Dual roles for Spt5 in pre-mRNA processing and transcription elongation revealed by identification of Spt5-associated proteins
Dual roles for Spt5 in pre-mRNA processing and transcription elongation revealed by identification of Spt5-associated proteins
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DOI：
10.1128/mcb.23.4.1368-1378.2003
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发表时间：
2003-02-01
影响因子：
5.3
通讯作者：
Hartzog, GA
中科院分区：
文献类型：
--
作者：
Lindstrom, DL;Squazzo, SL;Hartzog, GA
在转录延伸过程中，真核RNA聚合酶II （Pol II）必须与核小体提供的屏障相抗衡。保守的Spt4-Spt5复合物被认为通过核小体通过Pol II调节延伸。为了帮助确定Spt5的功能机制，我们对与Spt5共免疫纯化的蛋白进行了表征。其中包括一般的延伸因子TFIIF和TFIIS以及Spt6和FACT，这些因子被认为是通过核小体调节延伸的。Spt5还与mRNA盖帽酶和帽甲基转移酶共免疫纯化，并且spt4和Spt5突变与盖帽酶基因突变表现出遗传相互作用。此外，我们发现spt4和spt5突变导致未剪接的pre-mRNA积累。Spt5还与一些以前未研究过的蛋白质进行了共化；我们证明其中一个是由SPT基因家族的新成员编码的。最后，通过免疫沉淀这些因子，我们发现Spt5参与至少三个Pol II复合物的证据。这些观察结果为Spt4-Spt5在mrna前加工和转录伸长中的作用提供了新的证据。
During transcription elongation, eukaryotic RNA polymerase II (Pol II) must contend with the barrier presented by nucleosomes. The conserved Spt4-Spt5 complex has been proposed to regulate elongation through nucleosomes by Pol II. To help define the mechanism of Spt5 function, we have characterized proteins that coimmunopurify with Spt5. Among these are the general elongation factors TFIIF and TFIIS as well as Spt6 and FACT, factors thought to regulate elongation through nucleosomes. Spt5 also coimmunopurified with the mRNA capping enzyme and cap methyltransferase, and spt4 and spt5 mutations displayed genetic interactions with mutations in capping enzyme genes. Additionally, we found that spt4 and spt5 mutations lead to accumulation of unspliced pre-mRNA. Spt5 also copurified with several previously unstudied proteins; we demonstrate that one of these is encoded by a new member of the SPT gene family. Finally, by immunoprecipitating these factors we found evidence that Spt5 participates in at least three Pol II complexes. These observations provide new evidence of roles for Spt4-Spt5 in pre-mRNA processing and transcription elongation.