Department of Life Science, Sogang University, Seoul, South Korea.
mBio. 2022 Apr 26;13(2):e0383921. doi: 10.1128/mbio.03839-21. Epub 2022 Mar 21.
Exopolysaccharides (EPSs), biofilm-maturing components of Vibrio vulnificus, are abundantly produced when the expression of two major EPS gene clusters is activated by an enhancer-binding transcription factor, DctD, whose expression and phosphorylation are induced by dicarboxylic acids. Surprisingly, when glucose was supplied to V. vulnificus, similar levels of expression of these clusters occurred, even in the absence of dicarboxylic acids. This glucose-dependent activation was also mediated by DctD, whose expression was sequentially activated by the transcription regulator NtrC. Most DctD in cells grown without dicarboxylic acids was present in a dephosphorylated state, known as the transcriptionally inactive form. However, in the presence of glucose, a dephosphorylated component of the glucose-specific phosphotransferase system, d-IIA, interacted with dephosphorylated DctD (d-DctD). While d-DctD did not show any affinity to a DNA fragment containing the DctD-binding sequences, the complex of d-DctD and d-IIA exhibited specific and efficient DNA binding, similar to the phosphorylated DctD. The d-DctD-mediated activation of the EPS gene clusters' expression was not fully achieved in cells grown with mannose. Furthermore, the degrees of expression of the clusters under glycerol were less than those under mannose. This was caused by an antagonistic and competitive effect of GlpK, whose expression was increased by glycerol, in forming a complex with d-DctD by d-IIA. The data demonstrate a novel regulatory pathway for V. vulnificus EPS biosynthesis and biofilm maturation in the presence of glucose, which is mediated by d-DctD through its transition to the transcriptionally active state by interacting with available d-IIA. Transcription regulation by bacterial two-component systems is achieved by a response regulator upon its transition to the transcriptionally active form via kinase activity of its cognate sensor under specific conditions. A well-known response regulator, DctD, is converted to its phosphorylated form when DctB senses ambient dicarboxylic acids. Phospho-DctD induces expression of its regulon, including the gene clusters for biosynthesis of exopolysaccharides (EPSs), the essential constituents of biofilm matrix. In the absence of dicarboxylic acids, however, DctD-mediated induction of these EPS gene clusters and biofilm maturation was observed if glucose was supplied. This suggests that dephospho-DctD could play a role in activating the transcription of target genes. A component of glucose-phosphotransferase system, IIA, was present in a dephosphorylated state in the presence of glucose. Dephospho-DctD formed a complex with dephospho-IIA and was converted to a transcriptionally active state. These findings suggest the other response regulators could also have alternative pathways of activation independent of phosphorylation.
外多糖 (EPSs) 是创伤弧菌生物膜成熟的组成部分，当两个主要的 EPS 基因簇的表达被增强结合转录因子 DctD 激活时，大量产生，DctD 的表达和磷酸化由二羧酸诱导。令人惊讶的是，当葡萄糖被供应给创伤弧菌时，即使没有二羧酸，这些簇的表达水平也相似。这种葡萄糖依赖性激活也由 DctD 介导，其表达由转录调节剂 NtrC 依次激活。在没有二羧酸的情况下生长的细胞中，大多数 DctD 以去磷酸化状态存在，称为转录非活性形式。然而，在葡萄糖存在下，葡萄糖特异性磷酸转移酶系统的去磷酸化成分 d-IIA 与去磷酸化的 DctD (d-DctD) 相互作用。虽然 d-DctD 与包含 DctD 结合序列的 DNA 片段没有任何亲和力，但 d-DctD 和 d-IIA 的复合物表现出特异性和高效的 DNA 结合，类似于磷酸化的 DctD。在细胞生长过程中，d-DctD 介导的 EPS 基因簇表达的激活并没有完全实现用甘露糖。此外，在甘油条件下，这些簇的表达程度低于用甘露糖。这是由于甘油增加的 GlpK 的拮抗和竞争作用造成的，GlpK 通过 d-IIA 与 d-DctD 形成复合物。数据表明，在葡萄糖存在的情况下，创伤弧菌 EPS 生物合成和生物膜成熟存在一种新的调控途径，该途径通过其与可用的 d-IIA 相互作用，由 d-DctD 介导，使其向转录活性状态转变。细菌双组分系统的转录调控是通过其在特定条件下通过其同源传感器的激酶活性转变为转录活性形式的响应调节剂来实现的。众所周知的响应调节剂 DctD 在 DctB 感应环境中二羧酸时被转化为磷酸化形式。磷酸化的 DctD 诱导其调节子的表达，包括生物膜基质的必需成分外多糖 (EPS) 的生物合成基因簇。然而，在没有二羧酸的情况下，如果供应葡萄糖，则观察到 DctD 介导的这些 EPS 基因簇和生物膜成熟的诱导。这表明去磷酸化的 DctD 可能在激活靶基因的转录中发挥作用。葡萄糖磷酸转移酶系统的一个成分 IIA 在存在葡萄糖时处于去磷酸化状态。去磷酸化的 DctD 与去磷酸化的 IIA 形成复合物，并被转化为转录活性状态。这些发现表明其他响应调节剂也可能存在独立于磷酸化的激活的替代途径。
