Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham, Birmingham, Alabama 35294-0024, USA.
J Biol Chem. 2011 May 27;286(21):18816-24. doi: 10.1074/jbc.M110.202101. Epub 2011 Apr 5.
Spt5p is a universally conserved transcription factor that plays multiple roles in eukaryotic transcription elongation. Spt5p forms a heterodimer with Spt4p and collaborates with other transcription factors to pause or promote RNA polymerase II transcription elongation. We have shown previously that Spt4p and Spt5p also influence synthesis of ribosomal RNA by RNA polymerase (Pol) I; however, previous studies only characterized defects in Pol I transcription induced by deletion of SPT4. Here we describe two new, partially active mutations in SPT5 and use these mutant strains to characterize the effect of Spt5p on Pol I transcription. Genetic interactions between spt5 and rpa49Δ mutations together with measurements of ribosomal RNA synthesis rates, rDNA copy number, and Pol I occupancy of the rDNA demonstrate that Spt5p plays both positive and negative roles in transcription by Pol I. Electron microscopic analysis of mutant and WT strains confirms these observations and supports the model that Spt4/5 may contribute to pausing of RNA polymerase I early during transcription elongation but promotes transcription elongation downstream of the pause(s). These findings bolster the model that Spt5p and related homologues serve diverse critical roles in the control of transcription.
Spt5p 是一种普遍保守的转录因子，在真核转录延伸中发挥多种作用。Spt5p 与 Spt4p 形成异二聚体，并与其他转录因子合作，使 RNA 聚合酶 II 转录延伸暂停或促进。我们之前已经表明，Spt4p 和 Spt5p 也会影响 RNA 聚合酶 (Pol) I 合成核糖体 RNA；然而，之前的研究仅描述了 SPT4 缺失引起的 Pol I 转录缺陷。在这里，我们描述了 SPT5 中的两个新的、部分活跃突变，并使用这些突变株来描述 Spt5p 对 Pol I 转录的影响。spt5 和 rpa49Δ 突变之间的遗传相互作用以及核糖体 RNA 合成速率、rDNA 拷贝数和 rDNA 上 Pol I 占有率的测量表明，Spt5p 在 Pol I 转录中既发挥正调控作用，也发挥负调控作用。突变株和 WT 株的电子显微镜分析证实了这些观察结果，并支持了这样一种模型，即 Spt4/5 可能在转录延伸早期的 RNA 聚合酶 I 暂停中发挥作用，但在暂停（多个）之后促进转录延伸。这些发现支持了 Spt5p 和相关同源物在转录控制中发挥多种关键作用的模型。
