The role of two novel proteins YvcJ/YhbJ and YvcK/YbhK in cell wall biosynthesis in presumable connection to the PTS in Bacillus subtilis and Escherichia coli

细菌磷酸转移酶系统(PTS)通过由多个蛋白质组成的磷酸化级联反应催化糖的同时运输和磷酸化。PTS的磷酸化状态调节细胞中的一些信号转导链，这些信号转导链涉及分解代谢抑制、趋化和许多基因的调节。因此，PTS可以被视为细菌的大脑。我们在枯草芽孢杆菌中发现了两个新的基因yvcJ和yvcK，它们在细胞壁的生物合成或其调节中起着重要作用。这两个基因存在于许多进化分化的细菌中，并经常与PTS相关基因一起聚集在基因组上，表明它们之间存在功能联系。在芽孢杆菌中，它们存在于编码CRH的yvd-N-操纵子中，CRH是PTS的磷酸化载体蛋白HPR的同源物。我们发现YvcK缺陷的枯草杆菌突变株获得了类似L的细胞形态，并最终在利用需要糖异生的碳源进行代谢时裂解。我们的数据表明，该突变体不能提供合成细胞壁前体分子所需的足够水平的糖酵解中间体。在大肠杆菌中，yhbJ基因(与yvcJ同源)存在于rpoN操纵子中，rpoN操纵子也编码另一种54因子，以及蛋白质IIANtR和NPR，它们是PTS酶的同源蛋白，功能未知。我们发现，yhbJ的缺失导致氨基葡萄糖合成酶GLMS的大量过量生产，该GLMS催化形成氨基葡萄糖-6-磷酸，这是细胞壁合成中的一个关键反应。异常高的GLMS水平也可以很好地解释枯草杆菌yvcK缺失的影响，这表明这两个基因产物在GLMS数量的控制下是相互关联的。在大肠杆菌中，与yvcK同源的BiphK是单独编码的，可能是由一个依赖于54的启动子表达的。数据指向一个模型，在该模型中，YhbJ控制依赖于54的YbhK的表达，而YbhK反过来又调节大肠杆菌中GLMS的水平。在这里，我们建议在两个模式细菌枯草杆菌和大肠杆菌中研究这一信号转导途径。首先，我们想要探讨YhbJ(YvcJ)是否影响GLMS的转录、翻译或降解。其次，我们将研究YhbJ是直接作用于GLMS，还是间接作用于？54，从而调节YbhK的表达，进而调节大肠杆菌中GLMS的数量。最后，我们想要解决的是与yhbJ(YvcJ)共同表达的PTS-同源基因的作用，以及它们是否调节了这一通往GLMS的信号转导途径，从而调节细胞壁的生物合成以适应细胞的代谢状态。

The bacterial phosphotransferase System (PTS) catalyzes the simultaneous transport and phosphorylation of sugars using a phosphorylation cascade composed of several proteins. The phosphorylation state of the PTS regulates a number of signal transduction chains in the cell implicated in catabolite repression, in chemotaxis and in regulation of numerous genes. The PTS can therefore be regarded as the bacterial brain . We identified two novel genes named yvcJ and yvcK in Bacillus subtilis with a fundamental role in biosynthesis of the cell wall or its regulation. These two genes exist in many evolutionary divergent bacteria and often cluster on the genome together with genes related to the PTS, suggesting a functional connection. In Bacilli they are present within the yvd-N-operon encoding also Crh, a homologue of the phosphocarrier protein HPr of the PTS. We found that B. subtilis mutants defective for yvcK acquire an L-form-like cell shape and finally lyse when carbon sources are utilized that require gluconeogenesis for their metabolism. Our data suggest that this mutant is unable to provide a sufficient level of glycolytic intermediates required for the synthesis of cell wall precursor molecules. In Escherichia coli gene yhbJ (homologous to yvcJ) is present within the rpoN operon that also encodes the alternative ¿54-factor and the proteins IIANtr and NPr which are orthologues of PTS-enzymes and of unknown function. We found that deletion of yhbJ leads to huge overproduction of the glucosamine synthase GlmS that catalyzes the formation of glucosamine-6-phosphate, a key reaction in cell wall synthesis. An abnormal high GlmS level would also perfectly explain the effects of an yvcK deletion in B. subtilis suggesting that both gene products are interconnected in the control of the GlmS amount. In E. coli, ybhK (homologous to yvcK) is separately encoded and probably expressed from a ¿54-dependent Promoter. The data point to a model in which YhbJ controls the ¿54-dependent expression of YbhK which in turn regulates the GlmS level in E. coli. Here we propose to investigate this signal transduction pathway in the two model bacteria B. subtilis and E. coli. First, we want to explore whether glmS transcription or translation or degradation of its product is affected by YhbJ (YvcJ). Second, we will study whether YhbJ acts directly on GlmS or indirectly perhaps by interaction with ¿54 thereby modulating the expression of YbhK which in turn could regulate the GlmS amount in E. coli. Finally, we want to address the role of the PTS-orthologues co-expressed together with yhbJ (yvcJ) and whether they modulate this signal transduction pathway towards GlmS and thereby adjust cell wall biosynthesis to the metabolic state of the cell.