Li Yunghua, Burne Robert A
Center for Oral Biology and Department of Microbiology and Immunology, University of Rochester School of Medicine and Dentistry, 601 Elmwood Avenue, Rochester, NY 14642, USA1.
Microbiology (Reading). 2001 Oct;147(Pt 10):2841-2848. doi: 10.1099/00221287-147-10-2841.
Streptococcus mutans produces a number of extracellular sucrose-metabolizing enzymes that contribute to the ability of the organism to cause dental caries, including three glucosyltransferases, the products of the gtfB, gtfC and gtfD genes, and a fructosyltransferase, encoded by the ftf gene. To better understand the regulation of the expression of these genes under environmental conditions that more closely mimic those in dental plaque, two strains of S. mutans harbouring fusions of the gtfBC (SMS102) and ftf (SMS101) promoters to a chloramphenicol acetyltransferase (CAT) gene were examined in biofilms formed in vitro. The strains were grown in a Rototorque biofilm reactor in a tryptone-yeast extract-sucrose medium. CAT specific activity in biofilm cells was measured at quasi-steady state or following additions of 25 mM sucrose or glucose, with or without pH control. After approximately 10 generations of biofilm growth, the ftf and gtfBC genes of S. mutans were found to be expressed at levels different from those reported for planktonic cells growing under otherwise similar conditions. The expression of these genes was induced by the addition of sucrose to the quasi-steady-state cultures. Expression of the gtfBC genes was influenced by environmental pH, since CAT specific activities in quasi-steady-state biofilms of strain SMS102 grown without pH control were twice those produced by cells grown with pH control. Moreover, addition of glucose to quasi-steady-state biofilms resulted in increased expression of the gtfBC-cat fusion, although the magnitude of the induction was less than that seen with sucrose. The effect of pH on ftf expression was negligible. A modest and transient induction of ftf was observed in biofilms pulsed with excess glucose and the kinetics and level of induction of ftf by excess carbohydrate were dependent on the pH of the biofilms. This study demonstrates that the type and amount of carbohydrate and the environmental pH have a major influence on transcription of the gtfBC and ftf genes when the organisms are growing in biofilms, and provides evidence for previously undisclosed regulatory circuits for exopolysaccharide gene expression in S. mutans.
变形链球菌产生多种胞外蔗糖代谢酶，这些酶有助于该生物体引发龋齿，其中包括三种葡糖基转移酶，即gtfB、gtfC和gtfD基因的产物，以及一种由ftf基因编码的果糖基转移酶。为了在更接近牙菌斑环境条件下更好地理解这些基因表达的调控机制，对两株携带gtfBC（SMS102）和ftf（SMS101）启动子与氯霉素乙酰转移酶（CAT）基因融合体的变形链球菌菌株在体外形成的生物膜中进行了研究。这些菌株在Rototorque生物膜反应器中于胰蛋白胨 - 酵母提取物 - 蔗糖培养基中生长。在准稳态或添加25 mM蔗糖或葡萄糖后，测量生物膜细胞中的CAT比活性，添加时进行或不进行pH控制。在生物膜生长约10代后，发现变形链球菌的ftf和gtfBC基因的表达水平与在其他类似条件下生长的浮游细胞所报道的水平不同。向准稳态培养物中添加蔗糖可诱导这些基因的表达。gtfBC基因的表达受环境pH影响，因为在无pH控制条件下生长的SMS102菌株的准稳态生物膜中的CAT比活性是有pH控制条件下生长的细胞所产生的两倍。此外，向准稳态生物膜中添加葡萄糖会导致gtfBC - cat融合体的表达增加，尽管诱导幅度小于蔗糖诱导的情况。pH对ftf表达的影响可忽略不计。在用过量葡萄糖脉冲处理的生物膜中观察到ftf有适度且短暂的诱导，并且过量碳水化合物对ftf的诱导动力学和水平取决于生物膜的pH。这项研究表明，当生物体在生物膜中生长时，碳水化合物的类型和量以及环境pH对gtfBC和ftf基因的转录有重大影响，并为变形链球菌中以前未公开的胞外多糖基因表达调控回路提供了证据。
