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In this study, the fast-growing, non-pathogenic model mycobacteria\n                      <jats:italic toggle=\"yes\">Mycobacterium smegmatis<\/jats:italic>\n                      mc\n                      <jats:sup>2<\/jats:sup>\n                      -155 was engineered to express the\n                      <jats:italic toggle=\"yes\">M. tb<\/jats:italic>\n                      T7SS ESX-1 system. We found that\n                      <jats:italic toggle=\"yes\">M. smegmatis<\/jats:italic>\n                      transformed with\n                      <jats:italic toggle=\"yes\">M. tb esx-1<\/jats:italic>\n                      locus genes only, as well as\n                      <jats:italic toggle=\"yes\">M. smegmatis<\/jats:italic>\n                      transformed with\n                      <jats:italic toggle=\"yes\">M. tb esx-1<\/jats:italic>\n                      and\n                      <jats:italic toggle=\"yes\">espACD<\/jats:italic>\n                      operon genes (designated MSX-1), produces and secretes the\n                      <jats:italic toggle=\"yes\">M. tb<\/jats:italic>\n                      ESX-1 protein effectors EsxA, EsxB, and EspB. However, the abundance of these proteins was higher inside the cell and culture filtrate of the MSX-1 strain. Although ESX-1 is critical for\n                      <jats:italic toggle=\"yes\">M. tb<\/jats:italic>\n                      pathogenesis, expression of\n                      <jats:italic toggle=\"yes\">M. tb<\/jats:italic>\n                      ESX-1 did not make the recombinant\n                      <jats:italic toggle=\"yes\">M. smegmatis<\/jats:italic>\n                      strains virulent in macrophages. Serendipitously, transformation of\n                      <jats:italic toggle=\"yes\">M. smegmatis<\/jats:italic>\n                      with a modified\n                      <jats:italic toggle=\"yes\">esx-1<\/jats:italic>\n                      locus in this study revealed\n                      <jats:italic toggle=\"yes\">rv3860<\/jats:italic>\n                      , a gene of previously unknown function, to be required for the transcription of\n                      <jats:italic toggle=\"yes\">pe35<\/jats:italic>\n                      ,\n                      <jats:italic toggle=\"yes\">ppe68<\/jats:italic>\n                      ,\n                      <jats:italic toggle=\"yes\">esxB,<\/jats:italic>\n                      and\n                      <jats:italic toggle=\"yes\">esxA<\/jats:italic>\n                      genes. Finally, mice vaccinated with MSX-1 were found to be as protected as mice vaccinated with\n                      <jats:italic toggle=\"yes\">Mycobacterium bovis<\/jats:italic>\n                      BCG against\n                      <jats:italic toggle=\"yes\">M. tb<\/jats:italic>\n                      infection, without becoming sensitized to tuberculin. These results show that a functional\n                      <jats:italic toggle=\"yes\">M. tb<\/jats:italic>\n                      ESX-1 system can be assembled in\n                      <jats:italic toggle=\"yes\">M. smegmatis<\/jats:italic>\n                      to uncover novel facets of the secretion machinery and that the modified\n                      <jats:italic toggle=\"yes\">M. smegmatis<\/jats:italic>\n                      strain can function as a tuberculosis (TB) vaccine. Unlike BCG, however, its deployment may be compatible with tests currently used to diagnose TB.\n                    <\/jats:p>\n                    <jats:sec>\n                      <jats:title>IMPORTANCE<\/jats:title>\n                      <jats:p>\n                        In this study, we modified\n                        <jats:italic toggle=\"yes\">Mycobacterium smegmatis<\/jats:italic>\n                        , which is often used as a surrogate model organism in mycobacterial research, to produce and assemble a functional\n                        <jats:italic toggle=\"yes\">Mycobacterium tuberculosis<\/jats:italic>\n                        (\n                        <jats:italic toggle=\"yes\">M. tb<\/jats:italic>\n                        ) ESX-1 protein secretion system. One such\n                        <jats:italic toggle=\"yes\">M. smegmatis<\/jats:italic>\n                        strain named MSX-1 was found to make a functional\n                        <jats:italic toggle=\"yes\">M. tb<\/jats:italic>\n                        ESX-1 system without becoming virulent. And in using\n                        <jats:italic toggle=\"yes\">M. smegmatis<\/jats:italic>\n                        as a chassis to study the ESX-1 system, we found that\n                        <jats:italic toggle=\"yes\">rv3860<\/jats:italic>\n                        , an\n                        <jats:italic toggle=\"yes\">M. tb<\/jats:italic>\n                        gene of previously unknown function, is needed for the production of key ESX-1 proteins. Finally, mice vaccinated with MSX-1 were as protected from tuberculosis (TB) as mice given BCG, the only approved TB vaccine. Notably, we found that unlike BCG, MSX-1 does not sensitize mice to the antigens used in existing TB diagnostic tests. These observations, taken together, highlight the utility of\n                        <jats:italic toggle=\"yes\">M. smegmatis<\/jats:italic>\n                        as a chassis to study the\n                        <jats:italic toggle=\"yes\">M. tb<\/jats:italic>\n                        ESX-1 secretion machinery.\n                      <\/jats:p>\n                    <\/jats:sec>\n                  <\/jats:sec>","DOI":"10.1128\/spectrum.01131-25","type":"journal-article","created":{"date-parts":[[2025,9,12]],"date-time":"2025-09-12T13:00:26Z","timestamp":1757682026000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":4,"title":["Assembly of the\n                    <i>Mycobacterium tuberculosis<\/i>\n                    type VII ESX-1 secretion system in\n                    <i>Mycobacterium smegmatis<\/i>\n                    identifies a new transcriptional activator of\n                    <i>esx-1<\/i>\n       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