Pervasive promoter-proximal regulation in archaea: ChIP-seq stationary phase. Pervasive promoter-proximal regulation in archaea: ChIP-seq stationary phase

Recruitment of RNA polymerase and initiation factors to the promoter is the only known mechanisms for transcription activation and repression in archaea. Whether any of the subsequent steps towards productive transcription elongation is involved in regulation is not known. We characterised how the basal transcription machinery is distributed along genes in the archaeon Sulfolobus solfataricus. We discovered a distinct early elongation phase where RNA polymerases sequentially recruit the elongation factors Spt4/5 and Elf1 to form the transcription elongation complex (TEC) before the TEC escapes into productive transcription. TEC escape is rate-limiting for transcription output during exponential growth. Oxidative stress causes changes in TEC escape that correlate with changes in the transcriptome. Our results thus establish that TEC escape contributes to the basal promoter strength and facilitates transcription regulation. Impaired TEC escape coincides with the accumulation of initiation factors at the promoter and recruitment of termination factor aCPSF1 to the early TEC. This suggests two possible mechanisms for how TEC escape limits transcription, physically blocking upstream RNA polymerases during transcription initiation and premature termination of early TECs. Overall design: ChIP-seq with polyclonal antibodies

在古菌中，将RNA聚合酶与起始因子招募至启动子区域，是目前已知的唯一转录激活与抑制机制。目前尚不清楚朝着有效转录延伸的后续步骤是否参与转录调控过程。我们针对硫化叶菌（Sulfolobus solfataricus）中基础转录机器沿基因的分布特征展开了系统表征。研究发现一类独特的早期延伸阶段：在此阶段中，RNA聚合酶会依次招募延伸因子Spt4/5与Elf1，组装形成转录延伸复合物（Transcription Elongation Complex, TEC），随后该复合物方能逃逸进入有效转录状态。在指数生长期，TEC逃逸是转录产出的限速步骤。氧化应激可引发TEC逃逸的改变，且该改变与转录组的变化存在显著相关性。综上，本研究结果证实，TEC逃逸可增强基础启动子的转录强度，并助力转录调控过程。受损的TEC逃逸会伴随起始因子在启动子区域的积累，以及终止因子aCPSF1向早期TEC的招募。这提示TEC逃逸限制转录存在两种潜在机制：一是在转录起始阶段物理阻断上游的RNA聚合酶，二是引发早期TEC的提前终止。实验整体设计：使用多克隆抗体进行染色质免疫沉淀测序（ChIP-seq）