metabolism, and of microRNAs. They also participate in the production of small interfering RNAs in the RNA interference process. Our long term goals are (i) to exhaustively identify the gene expression pathways controlled by eukaryotic members of the RNase III family of endonucleases, (ii) to understand the mechanisms by which these enzymes bind and cleave dsRNA and (iii) to understand how the activity of these enzymes is integrated in the cell metabolism. Using the S.cerevisiae enzyme Rntlp as a model eukaryotic RNase III enzyme and functional genomics, we will investigate novel gene expression pathways controlled by this enzyme. The functions of Rntlp in the surveillance and regulation of mRNAs encoding iron uptake proteins and for a protein involved in the methionine salvage pathway will be investigated. Novel genomic strategies will be developed to identify additional gene expression pathways regulated by eukaryotic RNases III. The recognition of double-stranded RNA and the mechanism of cleavage site selection by Rnt1 p will be studied using site directed mutagenesis and site-specific modifications of the RNA substrate. The integration of RNase III activity in the cellular context will be investigated. In particular, we will decipher the mechanism by which RNase III is recruited co-transcriptionally to its substrates, and the mechanism by which RNase III is down-regulated in low iron conditions. These studies will contribute to our understanding of the roles of RNase III in the control of gene expression and of the mechanisms of adaptation of eukaryotic cell metabolism to iron deficiency.