Elucidation of a unique regulatory mechanism for scrA gene expression in S.mutans

我们已经调查了S. mutans的蔗糖PTS的EnzymeII编码基因的调控机制。该基因的表达不仅受到蔗糖的调控，而且还受到山梨醇的调控。为了阐明这一特征性调控的分子机制，我们开展了两个项目：1）鉴定推定的ORFs标记的ESTAIA和ESTAIB基因。2)山梨糖醇转运基因的克隆：将该基因下游的开放阅读框鉴定为分别编码果糖激酶和磷酸甘露糖异构酶的pmi和pMK基因。北方分析表明，这些基因组成的操纵子转录独立的MANIA基因。另一个ORF（ds 2 ORF 3）下游的BMPB基因不涉及BMPA或BMPB的调节，但有趣的是，涉及应力诱导的葡聚糖依赖性聚集。虽然聚集的详细机制仍有待解决，我们可以建立一种方法来克隆直接参与聚集的基因。在克隆山梨醇转运基因的过程中，另外两个基因已经被克隆。其中一个被鉴定为编码链球菌糖代谢关键酶丙酮酸甲酸裂解酶的pfl基因。另一个与戈登链球菌葡萄糖基转移酶的正调控基因相似。由于这些基因在变形链球菌的致龋性中起着重要作用，因此，今后应与ds 2 ORF 3一起鉴定这些基因，以确定变形链球菌蔗糖代谢所必需的基因。虽然我们不能阐明这一时期的独特的ESTA-A表达的机制，研究已经扩大到意想不到的方向。

We have investigated regulatory mechanisms of the scrA gene encoding EnzymeII of sucrose PTS from S.mutans. Expression of the scrA gene was regulated not only by sucrose but also by sorbitol. In order to elucidate molecular mechanisms of this characteristic regulation, we launched two projects :1) Characterization of putative ORFs franking the scrA and scrB genes. 2) Cloning of sorbitol transport genes.ORFs downstream from the scrA gene were identified as the scrK and pmi genes encoding fructokinase and phosphomannose isomerase respectively. Northern analysis revealed that these genes consisting an operon were transcribed independently of the scrA gene. Another ORF (ds2ORF3) downstream from the scrB gene did not involve in scrA or scrB regulation but interestingly involved stress-induced glucan-dependent aggregation. Although the mechanism of the aggregation in detail remains to be resolved, we could have established a method to clone genes directly involved in the aggregation.During a process to clone sorbitol transport genes, two other genes have been cloned. One was identified as the pfl gene encoding pyruvate formate-lyase that is the key enzyme of sugar metabolism of streptococci. The other was similar to the putative positive regulator gene of S.gordonii glucosyltransferase. Since these genes should be important for cariogenisity of S.mutans, they have to be characterized in future together with ds2ORF3.We could identify all genes essential for sucrose metabolism of S.mutans. Although we could not elucidate mechanisms of the unique scrA expression within this period, study has expanded to unexpected derections.