Molecular analysis of the urease regulatory circuit in Streptococcus salivarius

• Main Authors: Szu Chuan Huang, 黃司權
• Other Authors: Y. Y. M. Chen
• Format: Others
• Language: en_US
• Published: 2017
• Online Access: http://ndltd.ncl.edu.tw/handle/5v666v

博士 === 長庚大學 === 生物醫學研究所 === 105 === Ureolysis by Streptococcus salivarius is critical for pH homeostasis of dental plaque and prevention of dental caries. The expression of S. salivarius urease is enhanced by acidic pH and excess amounts of carbohydrate. Earlier studies have demonstrated that the differential expression is regulated mainly at the transcriptional level on the promoter 5’ to ureI, pureI. Meanwhile, sequence analysis revealed a CodY box, a VicR box and two GlnR boxes 5’ to the ‒35 element of pureI. The results of electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation assay indicated that CodY, VicR and GlnR interact with the predicted binding boxes on pureI. By using various pureI-chloramphenicol acetyltransferase gene (cat) fusion constructs and mutant strains, it was found that transcription from pureI was repressed by both CodY and VicR and activated by GlnR. Studies using chemostat-grown cultures under glucose limiting (20 mM) and excess (100 mM), at pH 7 and pH 5.5 revealed that the regulation by CodY and VicR was more pronounced at pH 7, whereas GlnR was more active at pH 5.5. Furthermore, the results of EMSA and promoter-cat fusion analysis indicated that the C-terminal domain of the RNA polymerase α subunit could interact with the AT tracks within the CodY and VicR box to enhance pureI expression. The overall regulation by CodY, VicR and GlnR in response to pH ensures optimal expression of urease in S. salivarius when the enzyme is most needed.