Polymerase pausing induced by sequence-specific RNA binding protein drives heterochromatin assembly (NET-Seq). Polymerase pausing induced by sequence-specific RNA binding protein drives heterochromatin assembly (NET-Seq)

In S. pombe, transcripts derived from the pericentromeric dg and dh repeats promote heterochromatin formation via RNAi as well as an RNAi-independent mechanism involving the RNAPII-associated RNA-binding protein Seb1 and RNA processing activities. We show that Seb1 promotes long-lived RNAPII pauses at pericentromeric repeat regions and their presence correlates with the heterochromatin-triggering activities of the corresponding dg and dh DNA fragments. Globally increasing RNAPII stalling by other means induces the formation of novel large ectopic heterochromatin domains. Such ectopic heterochromatin occurs even in cells lacking RNAi. These results uncover Seb1-mediated polymerase stalling as a signal necessary for heterochromatin nucleation. Overall design: NET-Seq was conducted on 3 samples in replicate in S. pombe.

在粟酒裂殖酵母（S. pombe）中，源自着丝粒旁侧dg和dh重复序列（pericentromeric dg and dh repeats）的转录本，可通过RNA干扰（RNAi）以及涉及RNA聚合酶II（RNAPII）结合的RNA结合蛋白Seb1和RNA加工活性的非RNA依赖途径，促进异染色质形成。本研究表明，Seb1可介导RNA聚合酶II在着丝粒旁侧重复区域产生长时停顿，且此类停顿的存在与对应dg和dh DNA片段的异染色质诱导活性密切相关。通过其他手段全局增强RNA聚合酶II的停滞效应，可诱导形成新型大型异位异染色质结构域。此类异位异染色质甚至可在缺失RNAi的细胞中形成。上述研究结果揭示，Seb1介导的聚合酶停滞是异染色质成核所必需的关键信号。整体实验设计：对粟酒裂殖酵母设置3个生物学重复样本进行新生转录物测序（Native Elongating Transcript Sequencing，NET-Seq）。