During transcription initiation, RNA polymerase (RNAP) binds to promoter DNA, unwinds promoter DNA to form an RNAP-promoter open complex (RPo) containing a single-stranded ‘transcription bubble,’ and selects a transcription start site (TSS). TSS selection occurs at different positions within the promoter region, depending on promoter sequence and initiating-substrate concentration. Variability in TSS selection has been proposed to involve DNA ‘scrunching’ and ‘anti-scrunching,’ the hallmarks of which are: (i) forward and reverse movement of the RNAP leading edge, but not trailing edge, relative to DNA, and (ii) expansion and contraction of the transcription bubble. Here, using in vitro and in vivo protein-DNA photocrosslinking and single-molecule nanomanipulation, we show bacterial TSS selection exhibits both hallmarks of scrunching and anti-scrunching, and we define energetics of scrunching and anti-scrunching. The results establish the mechanism of TSS selection by bacterial RNAP and suggest a general mechanism for TSS selection by bacterial, archaeal, and eukaryotic RNAP.

中文翻译：

---

转录起始位点选择的变异机制

在转录起始过程中，RNA 聚合酶 (RNAP) 与启动子 DNA 结合，解开启动子 DNA 以形成包含单链“转录泡”的 RNAP 启动子开放复合物 (RPo)，并选择转录起始位点 (TSS)。TSS 选择发生在启动子区域内的不同位置，具体取决于启动子序列和起始底物浓度。已经提出 TSS 选择的可变性涉及 DNA 'scrunching' 和 'anti-scrunching'，其特征是：（i）RNAP 前缘的向前和向后运动，但不是后缘，相对于 DNA，和（ ii) 转录气泡的膨胀和收缩。在这里，使用体外和体内蛋白质-DNA 光交联和单分子纳米操作，我们展示了细菌 TSS 选择表现出紧缩和反紧缩的两个特征，并且我们定义了紧缩和反紧缩的能量学。结果建立了细菌 RNAP 选择 TSS 的机制，并提出了细菌、古细菌和真核生物 RNAP 选择 TSS 的一般机制。