Pleiotropic Regulation of Virulence Genes in Streptococcus mutans by the Conserved Small Protein SprV

Abstract

Streptococcus mutans, an oral pathogen associated with dental caries, colonizes tooth surfaces as polymicrobial biofilms known as dental plaque. S. mutans expresses several virulence factors that allow the organism to tolerate environmental fluctuations and compete with other microorganisms. We recently identified a small hypothetical protein (90 amino acids) essential for the normal growth of the bacterium. Inactivation of the gene, SMU.2137, encoding this protein caused a significant growth defect and loss of various virulence-associated functions. An S. mutans strain lacking this gene was more sensitive to acid, temperature, osmotic, oxidative, and DNA damage-inducing stresses. In addition, we observed an altered protein profile and defects in biofilm formation, bacteriocin production, and natural competence development, possibly due to the fitness defect associated with SMU.2137 deletion. Transcriptome sequencing revealed that nearly 20% of the S. mutans genes were differentially expressed upon SMU.2137 deletion, thereby suggesting a pleiotropic effect. Therefore, we have renamed this hitherto uncharacterized gene as sprV (streptococcal pleiotropic regulator of virulence). The transcript levels of several relevant genes in the sprV mutant corroborated the phenotypes observed upon sprV deletion. Owing to its highly conserved nature, inactivation of the sprV ortholog in Streptococcus gordonii also resulted in poor growth and defective UV tolerance and competence development as in the case of S. mutans. Our experiments suggest that SprV is functionally distinct from its homologs identified by structure and sequence homology. Nonetheless, our current work is aimed at understanding the importance of SprV in the S. mutans biology.IMPORTANCE Streptococcus mutans employs several virulence factors and stress resistance mechanisms to colonize tooth surfaces and cause dental caries. Bacterial pathogenesis is generally controlled by regulators of fitness that are critical for successful disease establishment. Sometimes these regulators, which are potential targets for antimicrobials, are lost in the genomic context due to the lack of annotated homologs. This work outlines the regulatory impact of a small, highly conserved hypothetical protein, SprV, encoded by S. mutans. We show that SprV affects the transcript levels of various virulence factors required for normal growth, biofilm formation, stress tolerance, genetic competence, and bacteriocin production.KEYWORDS pleiotropic regulator, Streptococcus mutans, bacteriocins, biofilms, global virulence, stress response S treptococcus mutans expresses various stress tolerance mechanisms and virulence factors, which allow the organism to endure the detrimental oral environment, colonize tooth surfaces as dental plaque, and eventually cause caries by demineralization of the enamel (1). The coordinated expression of stress responsive and virulencerelated genes is critical for the persistence of S. mutans in the oral environment that

Involvement of ClpE ATPase in Physiology of Streptococcus mutans

“…The sensitivity of various S. mutans strains to different stressors was measured as described previously, with some modifications ( 52 – 54 ). Briefly, cells were grown from a 1:20 dilution in BHI overnight at 37°C.…”

MapZ deficiency leads to defects in the envelope structure and changes stress tolerance of Streptococcus mutans

“…The osmotic stress assay was performed as previously described (Shankar et al., 2017). Briefly, an overnight culture grown in BHI medium was diluted (1:100) in the same medium and incubated aerobically at 37°C to OD 600 = 0.1.…”

Regulatory mechanisms of exopolysaccharide synthesis and biofilm formation in Streptococcus mutans

“…The streptococcal pleiotropic regulator of virulence (SprV) is a small hypothetical protein that is encoded by smu.2137 in S. mutans . The deletion of SprV leads to an impaired biofilm formation and various virulence-related functions in S. mutans [ 106 ]. Transcriptome sequencing revealed that gtfB, gbpA, gbpB , and gbpC , which are genes related to biofilm formation, are downregulated by 2.6-, 2.4-, 1.7-, and 2.6-fold, respectively [ 106 ].…”

“… [ 105 ] sprV Smu.2137 Deletion of sprV inhibited exopolysaccharides synthesis and biofilm formation in S. mutans. [ 106 ] recA Smu.2085 Deletion of recA in S. mutans produced lower density biofilm. [ 107 ] …”