Recently, our group showed that the dental pathogen Streptococcus mutans is inherently more tolerant to high zinc stress than other streptococci, a phenotype associated with the presence of a P-type ATPase exporter named ZccE, virtually unique to S. mutans. In addition to zccE, a previous transcriptome analysis revealed that S. mutans upregulates genes involved in glutathione uptake during initial exposure to zinc stress. Glutathione, a major supplier of organic sulfur that also plays key roles in antioxidant defense and xenobiotic detoxification, forms coordination complexes with a variety of metals, including zinc, thereby functioning as a buffer that protects cells from metal intoxication. To investigate the contribution of glutathione zinc tolerance in S. mutans, the gshT gene, which encodes the substrate-binding subunit of a glutathione transporter, was deleted in both the parent and {Delta}zccE strains and the ability of these mutants to overcome zinc stress through intracellular glutathione accumulation determined. Targeted metabolomics revealed that S. mutans accumulates glutathione in a GshT-dependent manner following zinc stress, a response that was strikingly amplified in the {Delta}zccE strain. Although glutathione supplementation had a minimal and non-significant impact on growth of either parent or mutant strains in sub-inhibitory zinc concentrations, the {Delta}gshT strain exhibited increased zinc sensitivity in a plate-based assay. However, the {Delta}zccE{Delta}gshT mutant displayed enhanced zinc tolerance compared to the {Delta}zccE single mutant. While glutathione alone did not alter zinc levels in the UA159 or {Delta}zccE strains, the combination of zinc and glutathione nearly doubled intracellular zinc levels in {Delta}zccE compared to cells grown in zinc only. We conclude that while glutathione may play a minor role in S. mutans zinc tolerance, uncontrolled glutathione uptake observed in {Delta}zccE facilitates zinc entry, as glutathione:Zn2+ complexes inadvertently promote zinc intoxication via a Trojan horse mechanism.
Carter, M. C., Womack, E., Khatib, M., Peterson, A. M., Saengpet, I. S., Lemos, J. A.
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