Canonical bacterial transcription activators bind to non-transcribed promoter elements to increase transcription of their target genes. Here we report crystal structures of binary complexes comprising domains of Caulobacter crescentus GcrA, a noncanonical bacterial transcription factor, that support a novel mechanism for transcription activation through the preferential binding of methylated cis-regulatory elements and the promotion of open complex formation through an interaction with region 2 of the principal σ factor, σ⁷⁰. We present crystal structures of the C-terminal, σ factor-interacting domain (GcrA-SID) in complex with domain 2 of σ⁷⁰ (σ⁷⁰₂), and the N-terminal, DNA-binding domain (GcrA-DBD) in complex with methylated double-stranded DNA (dsDNA). The structures reveal interactions essential for transcription activation and DNA recognition by GcrA. These structures, along with mutational analyses, support a mechanism of transcription activation in which GcrA associates with RNA polymerase (RNAP) prior to promoter binding through GcrA-SID, arming RNAP with a flexible GcrA-DBD. The RNAP–GcrA complex then binds and activates target promoters harboring a methylated GcrA binding site either upstream or downstream of the transcription start site.

中文翻译：

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新月茎杆菌 GcrA 转录激活独特机制的结构洞察

典型的细菌转录激活剂与非转录启动子元件结合以增加其靶基因的转录。在这里，我们报告了包含Caulobacter crescentus GcrA结构域的二元复合物的晶体结构，GcrA 是一种非典型的细菌转录因子，它支持一种新的转录激活机制，通过甲基化顺式调控元件的优先结合和通过与主要 σ 因子 σ ⁷⁰的区域 2 。我们展示了 C 端的晶体结构，σ 因子相互作用结构域 (GcrA-SID) 与结构域 2 的 σ ⁷⁰ (σ ⁷⁰ ₂)，以及与甲基化双链 DNA (dsDNA) 复合的 N 端 DNA 结合域 (GcrA-DBD)。这些结构揭示了 GcrA 转录激活和 DNA 识别所必需的相互作用。这些结构以及突变分析支持转录激活机制，其中 GcrA 在启动子通过 GcrA-SID 结合之前与 RNA 聚合酶 (RNAP) 结合，为 RNAP 提供灵活的 GcrA-DBD。然后 RNAP-GcrA 复合物结合并激活在转录起始位点上游或下游含有甲基化 GcrA 结合位点的目标启动子。