Regulation of NAD+ Homeostasis by SsNrtR in Streptococcus Sobrinus: A Critical Determinant of Its Cariogenic Potential.

Nicotinamide adenine dinucleotide (NAD+) serves as a critical cofactor in redox reactions and metabolic transformations catalyzed by NAD-dependent enzymes and is essential for bacterial survival and virulence. The biosynthesis of NAD+ in the cariogenic pathogen Streptococcus Sobrinus (S. sobrinus), a pivotal participant in oral cavities of children and adolescents with a history of caries, has yet to be explored. Bioinformatics, genetics, and biochemical techniques were used to identify NAD+ biosynthesis pathways and corresponding regulator in S. Sobrinus. S. sobrinus lacks de novo NAD+ synthesis pathway but comprises NA and Nam salvage pathway I (PncA-PncB-NadD-NadE) and PnuC-NadR salvage pathway III. NiaY and PnuC were involved in the salvage pathways. N-terminal domain of SsNrtR regulator was identified as DNA-binding domain binding to the pnuC and pncB probe, and addition of ADP-ribose reversed the binding of SsNrtR to the target promoters to regulate NAD+ salvage pathways. C-terminal domain of SsNrtR was non-catalytic, consistent with loss of Nudix motif conservation. Furthermore, the abrogation of nrtR compromised multiple pathogenic traits, including cellular proliferation, acidogenesis, and the architecture/mechanical integrity of biofilms. Consequently, this mutant exhibited attenuated virulence in a rat caries model. Our findings conclusively demonstrate that SsNrtR-mediated regulation of NAD+ homeostasis is a critical determinant of the cariogenic potential of S. sobrinus. This study identifies SsNrtR as a previously uncharacterized NAD+-responsive regulator that integrates metabolic homeostasis with the control of virulence in Streptococcus sobrinus. These findings elucidate a novel metabolic-virulence regulatory axis in this species and position SsNrtR as a promising target for the development of anti-caries interventions.