CLONING OF THE RNA8-GENE OF SACCHAROMYCES-CEREVISIAE, DETECTION OF THE RNA8-PROTEIN, AND DEMONSTRATION THAT IT IS ESSENTIAL FOR NUCLEAR PRE-MRNA SPLICING
CLONING OF THE RNA8-GENE OF SACCHAROMYCES-CEREVISIAE, DETECTION OF THE RNA8-PROTEIN, AND DEMONSTRATION THAT IT IS ESSENTIAL FOR NUCLEAR PRE-MRNA SPLICING
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DOI：
10.1128/mcb.8.3.1067
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发表时间：
1988-03-01
影响因子：
5.3
通讯作者：
BEGGS, JD
中科院分区：
文献类型：
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作者：
JACKSON, SP;LOSSKY, M;BEGGS, JD
携带温度敏感突变rna8-1的酿酒酵母菌株在限制性温度（36℃）下核前mrna剪接缺陷。C)，表明RNA8基因编码剪接机制的一个组成部分。利用RNA8 -1突变菌株的温度敏感生长缺陷，克隆了RNA8基因。整合转化和基因破坏实验证实了克隆DNA的身份，并证明了RNA8基因编码的基本功能。RNA8基因在酿酒酵母单倍体基因组中只出现一次，编码约7.4千碱基的低丰度转录物。通过使用抗血清。-半乳糖苷酶-RNA8功能蛋白，RNA8基因产物在酿酒酵母细胞提取物中鉴定为低丰度蛋白，约260千顿。免疫缺失RNA8蛋白特异性地消除了酿酒酵母体外剪接提取物的活性，证实了RNA8在剪接中起重要作用。
Strains of Saccharomyces cerevisiae that bear the temperature-sensitive mutation rna8-1 are defective in nuclear pre-mRNA splicing at the restrictive temperature (36.degree. C), suggesting that the RNA8 gene encodes a component of the splicing machinery. The RNA8 gene was cloned by complementation of the temperature-sensitive growth defect of an rna8-1 mutation strain. Integrative transformation and gene disruption experiments confirmed the identity of the cloned DNA and demonstrated that the RNA8 gene encodes an essential function. The RNA8 gene was shown to be represented once per S. cerevisiae haploid genome and to encode a low-abundance transcript of approximately 7.4 kilobases. By using antisera raised against .beta.-galactosidase-RNA8 function proteins, the RNA8 gene product was identified in S. cerevisiae cell extracts as a low-abundance protein of approximately 260 kilodaltons. Immunodepletion of the RNA8 protein specifically abolished the activity of S. cerevisiae in vitro splicing extracts, confirming that RNA8 plays an essential role in splicing.