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The specific gene(s) that participates in the mtROS-mediated antibacterial process remains unclear. We showed that Listeria and Salmonella infections in human and mouse macrophages increased mtDNA copy number with which dictates antibacterial activity. Interestingly, adenylate kinase 4 (Ak4) expression was upregulated in macrophages after infection. Ak4 KO mice as well as macrophage-specific Ak4 KO mice became highly susceptible to bacterial infections. Ak4 is critical for the increase of mtDNA synthesis and mitochondrial mass in macrophages after bacterial infection. Biochemically, Ak4 transfers a phosphate group from ATP\/GTP to (d)AMP for (d)ADP formation, and the K18A and G89S\/A166D mutations abolished this function. Our results suggest that induction of Ak4 after infection produces more dADP, whose conversion to dATP in mitochondria supports mtDNA synthesis and the subsequent increase of mtROS production. Loss of this metabolic coupling in Ak4 KO macrophages diminishes antibacterial activity. Our findings highlight the vital role of Ak4 in macrophage defense against pathogenic bacteria.<\/jats:p>","DOI":"10.1084\/jem.20250978","type":"journal-article","created":{"date-parts":[[2026,1,30]],"date-time":"2026-01-30T07:20:52Z","timestamp":1769757652000},"update-policy":"https:\/\/doi.org\/10.1084\/jem.crossmarkpolicy","source":"Crossref","is-referenced-by-count":5,"title":["Macrophage anti-bacterial activity is controlled by adenylate kinase 4\u2013mediated mitochondrial DNA synthesis"],"prefix":"10.1084","volume":"223","author":[{"ORCID":"https:\/\/orcid.org\/0009-0003-0765-8868","authenticated-orcid":true,"given":"Wei-Yao","family":"Chin","sequence":"first","affiliation":[{"id":[{"id":"https:\/\/ror.org\/05bqach95","id-type":"ROR","asserted-by":"publisher"}],"name":"Graduate Institute of Immunology, National Taiwan University College of Medicine 1 , Taipei,","place":["Taiwan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0000-0128-6811","authenticated-orcid":true,"given":"Ching-Tung","family":"Wu","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/05bqach95","id-type":"ROR","asserted-by":"publisher"}],"name":"Graduate Institute of Immunology, National Taiwan University College of Medicine 1 , Taipei,","place":["Taiwan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2619-8516","authenticated-orcid":true,"given":"Gunn-Guang","family":"Liou","sequence":"additional","affiliation":[{"name":"Cryo-EM Core Facility of the First Core Laboratory, National Taiwan University College of Medicine 2 , Taipei,","place":["Taiwan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5145-258X","authenticated-orcid":true,"given":"Si-Tse","family":"Jiang","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/05wcstg80","id-type":"ROR","asserted-by":"publisher"}],"name":"National Laboratory Animal Center, National Applied Research Laboratories 3 , Taipei,","place":["Taiwan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4423-4381","authenticated-orcid":true,"given":"Yi-Sheng","family":"Cheng","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/05bqach95","id-type":"ROR","asserted-by":"publisher"}],"name":"Genome and Systems Biology Degree Program, College of Life Science, National Taiwan University 4 , Taipei,","place":["Taiwan"]},{"id":[{"id":"https:\/\/ror.org\/05bqach95","id-type":"ROR","asserted-by":"publisher"}],"name":"College of Life Science, National Taiwan University 5 Department of Life Science, , Taipei,","place":["Taiwan"]},{"id":[{"id":"https:\/\/ror.org\/05bqach95","id-type":"ROR","asserted-by":"publisher"}],"name":"Institute of Plant Biology, College of Life Science, National Taiwan University 6 , Taipei,","place":["Taiwan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0583-148X","authenticated-orcid":true,"given":"Jr-Shiuan","family":"Lin","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/05bqach95","id-type":"ROR","asserted-by":"publisher"}],"name":"Graduate Institute of Immunology, National Taiwan University College of Medicine 1 , Taipei,","place":["Taiwan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1522-2055","authenticated-orcid":true,"given":"Betty A.","family":"Wu-Hsieh","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/05bqach95","id-type":"ROR","asserted-by":"publisher"}],"name":"Graduate Institute of Immunology, National Taiwan University College of Medicine 1 , Taipei,","place":["Taiwan"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1347-9367","authenticated-orcid":true,"given":"Shi-Chuen","family":"Miaw","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/05bqach95","id-type":"ROR","asserted-by":"publisher"}],"name":"Graduate Institute of Immunology, National Taiwan University College of Medicine 1 , Taipei,","place":["Taiwan"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"291","published-online":{"date-parts":[[2026,3,12]]},"reference":[{"key":"2026061818115919100_bib1","doi-asserted-by":"publisher","first-page":"493","DOI":"10.1038\/s41586-024-07487-w","article-title":"Accurate structure prediction of biomolecular interactions with AlphaFold 3","volume":"630","author":"Abramson","year":"2024","journal-title":"Nature"},{"key":"2026061818115919100_bib2","doi-asserted-by":"publisher","first-page":"14190","DOI":"10.1038\/s41598-018-31170-6","article-title":"Mitochondrial transcription factor A (TFAM) shapes metabolic and invasion gene signatures in melanoma","volume":"8","author":"Araujo","year":"2018","journal-title":"Sci. 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