A unique nine-gene comY operon in Streptococcus mutans

Abstract

Many Gram-positive and Gram-negative bacteria possess natural competence mechanisms for DNA capture and internalization. In Bacillus subtilis, natural competence is absolutely dependent upon the presence of a seven-gene operon known as the comG operon (comGA-G). In species of Streptococcus, this function has been described for a four-gene operon (comYA-D in Streptococcus gordonii and cglA-D in Streptococcus pneumoniae). In this study, a nine-orf operon (named comYA-I) required for natural competence in Streptococcus mutans was identified and characterized. Orf analysis of this operon indicates that the first four Orfs (ComYA-D) share strong homology with ComYA-D of S. gordonii and CglA-D of S. pneumoniae, the fifth to seventh Orfs (ComYE-G) match conserved hypothetical proteins from various species of Streptococcus with ComYF possessing a predicted ComGF domain, the eighth Orf (ComYH) shows a strong homology to numerous DNA methyltransferases from restriction/modification systems, and the ninth Orf (ComYI) is homologous to acetate kinase (AckA). RT-PCR analysis of the orf junctions confirmed that all nine orfs were present in a single transcript, while real-time RT-PCR analysis demonstrated that these orfs were expressed at a level very similar to that of the first orf in the operon. Mutations were constructed in all nine putative orfs. The first seven genes (comYA-G) were found to be essential for natural competence, while comYH and comYI had reduced and normal natural competence ability, respectively. Analyses of S. mutans comY-luciferase reporter fusions indicated that comY expression is growth-phase dependent, with maximal expression at an OD 600 of about 0?2, while mutations in ciaH, comC and luxS reduced the level of comY expression. In addition, comY operon expression appears to be correlated with natural competence ability.

Heterologous Expression of sahH Reveals That Biofilm Formation Is Autoinducer-2-independent in Streptococcus sanguinis but Is Associated with an Intact Activated Methionine Cycle

“…This has been demonstrated for S. mutans and is consistent with the findings of the present study (47). An influence on expression of late competence genes was also found in LuxS mutants of S. pneumonia (22,62); however, in those studies the direct influence of AI-2 was not investigated.…”

“…Subsequently, the cultures were allowed to grow for an additional 2 h and were then plated on BHI agar supplemented with tetracycline (5 g ml Ϫ1 ) as well as on nonselective BHI plates. Transformation efficiency was defined as the ratio of transformants/total viable cells (47).…”

Genetic characterization of the hdrRM operon: a novel high-cell-density-responsive regulator in Streptococcus mutans

“…While there are numerous examples of mutations that lower the natural competence ability of S. mutans (Abranches et al, 2006;Ahn et al, 2005;Hussain et al, 2006;Merritt et al, 2005c;Qi et al, 2004;Rolerson et al, 2006;Senadheera et al, 2005Senadheera et al, , 2007Wang & Kuramitsu, 2006;Wen et al, 2005), to our knowledge, there is only one other report of a mutation that increases natural competence (Tao et al, 1993). This mutation was generated by chemical mutagenesis and its identity has, so far, not been published.…”

“…Determination of natural transformation efficiency was performed using a method similar to that described previously (Li et al, 2001;Merritt et al, 2005c). UA140 and the various hdrR and hdrM mutants were grown anaerobically to OD 600 0.2-0.3 in TH broth supplemented with 0.4 %, w/v, BSA.…”

Global transcriptional analysis of acid-inducible genes in Streptococcus mutans: multiple two-component systems involved in acid adaptation

“…To confirm these results, we examined two late competence genes, comYA and comX (Merritt et al, 2005), an early competence gene, comE, and its controlled bacteriocin-producing and immunity genes, nlmC (SMU.1914) and immA (SMU.1913), by real-time qRT-PCR. The results showed that, similar to the microarray data, comYA and comX were indeed downregulated at pH 5.5, but comE and the two bacteriocin-related genes were upregulated at this pH (Table 2).…”