Bidirectional transcription initiation occurs at promoters where two RNA polymerase II (RNAPII) pre-initiation complexes (PICs) are organized in opposite directions. For coding genes, this results in the production of precursor mRNAs (pre-mRNAs) in the sense direction and short unstable promoter upstream transcripts (PROMPTs) in the antisense direction. However, only transcription in the sense direction continues to produce complete pre-mRNAs. Although it has been reported that U1 sites enriched in the sense direction are associated with transcription directionality, the detailed regulatory mechanism is unclear. Now, using chemical-induced protein degradation and various sequencing methods, Yang, Li et al. found that Integrator, a protein complex associated with the RNAPII C-terminal domain, regulates transcription directionality by cleaving PROMPTs through its endonuclease activity.

Acute depletion of Integrator by the mini-auxin-inducible degron system did not affect transcription loading or PIC assembly, but promoted transcription bias toward the antisense direction. This is manifested by inducing the accumulation of PROMPTs and reducing pre-mRNA transcription at actively transcribed genes. This transcriptional deviation effect could be rescued by the re-expression of INTS11, an Integrator component functioning as an endonuclease, but not its catalytically dead mutant. Cleavage of PROMPTs by antisense oligonucleotides mimicked the rescue effect of INTS11, suggesting the essential role of endonuclease activity in determining transcriptional direction. Further studies showed that U1 sites suppressed the cleavage of PROMPTs by Integrator. Introducing U1 sites into the PROMPT regions increased their transcription. Overall, this study reports a new player in regulating bidirectional transcription, and deepens the understanding of Integrator function.

双向转录起始发生在启动子处，其中两个 RNA 聚合酶 II (RNAPII) 预起始复合物 (PIC) 以相反方向组织。对于编码基因，这会导致有义方向产生前体 mRNA (pre-mRNA) 和反义方向产生短的不稳定启动子上游转录物 (PROMPT)。然而，只有正义方向的转录才能继续产生完整的前 mRNA。尽管有报道称有义方向富集的U1位点与转录方向性相关，但详细的调控机制尚不清楚。现在，Yang、Li 等人利用化学诱导的蛋白质降解和各种测序方法。发现 Integrator 是一种与 RNAPII C 端结构域相关的蛋白质复合物，通过其核酸内切酶活性切割 PROMPT 来调节转录方向。

微型生长素诱导型降解决定子系统对 Integrator 的急性消耗并不影响转录加载或 PIC 组装，但会促进转录偏向反义方向。这通过诱导 PROMPT 的积累和减少活跃转录基因的前 mRNA 转录来体现。这种转录偏差效应可以通过 INTS11 的重新表达来挽救，INTS11 是一种充当核酸内切酶功能的整合器组件，但不能通过其催化死亡突变体来挽救。反义寡核苷酸对 PROMPT 的切割模拟了 INTS11 的救援作用，表明核酸内切酶活性在决定转录方向中的重要作用。进一步的研究表明，U1 位点抑制了 Integrator 对 PROMPT 的切割。将 U1 位点引入 PROMPT 区域会增加其转录。总体而言，本研究报告了调节双向转录的新参与者，并加深了对积分器功能的理解。