Manganese Uptake, Mediated by SloABC and MntH, Is Essential for the Fitness of Streptococcus mutans

Abstract

Early epidemiological studies implicated manganese (Mn) as a possible caries-promoting agent, while laboratory studies have indicated that manganese stimulates the expression of virulence-related factors in the dental pathogen Streptococcus mutans. To better understand the importance of manganese homeostasis to S. mutans pathophysiology, we first used RNA sequencing to obtain the global transcriptional profile of S. mutans UA159 grown under Mn-restricted conditions. Among the most highly expressed genes were those of the entire sloABC operon, encoding a dual iron/manganese transporter, and an uncharacterized gene, here mntH, that codes for a protein bearing strong similarity to Nramp-type transporters. While inactivation of sloC, which encodes the lipoprotein receptor of the SloABC system, or of mntH alone had no major consequence for the overall fitness of S. mutans, simultaneous inactivation of sloC and mntH (ΔsloC ΔmntH) impaired growth and survival under Mn-restricted conditions, including in human saliva or in the presence of calprotectin. Further, disruption of Mn transport resulted in diminished stress tolerance and reduced biofilm formation in the presence of sucrose. These phenotypes were markedly improved when cells were provided with excess Mn. Metal quantifications revealed that the single mutant strains contained intracellular levels of Mn similar to those seen with the parent strain, whereas Mn was nearly undetectable in the ΔsloC ΔmntH strain. Collectively, these results reveal that SloABC and MntH work independently and cooperatively to promote cell growth under Mn-restricted conditions and that maintenance of Mn homeostasis is essential for the expression of major virulence attributes in S. mutans. IMPORTANCE As transition biometals such as manganese (Mn) are essential for all forms of life, the ability to scavenge biometals in the metal-restricted host environment is an important trait of successful cariogenic pathobionts. Here, we showed that the caries pathogen Streptococcus mutans utilizes two Mn transport systems, namely, SloABC and MntH, to acquire Mn from the environment and that the ability to maintain the cellular levels of Mn is important for the manifestation of characteristics that associate S. mutans with dental caries. Our results indicate that the development of strategies to deprive S. mutans of Mn hold promise in the combat against this important bacterial pathogen.

Manganese Depletion Leads to Multisystem Changes in the Transcriptome of the Opportunistic Pathogen Streptococcus sanguinis

“…Mn levels have been found to be related to each of these systems in other gram-positive bacteria (Kehres and Maguire, 2003;Colomer-Winter et al, 2017). Mn depletion does not appear to induce a traditional stress response, although expression of an oxidative stress tolerance protein, Dpr, significantly increased, which is consistent with a recent study on Mn depletion in S. mutans (Kajfasz et al, 2020). Here we highlight Mn-related systems we identified in this study of S. sanguinis for future investigation.…”

“…The connection between Mn and sugar catabolism is not unprecedented, as previous studies have implicated Mn as important for the activity of carbon catabolism enzymes in other bacteria (Kehres and Maguire, 2003). Additionally, recent studies in S. pneumoniae and S. mutans have shown that fluctuations in metal homeostasis influence the regulation of carbohydrate metabolism (Burcham et al, 2020;Kajfasz et al, 2020). One possible explanation for the Mn-dependent, glucose-independent CcpA repression observed in our study is the accumulation of the glycolytic intermediate, fructose-1,6-bisphosphate (FBP) during Mn depletion (Figure 7).…”

Increased oxidative stress tolerance of a spontaneously-occurringperRgene mutation inStreptococcus mutansUA159

“…A number of the differentially expressed genes belong to two large polycistronic transcriptional units (smu193c-smu217c) and (smu1750c-1764c) coding, respectively, for hypothetical proteins of unknown function and the CRISPR2-cas system, a defense mechanism believed to provide immunity against phage invasion (32). In S. mutans UA159, the two CRISPR systems are also linked to stress responses, though the reason for this association remains to be determined (33). For visualization purposes, a dot plot graph showing all genes differently expressed in UA∆perR is provided (Fig.…”

“…For RNA-Seq analysis, samples were prepared as previously described (33). Sample quality and quantity were assessed on an Agilent 2100 Bioanalyzer at the University of Florida Interdisciplinary Center for Biotechnology research (UF-ICBR).…”

Regulatory involvement of the PerR and SloR metalloregulators in theStreptococcus mutansoxidative stress response

“…Specifically, S. mutans SloR regulates an ABC-type metal ion transport operon comprised of the sloABC genes, by mechanisms that have been previously described (23,9,24,22). A more recent report describes an ancillary metal ion transporter, called MntH, that helps maintain intracellular metal ion homeostasis in S. mutans, and which bears similarity to Nramptype transporters (25). Taken together, the SloABC and MntH transporters as well as their regulators are essential for ensuring S. mutans fitness and survival in both healthy and dysbiotic plaque environments (9,25,26).…”