Characterization of Streptococcus mutans Strains Deficient in EIIAB ^Man of the Sugar Phosphotransferase System

Abstract

The phosphoenolpyruvate:sugar phosphotransferase system (PTS) is the major sugar uptake system in oral streptococci. The role of EIIAB Man (encoded by manL) in gene regulation and sugar transport was investigated in Streptococcus mutans UA159. The manL knockout strain, JAM1, grew more slowly than the wild-type strain in glucose but grew faster in mannose and did not display diauxic growth, indicating that EIIAB Man is involved in sugar uptake and in carbohydrate catabolite repression. PTS assays of JAM1, and of strains lacking the inducible (fruI) and constitutive (fruCD) EII fructose, revealed that S. mutans EIIAB Man transported mannose and glucose and provided evidence that there was also a mannose-inducible or glucose-repressible mannose PTS. Additionally, there appears to be a fructose PTS that is different than FruI and FruCD. To determine whether EIIAB Man controlled expression of the known virulence genes, glucosyltransferases (gtfBC) and fructosyltransferase (ftf) promoter fusions of these genes were established in the wild-type and EIIAB Man -deficient strains. In the manL mutant, the level of chloramphenicol acetyltransferase activity expressed from the gtfBC promoter was up to threefold lower than that seen with the wild-type strain at pH 6 and 7, indicating that EIIAB Man is required for optimal expression of gtfBC. No significant differences were observed between the mutant and the wild-type background in ftf regulation, with the exception that under glucose-limiting conditions at pH 7, the mutant exhibited a 2.1-fold increase in ftf expression. Two-dimensional gel analysis of batch-grown cells of the EIIAB Man -deficient strain indicated that the expression of at least 38 proteins was altered compared to that seen with the wild-type strain, revealing that EIIAB Man has a pleiotropic effect on gene expression.Oral streptococci depend on dietary carbohydrates and carbohydrates presented in oral secretions for growth and persistence in the mouth. The ability of oral streptococci to metabolize a wide variety of carbohydrates to produce organic acids is directly related to their ability to elicit dental caries. The phosphoenolpyruvate (PEP):sugar phosphotransferase system (PTS) is the primary sugar transport system in oral streptococci, especially under carbohydrate-limiting conditions, and plays important roles in global control of gene expression (20,29,30,37,38). The PTS consists of two proteins that are common to all PTS substrates, enzyme I (EI) and the heatstable phosphocarrier protein HPr, as well as a variety of sugar-specific permeases, known as EII complexes, which catalyze the transport and concomitant phosphorylation of the substrate. The EII complexes usually consist of three domains, A, B, and C, but sometimes a fourth domain, D, is required (20, 29). The A and B domains participate in phosphorylation of the cognate substrates, whereas the C and D domains comprise the membrane permeases. The EII domains can either be covalently linked as a single protein or can be present in various combi...

The complete genome sequence of Clostridium difficile phage ϕC2 and comparisons to ϕCD119 and inducible prophages of CD630

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Responses of Cariogenic Streptococci to Environmental Stresses

“…Using 2D gels, many alterations in expression of cytoplasmic and membrane proteins were observed in these mutants. More recently, it was shown that EIIAB Man plays a role in CCR by regulating diauxic growth in S. mutans and S. salivarius (Abranches et al, 2003;Pelletier et al, 1998;Vadeboncoeur and Pelletier, 1997). In S. mutans, EIIAB Man can also regulate the activity of other sugar-specific EII enzymes, as well as the expression of important virulence genes, including gtfBC and ftf (Abranches et al, 2003).…”

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