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While gram-negative bacteria use ApaH-family hydrolases to degrade Ap\n                      <jats:sub>4<\/jats:sub>\n                      A and other dinucleoside tetraphosphates (Ap\n                      <jats:sub>4<\/jats:sub>\n                      Ns), gram-positive bacteria, including\n                      <jats:italic toggle=\"yes\">Staphylococcus aureus,<\/jats:italic>\n                      use YqeK. Inactivation of Ap\n                      <jats:sub>4<\/jats:sub>\n                      A hydrolases and corresponding Ap\n                      <jats:sub>4<\/jats:sub>\n                      A accumulation cause diverse phenotypic effects in gram-negative and gram-positive species, ranging from increased antimicrobial sensitivity to reduced virulence. However, the physiological role of YqeK in\n                      <jats:italic toggle=\"yes\">S. aureus<\/jats:italic>\n                      remains uncharacterized. Here, we constructed an isogenic\n                      <jats:italic toggle=\"yes\">yqeK<\/jats:italic>\n                      mutant in\n                      <jats:italic toggle=\"yes\">S. aureus<\/jats:italic>\n                      and showed that \u2206\n                      <jats:italic toggle=\"yes\">yqeK<\/jats:italic>\n                      was sensitive to nitrosative and organic acid stress, with all phenotypes restored by complementation. We used a luminescence-based assay to quantify relative Ap\n                      <jats:sub>4<\/jats:sub>\n                      Ns and found that \u2206\n                      <jats:italic toggle=\"yes\">yqeK<\/jats:italic>\n                      had ~1,000 fold higher signal than wild-type even during unstressed growth. Transcriptomics revealed that \u2206\n                      <jats:italic toggle=\"yes\">yqeK<\/jats:italic>\n                      exhibited stress-specific dysregulation of translation, nucleotide metabolism, central metabolism, iron acquisition, and stress response genes. In contrast, \u2206\n                      <jats:italic toggle=\"yes\">yqeK<\/jats:italic>\n                      had few transcriptional differences relative to the wild-type during unstressed growth despite the large Ap\n                      <jats:sub>4<\/jats:sub>\n                      N accumulation, suggesting that the effects of Ap\n                      <jats:sub>4<\/jats:sub>\n                      Ns are contingent on the cellular stress state. Unexpectedly, the\n                      <jats:italic toggle=\"yes\">agr<\/jats:italic>\n                      quorum sensing operon and numerous virulence genes showed reduced expression in \u2206\n                      <jats:italic toggle=\"yes\">yqeK<\/jats:italic>\n                      , correlating with reduced hemolytic activity. This represents the first reported link between Ap\n                      <jats:sub>4<\/jats:sub>\n                      N accumulation and quorum sensing suppression in a gram-positive pathogen. Our data reveal YqeK to be a critical metabolic determinant of\n                      <jats:italic toggle=\"yes\">S. aureus<\/jats:italic>\n                      stress resistance and virulence and position this hydrolase as a promising candidate for anti-virulence drug development.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>IMPORTANCE<\/jats:title>\n                    <jats:p>\n                      <jats:italic toggle=\"yes\">Staphylococcus aureus<\/jats:italic>\n                      is a leading cause of antibiotic-resistant bacterial infections worldwide and is resistant to many components of the host immune response. Here, we discovered that deletion of YqeK, an enzyme that degrades a stress-associated nucleotide signaling molecule called Ap\n                      <jats:sub>4<\/jats:sub>\n                      A, rendered\n                      <jats:italic toggle=\"yes\">S. aureus<\/jats:italic>\n                      more susceptible to infection-relevant stress conditions but had little impact on normal growth. Ap\n                      <jats:sub>4<\/jats:sub>\n                      Ns accumulated in the\n                      <jats:italic toggle=\"yes\">yqeK<\/jats:italic>\n                      mutant and caused major stress-specific changes in gene expression, including reduced expression of key virulence genes. This correlated with a reduction in the destruction of red blood cells, a measure of bacterial toxicity toward host cells. Our data suggest that YqeK represents a promising target for new drugs aimed at reducing the virulence of\n                      <jats:italic toggle=\"yes\">S. aureus<\/jats:italic>\n                      .\n                    <\/jats:p>\n                  <\/jats:sec>","DOI":"10.1128\/jb.00265-26","type":"journal-article","created":{"date-parts":[[2026,8,20]],"date-time":"2026-08-20T13:00:37Z","timestamp":1787230837000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":0,"title":["Loss of the Ap\n                    <sub>4<\/sub>\n                    A hydrolase YqeK impairs stress adaptation and virulence gene expression in\n                    <i>Staphylococcus aureus<\/i>"],"prefix":"10.1128","volume":"208","author":[{"ORCID":"https:\/\/orcid.org\/0009-0005-1204-1795","authenticated-orcid":false,"given":"J.","family":"Vidaud","sequence":"first","affiliation":[{"name":"Department of Biology, University of North Carolina Asheville","place":["Asheville, USA"]},{"name":"Department of Chemistry and Biochemistry, University of North Carolina Asheville","place":["Asheville, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"J. F.","family":"Coker","sequence":"additional","affiliation":[{"name":"Department of Biology, University of North Carolina Asheville","place":["Asheville, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"J.","family":"Silva","sequence":"additional","affiliation":[{"name":"Department of Biology, University of North Carolina Asheville","place":["Asheville, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0000-0613-1173","authenticated-orcid":false,"given":"G. S.","family":"Davidson","sequence":"additional","affiliation":[{"name":"Department of Biology, University of North Carolina Asheville","place":["Asheville, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"C. C.","family":"Anderson","sequence":"additional","affiliation":[{"name":"Department of Biology, University of North Carolina Asheville","place":["Asheville, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"G. M.","family":"Bassett","sequence":"additional","affiliation":[{"name":"Department of Biology, University of North Carolina Asheville","place":["Asheville, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"A. A.","family":"Harry","sequence":"additional","affiliation":[{"name":"Department of Biology, University of North Carolina Asheville","place":["Asheville, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"T.","family":"Dusenbury","sequence":"additional","affiliation":[{"name":"Department of Biology, University of North Carolina Asheville","place":["Asheville, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"T. M.","family":"Gardner","sequence":"additional","affiliation":[{"name":"Department of Biology, University of North Carolina Asheville","place":["Asheville, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"M. L.","family":"Melear","sequence":"additional","affiliation":[{"name":"Department of Biology, University of North Carolina Asheville","place":["Asheville, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"N. J.","family":"Moraga","sequence":"additional","affiliation":[{"name":"Department of Biology, University of North Carolina Asheville","place":["Asheville, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"J. D.","family":"Fender","sequence":"additional","affiliation":[{"name":"Department of Biology, University of North Carolina Asheville","place":["Asheville, USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6619-3592","authenticated-orcid":true,"given":"C. 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