Noble J C, Pan Z Q, Prives C, Manley J L
Cell. 1987 Jul 17;50(2):227-36. doi: 10.1016/0092-8674(87)90218-2.
To explore the mechanism and control of alternative splicing, we have characterized the products formed by splicing of SV40 early pre-mRNA in vitro and in vivo. Large T and small t mRNAs are derived from this precursor by joining alternative 5' splice sites to a single shared 3' splice site. In contrast to pre-mRNAs studied previously, we have shown that splicing to large T RNA involves the utilization of multiple lariat branch sites, while small t splicing uses a single branch site. Interestingly, the predominant branch sites utilized in splicing of large T RNA in vitro were found to differ in nuclear extracts from HeLa and human 293 cells, correlated with previously observed differences in the ratio of large T to small t mRNAs produced in the two cell types. To test the significance of this correlation, we examined the products formed by splicing of an SV40 early precursor microinjected into X. laevis oocytes. Strikingly, both the pattern of branch sites used in large T splicing and the ratio of large T to small t mRNAs produced were found to be identical to those observed in 293 cells and extracts.
为了探究可变剪接的机制及其调控，我们对猴病毒40（SV40）早期前体信使核糖核酸（pre - mRNA）在体外和体内剪接所形成的产物进行了表征。大T和小t信使核糖核酸（mRNA）是通过将可变的5'剪接位点与单个共享的3'剪接位点连接，从该前体产生的。与先前研究的前体信使核糖核酸不同，我们发现剪接形成大T RNA涉及多个套索状分支位点的利用，而小t剪接则使用单个分支位点。有趣的是，在体外剪接大T RNA时所利用的主要分支位点，在来自HeLa细胞和人293细胞的核提取物中有所不同，这与之前观察到的两种细胞类型中产生的大T与小t mRNA比例的差异相关。为了测试这种相关性的重要性，我们检查了显微注射到非洲爪蟾卵母细胞中的SV40早期前体剪接所形成的产物。令人惊讶的是，发现大T剪接中使用的分支位点模式以及产生的大T与小t mRNA的比例，与在293细胞和提取物中观察到的完全相同。
