The rnc Gene Promotes Exopolysaccharide Synthesis and Represses the vicRKX Gene Expressions via MicroRNA-Size Small RNAs in Streptococcus mutans

Abstract

Dental caries is a biofilm-dependent disease that largely relies on the ability of Streptococcus mutans to synthesize exopolysaccharides. Although the rnc gene is suggested to be involved in virulence mechanisms in many other bacteria, the information regarding it in S. mutans is very limited. Here, using deletion or overexpression mutant assay, we demonstrated that rnc in S. mutans significantly positively regulated exopolysaccharide synthesis and further altered biofilm formation. Meanwhile, the cariogenecity of S. mutans was decreased by deletion of rnc in a specific pathogen-free (SPF) rat model. Interestingly, analyzing the expression at mRNA level, we found the downstream vic locus was repressed by rnc in S. mutans. Using deep sequencing and bioinformatics analysis, for the first time, three putative microRNA-size small RNAs (msRNAs) targeting vicRKX were predicted in S. mutans. The expression levels of these msRNAs were negatively correlated with vicRKX but positively correlated with rnc, indicating rnc probably repressed vicRKX expression through msRNAs at the post-transcriptional level. In all, the results present that rnc has a potential role in the regulation of exopolysaccharide synthesis and can affect vicRKX expressions via post-transcriptional repression in S. mutans. This study provides an alternative avenue for further research aimed at preventing caries.

The Regulator Gene <b><i>rnc</i></b> Is Closely Involved in Biofilm Formation in <b><i>Streptococcus mutans</i></b>

“…3c). This is consistent with the results from an anthrone assay in our previous study [Mao et al, 2016]. Besides, we also estimated the growth characteristics and bacterial viabilities of the rnc mutant and parent strains to exclude the possibility that the differences in biofilm formation were due to variations in growth capacity.…”

“…The cariogenic bacterial species S. mutans can metabolize and convert dietary sucrose into a glucan matrix, which in turn promotes adhesion of S. mutans to the tooth's surface, leading to the formation of highly adhesive and cohesive biofilms on the tooth's surface [de Sousa et al, 2015;Garcia et al, 2017]. A previous animal study showed that Smurnc could significantly decrease the amount of sulcal caries and that the downregulation of cariogenicity might be due to its capacity to disrupt biofilm formation [Mao et al, 2016].…”

“…Also, rnc is required for actinomycin production in Streptomyces antibioticus [Lee et al, 2013]. A previous study indicated that deletion of rnc could decrease the cariogenicity of S. mutans and repress its downstream vicRKX expression at the posttranscriptional level [Mao et al, 2016]. However, the related biofunction by which rnc triggers virulence expression in S. mutans remains relatively unexplored in the literature.…”

“…Among these regulator genes, the vicRKX operon-encoded VicRK signal transduction system is crucial to biofilm formation by VicR, specifically binding to the promoter regions of gtfB/ C/D that are responsible for synthesis of the glucan matrix [Senadheera et al, 2005]. In our previous study, we found that rnc could repress vicRKX expression by posttranscriptional regulation in the mid-exponential phase [Mao et al, 2016]. This was confirmed, as the results showed that the expression of vicR/K/X and gtfB/C/D in interaction with rnc presented the same increased decline in Smurnc before the maturation phase (online suppl.…”

“…Considering rnc as a typical regulator gene, it directly responds to stimulation and regulates its target genes by posttranscriptional repression [Viegas et al, 2011;Mao et al, 2016]. Thus, we hypothesize that rnc-mediated biofilm formation includes both formation-related and disintegration-related genes, and that this kind of regulation is a dynamic process dependent on the biofilm's growth phase.…”