Expression of a beta-galactosidase gene containing the ribosomal protein 51 intron is sensitive to the rna2 mutation of yeast.
Expression of a beta-galactosidase gene containing the ribosomal protein 51 intron is sensitive to the rna2 mutation of yeast.
复制标题
含有核糖体蛋白51内含子的β-半乳糖苷酶基因的表达对酵母的rna2突变敏感。
DOI：
10.1073/pnas.80.14.4403
复制
发表时间：
1983
影响因子：
11.1
通讯作者：
Rosbash,M
中科院分区：
文献类型：
--
作者：
Teem,JL;Rosbash,M
温度敏感突变 rna2 导致酵母核糖体蛋白 51 (rp51) 基因和许多其他酵母核糖体蛋白基因的较高分子量转录物的积累。我们已经确定了 rp51 基因的 DNA 序列，确认其含有内含子，并且较高分子量的转录物是含有内含子的前体 RNA。这些数据和其他实验表明，rna2 突变影响 mRNA 加工（剪接），并且内含子的存在足以使基因的表达对 rna2 突变敏感。为了测试这些假设，我们将 rp51 内含子插入杂合大肠杆菌 β-半乳糖苷酶基因的编码区，从而中断起始甲硫氨酸密码子之后的开放阅读框。尽管存在内含子，β-半乳糖苷酶基因仍在酵母中表达。因此，rp51 内含子被从正常无内含子的基因中正确切除。 rp51内含子的存在导致β-半乳糖苷酶活性对rna2突变敏感，这与该突变在剪接水平影响基因表达的观点一致。实验表明，含有内含子的 β-半乳糖苷酶基因可用于研究 mRNA 剪接的通用方法。
The temperature-sensitive mutation rna2 causes the accumulation of higher molecular weight transcripts from the ribosomal protein 51 (rp51) gene of yeast and many other yeast ribosomal protein genes. We have determined the DNA sequence of the rp51 gene, confirming that it contains an intron and that the higher molecular weight transcript is an intron-containing precursor RNA. These data and other experiments suggest that the rna2 mutation affects mRNA processing (splicing) and that the presence of an intron is sufficient to render expression of a gene sensitive to the rna2 mutation. To test these hypotheses, we have inserted the rp51 intron into the coding region of a hybrid Escherichia coli beta-galactosidase gene, thereby interrupting the open reading frame subsequent to the initiating methionine codon. Despite the presence of the intron, the beta-galactosidase gene is expressed in yeast. Thus, the rp51 intron is properly excised from the normally intronless gene. The presence of the rp51 intron causes the beta-galactosidase activity to be sensitive to the rna2 mutation, consistent with the notion that this mutation affects gene expression at the level of splicing. The experiments suggest that an intron-containing beta-galactosidase gene can be used in a general way to study mRNA splicing.