Viral nucleases reveal an mRNA degradation-transcription feedback loop in mammalian cells

Gamma-herpesviruses encode a cytoplasmic mRNA-targeting endonuclease, termed SOX, that cleaves the majority of mRNAs within a cell. Cleaved fragments are subsequently degraded by the cellular mRNA degradation machinery. Here, we reveal that mammalian cells respond to this widespread cytoplasmic mRNA decay by altering levels of RNA polymerase II (RNAPII) transcription in the nucleus. Measurements of both RNAPII recruitment to promoters and nascent mRNA synthesis revealed that the majority of affected genes are transcriptionally repressed in SOX-expressing cells. The transcriptional feedback does not occur in response to the initial endonuclease-induced cleavage, but instead to degradation of the cleaved fragments by cellular exonucleases. In particular, Xrn1 catalytic activity is required for transcriptional repression. Notably, viral mRNA transcription escapes decay-induced repression, and this escape requires Xrn1. Collectively, these results indicate that mRNA decay rates impact transcription in mammalian cells, and that gamma-herpesviruses have incorporated this feedback mechanism into their own gene expression strategy. NIH 3T3 cells were mock, WT, or ∆HS infected with MHV68 in duplicate and 4sU-labeled RNA isolated. 4sU-labeled RNA was submitted for sequencing and reads aligned to the mouse genome or MHV68 viral genome. Differential cellular gene expression was determined between mock and WT infected, mock and ∆HS infected, as well as differential viral gene expression between WT and ∆HS.

γ疱疹病毒（Gamma-herpesviruses）编码一种被称为SOX的靶向细胞质mRNA的核酸内切酶，该酶可切割细胞内绝大多数mRNA。切割产生的片段随后会被细胞的mRNA降解机制降解。本研究发现，哺乳动物细胞可通过改变细胞核内RNA聚合酶II（RNAPII）的转录水平，来响应这种广泛存在的细胞质mRNA降解过程。通过检测RNAPII在启动子（promoters）上的招募情况以及新生mRNA合成水平，研究人员发现，在表达SOX的细胞中，绝大多数受影响的基因均发生了转录抑制。该转录反馈并非由核酸内切酶最初引发的切割事件所触发，而是由细胞核酸外切酶（exonucleases）对切割片段的降解所介导。具体而言，Xrn1的催化活性是转录抑制得以发生的必要条件。值得注意的是，病毒mRNA的转录可逃脱降解诱导的抑制，且该逃脱过程依赖于Xrn1。综上，上述结果表明，mRNA的降解速率会影响哺乳动物细胞的转录过程，而γ疱疹病毒已将这种反馈机制整合至自身的基因表达策略中。将NIH 3T3细胞以空白对照（mock）、野生型（WT）或∆HS毒株进行MHV68感染并设置生物学重复，随后分离4-硫尿苷（4sU）标记的RNA。将4sU标记的RNA进行测序，并将测序读段（reads）比对至小鼠基因组或MHV68病毒基因组。分别确定了空白对照感染与野生型感染、空白对照感染与∆HS感染之间的细胞基因差异表达情况，以及野生型感染与∆HS感染之间的病毒基因差异表达情况。