Analyses of competent and non-competent subpopulations of Bacillus subtilis reveal yhfW, yhxC and ncRNAs as novel players in competence
Analyses of competent and non-competent subpopulations of Bacillus subtilis reveal yhfW, yhxC and ncRNAs as novel players in competence
复制标题
DOI：
10.1111/1462-2920.15005
复制
发表时间：
2020-04-15
影响因子：
5.1
通讯作者：
Kuipers, Oscar P.
中科院分区：
文献类型：
--
作者：
Boonstra, Mirjam;Schaffer, Marc;Kuipers, Oscar P.
在诱导能力的营养限制条件下，只有部分枯草芽孢杆菌种群具有能力。在这里，我们通过荧光辅助细胞分选（FACS）分离了两个亚群。使用RNA-seq，我们证实了前面描述的ComK调控子。我们也首次在胜任亚群中发现了显著下调的基因。下调基因不受ComK的直接控制，但在适态亚群中具有较高水平的相应反义rna。在竞争期间，细胞分裂和复制停止。通过对其蛋白质组的研究，我们发现细胞分裂、MinD和Noc的调节因子水平较高。外切酶SbcC和SbcD也主要在转录后水平受到调控。在有能力的亚群中，yhfW被新发现为高度上调。它的缺失降低了comG的表达，对comK的表达影响不大，但有统计学意义。虽然yhfW在正常亚群中表达较高，但在其启动子区域没有comk结合位点。yhfW突变体在转化过程中存在小而显著的缺陷。代谢组学分析显示，在Delta yhfW突变体中，三羧酸（TCA）循环代谢物和几种氨基酸显著减少。RNA-seq分析显示，Delta yhfW的NAD合成基因nadA、nadB和nadC的表达较高。
Upon competence-inducing nutrient-limited conditions, only part of the Bacillus subtilis population becomes competent. Here, we separated the two subpopulations by fluorescence-assisted cell sorting (FACS). Using RNA-seq, we confirmed the previously described ComK regulon. We also found for the first time significantly downregulated genes in the competent subpopulation. The downregulated genes are not under direct control by ComK but have higher levels of corresponding antisense RNAs in the competent subpopulation. During competence, cell division and replication are halted. By investigating the proteome during competence, we found higher levels of the regulators of cell division, MinD and Noc. The exonucleases SbcC and SbcD were also primarily regulated at the post-transcriptional level. In the competent subpopulation, yhfW was newly identified as being highly upregulated. Its absence reduces the expression of comG, and has a modest, but statistically significant effect on the expression of comK. Although expression of yhfW is higher in the competent subpopulation, no ComK-binding site is present in its promoter region. Mutants of yhfW have a small but significant defect in transformation. Metabolomic analyses revealed significant reductions in tricarboxylic acid (TCA) cycle metabolites and several amino acids in a Delta yhfW mutant. RNA-seq analysis of Delta yhfW revealed higher expression of the NAD synthesis genes nadA, nadB and nadC.