Interactions of the Metalloregulatory Protein SloR from Streptococcus mutans with Its Metal Ion Effectors and DNA Binding Site

Spatafora, Grace A.; Corbett, John; Cornacchione, Louis P.; Daly, William H.; Galán, Diego; Wysota, Michael; Tivnan, Patrick; Collins, Justin; Nye, Dillon B.; Levitz, Talya S.; Breyer, Wendy A.; Glasfeld, Arthur

doi:10.1128/jb.00612-15

Cited by 20 publications

(42 citation statements)

References 37 publications

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“…Transcriptional repression of the sloABC operon exerted by SloR has been thoroughly characterized by one of our laboratories [30, 31, 36]. In one of these studies a conserved SloR-binding palindrome was identified upstream of the mntH gene but the specificity of SloR binding to the mntH promoter region was not explored at that time [31].…”

Section: Resultsmentioning

confidence: 99%

Manganese uptake, mediated by SloABC and MntH, is essential for the fitness ofStreptococcus mutans

Kajfasz

Katrak

Ganguly

et al. 2019

Preprint

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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 the entire sloABC operon, encoding a dual iron/manganese transporter, and an uncharacterized gene, herein 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 mntH alone had no major consequence on the overall fitness of S. mutans, simultaneous inactivation of sloC and mntH (ΔsloCΔmntH) 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 contain similar intracellular levels of Mn as the parent strain, whereas Mn was nearly undetectable in the ΔsloCΔmntH strain. Collectively, these results reveal that SloABC and MntH work independently and cooperatively to promote cell growth under Mn-restricted conditions, and that mauitanence of Mn homeostasis is essential for the expression of major virulence attributes in S. mutans.IMPORTANCEAs trace biometals such as manganese (Mn) are important for all forms of life, the ability to regulate biometals availability during infection is an essential trait of successful bacterial pathogens. 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.

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Section: Resultsmentioning

confidence: 99%

Manganese uptake, mediated by SloABC and MntH, is essential for the fitness ofStreptococcus mutans

Kajfasz

Katrak

Ganguly

et al. 2019

Preprint

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“…Herein we present evidence that supports an updated model for SloR binding at this locus, consistent with the direct binding of SloR homodimers to binding sites that span a 72bp region of DNA which includes the −10 and −35 sloABC promoter. In previous work (22), we described a pattern for SloR binding at the sloABC promoter site that involved two SloR dimers binding to two sets of inverted repeats, each 6bp in length and separated by 8bp. Subsequent analysis of the 72bp region upstream of the sloABC operon suggested that, in fact, SloR binds to three distinct but abutting 22bp sites in that region, referred to herein as A, B, and C. Each binding site is comprised of two inverted hexameric repeats with an AATTAA consensus separated by 6bp, thereby defining a putative 6-6-6 motif for SloR binding (Figure 1).…”

Section: Discussionmentioning

confidence: 99%

“…This is not surprising given frequent reports of sequence variation in and around the −35 promoter region within and across prokaryotic species (39). Since promoter strength is, in part, determined by conservation of the −10 and −35 promoter sequences (36) one might expect RNAP to have only moderate binding affinity for the relatively divergent sloR promoter (TATAAT and TATCCA) as compared to RNAP binding at the more highly conserved sloABC promoter (TATATT and TTGACT) (22), and accordingly, weaker transcription from the former as compared to the latter. The results of DNA binding and expression profiling experiments reported herein support these predictions and suggest a role for divergent sloABC and sloR promoter sequences in fine-tuning metal ion transport and minimizing the toxic effects of metal ion hyper-accumulation.…”

Section: Discussionmentioning

confidence: 99%

“…We believe SloR mediates such metalloregulation by binding directly to Mn 2+ which, in turn, promotes SloR dimerization and a subsequent conformational change at the N-terminus of the protein to facilitate DNA binding. Specifically, in a previous report we describe SloR-DNA binding upstream of the sloABC locus to a promoter-proximal SloR Recognition Element (SRE) that represses sloABC transcription, presumably via a mechanism that involves promoter exclusion to RNA polymerase (RNAP) (22, 23). Hence, the sloABC -encoded metal ion uptake machinery in S. mutans is subject to transcriptional repression by SloR under conditions of Mn 2+ availability, and conversely to de-repression when Mn 2+ becomes limiting.…”

