YEAST RNA VIROLOGY

酵母RNA病毒学

• 批准号: 6161903
• 负责人: Reed B. WICKNER
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6161903
• 关键词: RNA virus; double stranded RNA; gene deletion mutation; gene mutation; genetic translation; messenger RNA; molecular cloning; nucleic acid sequence; virus RNA; virus genetics; virus protein; virus replication

We have described two dsRNA viruses (L-A and L-BC) and two ssRNA replicons (20S RNA and 23S RNA) in the yeast Saccharomyces cerevisiae. M dsRNA is a satellite of L-A encoding the killer toxin. We discovered 7 chromosomal genes, SKI1, 2, 3, 4, 6, 7, and 8, by their ability to prevent these replicons from causing pathogenicity to yeast cells. These four RNA replicons all make uncapped mRNAs and lack a 3' poly(A) structure. SKI1 is an exoribonuclease specific for uncapped mRNAs, while we showed that the SKI2, SKI3 and SKI8 gene products block the translation of non-poly(A) mRNAs. We showed that mutations in 20 chromosomal genes resulting in loss of M dsRNA are deficient in 60S ribosomal subunits. These mutations are suppressed by ski mutations without restoration of the 60S subunit deficiency. We proposed that SKI2, SKI3 and SKI8 block translation of non-poly(A) mRNA by an effect on ribosome biogenesis affecting the interaction of 60S subunits with the 3' poly(A) structure of the mRNA. We have now cloned the SKI6 and SKI7 genes. Ski6p has homology with RNase PH, an enzyme that removes 3' nucleotides from tRNA precursors in a phosphorolytic reaction (like polynucleotide phosphorylase). Ski6-2 mutants are hypersensitive to hygromycin, an indication of involvement of Ski6p in translation. These ski6-2 strains translate non-poly(A) mRNAs 10x better than isogenic wild type cells. They also accumulate a 38S particle with a partially degraded 25S rRNA, providing direct evidence for altered 60S subunits as the cause of the derepressed translation of non-poly(A) mRNAs. Ski7p is highly homologous to translation factor EF1alpha. The ski7 mutants are derepressed for translation of non-poly(A) mRNAs and overexpression of Ski7p results in loss of M dsRNA.

我们描述了两个DSRNA病毒（L-A和L-BC）和两个ssRNA 酿酒酵母中的复制子（20S RNA和23S RNA）。 M dsRNA是L-A编码杀手毒素的卫星。 我们发现了 7染色体基因，Ski1、2、3、4、6、7和8，其能力 防止这些复制子引起致病性对酵母细胞。 这四个RNA复制子都形成了未盖的mRNA，并且缺乏3'poly（a） 结构。 SKI1是针对非封闭mRNA的驱齿核酸酶， 虽然我们表明SKI2，SKI3和SKI8基因产品阻止了 非poly（a）mRNA的翻译。 我们证明了20种突变 导致M DSRNA损失的染色体基因在60s中缺乏 核糖体亚基。 这些突变被滑雪突变抑制 没有恢复60年代亚基缺乏症。 我们提出了这一点 SKI2，SKI3和SKI8块的非poly（a）mRNA的翻译通过效果 关于核糖体生物发生，影响60s亚基与 mRNA的3'poly（a）结构。 我们现在已经克隆了Ski6和 SKI7基因。 SKI6P与RNase PH具有同源性，该酶可去除 在磷酸反应中的tRNA前体的3'核苷酸（如 多核苷酸磷酸化酶）。 SKI6-2突变体对 湿霉素，Ski6p参与翻译的指示。 这些 SKI6-2菌株比同生野生菌株翻译非poly（a）mRNA 10x 类型单元格。 他们还积累了一个38S粒子 降级25S rRNA，提供了更改60s亚基的直接证据 作为非poly（a）mRNA的过压缩翻译的原因。 Ski7p 与翻译因子EF1alpha高度同源。 SKI7突变体 被解压缩，以翻译非poly（a）mRNA和过表达 SKI7P的损失。