NusG prevents transcriptional invasion of H-NS-silenced genes
NusG prevents transcriptional invasion of H-NS-silenced genes
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DOI：
10.1371/journal.pgen.1008425
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发表时间：
2019-10-01
期刊：
影响因子：
4.5
通讯作者：
Figueroa-Bossi, Nara
中科院分区：
文献类型：
--
作者：
Bossi, Lionello;Ratel, Mathilde;Figueroa-Bossi, Nara
进化上保守的NusG蛋白增强细菌RNA聚合酶的持续合成能力，但也可以通过结合Rho因子并刺激Rho因子的活性来促进转录终止。Rho在锚定到富含胞苷的基序（新生RNA中所谓的Rho利用位点（Rut））后终止转录。NusG和Rho都与类核结构蛋白H-NS沉默水平获得的富含A/T的DNA有关。然而，这两种蛋白在H-NS介导的基因沉默中的相对作用仍然不完全确定。在本研究中，使用携带受阿拉伯糖诱导型阻遏物控制的nusG基因的沙门氏菌菌株来评估对NusG耗竭的全基因组响应。来自两种互补方法的结果，i）筛选通过随机转座产生的lacZ蛋白融合体和ii）转录组学分析，收敛于显示NusG的缺失导致沙门氏菌致病岛（SPI）和其他H-NS沉默基因座的大量上调。在rho基因突变的菌株Rho K130 Q中观察到了类似的SPI上调谱，尽管不完全相同。令人惊讶的是，影响Rho的主要RNA结合结构域的Rho突变Y80 C要么没有影响，要么使H-NS介导的SPI沉默更加紧密。因此，虽然证实了结合的H-NS可以在体内触发Rho依赖性转录终止的概念，但这些数据表明H-NS引发的终止完全通过NusG依赖性途径发生，并且较少依赖于Rho对Rut位点的结合。我们提供的证据表明，通过Rho招聘，并可能通过其他尚未确定的机制，NusG防止普遍的成绩单延长到H-NS沉默的地区。不执行该功能会导致整个沙门氏菌毒力程序的前馈激活。这些发现进一步深入了解了NusG/Rho在H-NS介导的基因沉默中的贡献，并强调了这种贡献对于生长中细菌的全局调节反应的适当功能的重要性。通过用户友好的基因组浏览器界面可以免费查看完整的转录组数据。
Evolutionarily conserved NusG protein enhances bacterial RNA polymerase processivity but can also promote transcription termination by binding to, and stimulating the activity of, Rho factor. Rho terminates transcription upon anchoring to cytidine-rich motifs, the so-called Rho utilization sites (Rut) in nascent RNA. Both NusG and Rho have been implicated in the silencing of horizontally-acquired A/T-rich DNA by nucleoid structuring protein H-NS. However, the relative roles of the two proteins in H-NS-mediated gene silencing remain incompletely defined. In the present study, a Salmonella strain carrying the nusG gene under the control of an arabinose-inducible repressor was used to assess the genome-wide response to NusG depletion. Results from two complementary approaches, i) screening lacZ protein fusions generated by random transposition and ii) transcriptomic analysis, converged to show that loss of NusG causes massive upregulation of Salmonella pathogenicity islands (SPIs) and other H-NS-silenced loci. A similar, although not identical, SPI-upregulated profile was observed in a strain with a mutation in the rho gene, Rho K130Q. Surprisingly, Rho mutation Y80C, which affects Rho's primary RNA binding domain, had either no effect or made H-NS-mediated silencing of SPIs even tighter. Thus, while corroborating the notion that bound H-NS can trigger Rho-dependent transcription termination in vivo, these data suggest that H-NS-elicited termination occurs entirely through a NusG-dependent pathway and is less dependent on Rut site binding by Rho. We provide evidence that through Rho recruitment, and possibly through other still unidentified mechanisms, NusG prevents pervasive transcripts from elongating into H-NS-silenced regions. Failure to perform this function causes the feedforward activation of the entire Salmonella virulence program. These findings provide further insight into NusG/Rho contribution in H-NS-mediated gene silencing and underscore the importance of this contribution for the proper functioning of a global regulatory response in growing bacteria. The complete set of transcriptomic data is freely available for viewing through a user-friendly genome browser interface.