Promoter-proximal pausing of RNA polymerase II (RNAPII) is a critical step for the precise regulation of gene expression. Despite the apparent close relationship between promoter-proximal pausing and nucleosome, the role of chromatin remodeler governing this step has mainly remained elusive. Here, we report highly confined RNAPII enrichments downstream of the transcriptional start site in Saccharomyces cerevisiae using PRO-seq experiments. This non-uniform distribution of RNAPII exhibits both similar and different characteristics with promoter-proximal pausing in Schizosaccharomyces pombe and metazoans. Interestingly, we find that Ino80p knockdown causes a significant upstream transition of promoter-proximal RNAPII for a subset of genes, relocating RNAPII from the main pausing site to the alternative pausing site. The proper positioning of RNAPII is largely dependent on nucleosome context. We reveal that the alternative pausing site is closely associated with the + 1 nucleosome, and nucleosome architecture around the main pausing site of these genes is highly phased. In addition, Ino80p knockdown results in an increase in fuzziness and a decrease in stability of the + 1 nucleosome. Furthermore, the loss of INO80 also leads to the shift of promoter-proximal RNAPII toward the alternative pausing site in mouse embryonic stem cells. Based on our collective results, we hypothesize that the highly conserved chromatin remodeler Ino80p is essential in establishing intact RNAPII pausing during early transcription elongation in various organisms, from budding yeast to mouse.

中文翻译：

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染色质重塑因子 Ino80 介导 RNAPII 暂停位点测定

RNA 聚合酶 II (RNAPII) 的启动子近端暂停是基因表达精确调控的关键步骤。尽管启动子近端暂停与核小体之间存在明显的密切关系，但控制这一步骤的染色质重塑剂的作用主要仍然难以捉摸。在这里，我们使用 PRO-seq 实验报告了酿酒酵母转录起始位点下游的高度限制性 RNAPII 富集。RNAPII 的这种不均匀分布在粟酒裂殖酵母和后生动物中表现出相似和不同的启动子近端暂停特征。有趣的是，我们发现 Ino80p 敲低会导致一部分基因的启动子-近端 RNAPII 发生显着的上游转变，将 RNAPII 从主要暂停位点重新定位到替代暂停位点。RNAPII 的正确定位很大程度上取决于核小体背景。我们揭示了替代暂停位点与+1核小体密切相关，并且这些基因的主要暂停位点周围的核小体结构是高度定相的。此外，Ino80p 敲低会导致 +1 核小体的模糊性增加和稳定性降低。此外，INO80 的缺失还会导致启动子近端 RNAPII 向小鼠胚胎干细胞中的替代暂停位点转移。根据我们的集体结果，我们假设高度保守的染色质重塑剂 Ino80p 对于从芽殖酵母到小鼠等各种生物体的早期转录延伸过程中建立完整的 RNAPII 暂停至关重要。