Section: Introductionmentioning

confidence: 99%

“…Despite the central importance of SloR in promoting S. mutans survival and virulence gene expression, surprisingly little is known of the regulatory mechanism(s) that modulate SloR itself. To date, the regulation of SloR in S. mutans has been shown to be manganese-dependent and driven, in part, by the sloABC promoter via transcriptional read-through of a weak terminator that is located 3’ to the sloC gene (7, 20, 22). Hence, SloR levels that derive from sloABC promoter activity likely vary with Mn 2+ availability between and during meal times.…”

Section: Introductionmentioning

confidence: 99%

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Autoregulation of theS. mutansSloR metalloregulator is constitutive and driven by an independent promoter

Monette

Cowan

et al. 2018

Preprint

Self Cite

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Streptococcus mutans, one of ∼600 bacterial species in the human oral cavity, is among the most acidogenic constituents of the plaque biofilm. Considered to be the primary causative agent of dental caries, S. mutans harbors a 25kDa SloR metalloregulatory protein which controls metal ion transport across the bacterial cell membrane to maintain essential metal ion homeostasis. The expression of SloR derives, in part, from transcriptional readthrough of the sloABC operon which encodes a Mn2+/Fe2+ ABC transport system. Herein, we describe the details of the sloABC promoter that drives this transcription, as well as a novel independent promoter in an intergenic region (IGR) that contributes to downstream sloR expression. RT-PCR studies support sloR transcription that is independent of sloABC expression, and the results of 5′ RACE revealed a sloR transcription start site in the IGR from which the −10 and −35 promoter regions were predicted. The results of gel mobility shift assays support direct SloR binding to the IGR, albeit with lower affinity than SloR binding to the sloABCR promoter. Function of the sloR promoter was validated in qRT-PCR experiments. Interestingly, sloR expression was not significantly impacted when grown in the presence of high manganese, whereas expression of the sloABC operon was repressed under these conditions. The results of in vitro transcription studies support SloR-mediated transcriptional-activation of sloR and -repression of sloABC. Taken together, these findings implicate SloR as a bifunctional regulator that represses sloABC promoter activity and encourages sloR transcription from an independent promoter.ImportanceTooth decay is a ubiquitous infectious disease that is especially pervasive in underserved communities worldwide. S. mutans-induced carious lesions cause functional, physical, and/or aesthetic impairment in the vast majority of adults, and in 60-90% of schoolchildren in industrialized countries. Billions of dollars are spent annually on caries treatment, and productivity losses due to absenteeism from the workplace are significant. Research aimed at alleviating S. mutans-induced tooth decay is important because it can address the socioeconomic disparity that is associated with dental cavities and improve overall general health which is inextricably linked to oral health. Research focused on the S. mutans SloR metalloregulatory protein can guide the development of novel therapeutics and so alleviate the burden of dental cavities.

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Structural and dynamical changes of the Streptococcus gordonii metalloregulatory ScaR protein induced by Mn2+ ion binding

Radman,

Jelić Matošević,

Žilić

et al. 2023

International Journal of Biological Macromolecules

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Interactions of the Metalloregulatory Protein SloR from Streptococcus mutans with Its Metal Ion Effectors and DNA Binding Site

Cited by 20 publications

References 37 publications

Manganese uptake, mediated by SloABC and MntH, is essential for the fitness ofStreptococcus mutans

Manganese uptake, mediated by SloABC and MntH, is essential for the fitness ofStreptococcus mutans

Autoregulation of theS. mutansSloR metalloregulator is constitutive and driven by an independent promoter

Structural and dynamical changes of the Streptococcus gordonii metalloregulatory ScaR protein induced by Mn2+ ion binding

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