Department of Microbiology, University of Virginia Health System, P.O. Box 800734, Charlottesville, VA 22908-0734, USA.
Mol Cell Biol. 2011 May;31(9):1848-60. doi: 10.1128/MCB.01346-10. Epub 2011 Feb 28.
Nucleosomes containing histone variant H2A.Z (Htz1) serve to poise quiescent genes for activation and transcriptional initiation. However, little is known about their role in transcription elongation. Here we show that dominant mutations in the elongation genes SPT5 and SPT16 suppress the hypersensitivity of htz1Δ strains to drugs that inhibit elongation, indicating that Htz1 functions at the level of transcription elongation. Direct kinetic measurements of RNA polymerase II (Pol II) movement across the 9.5-kb GAL10p-VPS13 gene revealed that the elongation rate of polymerase is 24% slower in the absence of Htz1. We provide evidence for two nonexclusive mechanisms. First, we observed that both the phospho-Ser2 levels in the elongating isoform of Pol II and the loading of Spt5 and Elongator over the GAL1 open reading frame (ORF) depend on Htz1. Second, in the absence of Htz1, the density of nucleosome occupancy is increased over the GAL10p-VPS13 ORF and the chromatin is refractory to remodeling during active transcription. These results establish a mechanistic role for Htz1 in transcription elongation and suggest that Htz1-containing nucleosomes facilitate Pol II passage by affecting the correct assembly and modification status of Pol II elongation complexes and by favoring efficient nucleosome remodeling over the gene.
组蛋白变体 H2A.Z（Htz1）形成的核小体可以使静止基因处于激活和转录起始状态。然而，它们在转录延伸中的作用知之甚少。在这里，我们发现延伸基因 SPT5 和 SPT16 的显性突变可以抑制 htz1Δ 菌株对抑制延伸的药物的敏感性，这表明 Htz1 在转录延伸水平上起作用。对 Pol II（聚合酶 II）跨越 9.5kb GAL10p-VPS13 基因的直接动力学测量表明，在没有 Htz1 的情况下，聚合酶的延伸速度慢 24%。我们提供了两种非排他性机制的证据。首先，我们观察到延伸型 Pol II 的磷酸化 Ser2 水平以及 Spt5 和 Elongator 在 GAL1 开放阅读框（ORF）上的加载都依赖于 Htz1。其次，在没有 Htz1 的情况下，GAL10p-VPS13 ORF 上核小体占据的密度增加，并且在活跃转录过程中染色质不易重塑。这些结果确立了 Htz1 在转录延伸中的机制作用，并表明 Htz1 包含的核小体通过影响 Pol II 延伸复合物的正确组装和修饰状态，并有利于基因上有效核小体重塑，从而促进 Pol II 穿过。
