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The dividing form called the reticulate body (RB) must convert at late times into the infectious elementary body (EB) for spread to new host cells. Late genes are a temporal class of chlamydial genes believed to be responsible for RB-to-EB conversion, but late gene regulation is incompletely understood. In this study, we used chromatin immunoprecipitation (ChIP) to investigate two alternative sigma factors, \u03c3\n                      <jats:sup>28<\/jats:sup>\n                      and \u03c3\n                      <jats:sup>54<\/jats:sup>\n                      , that alter the promoter specificity of\n                      <jats:italic toggle=\"yes\">Chlamydia trachomatis<\/jats:italic>\n                      RNA polymerase. \u03c3\n                      <jats:sup>28<\/jats:sup>\n                      ChIP-seq identified\n                      <jats:italic toggle=\"yes\">hctB<\/jats:italic>\n                      and\n                      <jats:italic toggle=\"yes\">tsp<\/jats:italic>\n                      as the only promoters bound by \u03c3\n                      <jats:sup>28<\/jats:sup>\n                      , and binding only occurred late, around the time of RB-to-EB conversion. Overexpression of \u03c3\n                      <jats:sup>28<\/jats:sup>\n                      confirmed that these genes are transcribed in a \u03c3\n                      <jats:sup>28<\/jats:sup>\n                      -dependent manner. \u03c3\n                      <jats:sup>54<\/jats:sup>\n                      ChIP-seq showed that \u03c3\n                      <jats:sup>54<\/jats:sup>\n                      only bound\n                      <jats:italic toggle=\"yes\">ctl0021<\/jats:italic>\n                      and\n                      <jats:italic toggle=\"yes\">ctl0052<\/jats:italic>\n                      and only at late times. This \u03c3\n                      <jats:sup>54<\/jats:sup>\n                      regulon appears to be conserved as\n                      <jats:italic toggle=\"yes\">in silico<\/jats:italic>\n                      analysis identified \u03c3\n                      <jats:sup>54<\/jats:sup>\n                      promoter sequences upstream of\n                      <jats:italic toggle=\"yes\">ctl0021<\/jats:italic>\n                      and\n                      <jats:italic toggle=\"yes\">ctl0052<\/jats:italic>\n                      homologs in all\n                      <jats:italic toggle=\"yes\">Chlamydia<\/jats:italic>\n                      spp. The genes encoding \u03c3\n                      <jats:sup>28<\/jats:sup>\n                      and \u03c3\n                      <jats:sup>54<\/jats:sup>\n                      were only transcribed at late times, but ChIP analysis with the late regulator Euo showed that Euo only controls \u03c3\n                      <jats:sup>28<\/jats:sup>\n                      expression, and late transcription of \u03c3\n                      <jats:sup>54<\/jats:sup>\n                      is regulated in an Euo-independent manner. Thus, multiple mechanisms regulate late genes, including Euo and different forms of RNA polymerase. The dedicated use of two alternative RNA polymerases to control a small subset of late genes suggests that these genes and the independent control of their temporal expression are important for RB-to-EB conversion.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>IMPORTANCE<\/jats:title>\n                    <jats:p>\n                      In this study, we performed chromatin immunoprecipitation-seq to identify genes transcribed by alternative forms of RNA polymerases in\n                      <jats:italic toggle=\"yes\">Chlamydia trachomatis<\/jats:italic>\n                      . Under normal growth conditions, the sigma factors, \u03c3\n                      <jats:sup>28<\/jats:sup>\n                      and \u03c3\n                      <jats:sup>54<\/jats:sup>\n                      , bound only two genes each, and binding was only detected at late times. In addition, the late regulator Euo controls the expression of \u03c3\n                      <jats:sup>28<\/jats:sup>\n                      but not \u03c3\n                      <jats:sup>54<\/jats:sup>\n                      . Thus,\n                      <jats:italic toggle=\"yes\">Chlamydia<\/jats:italic>\n                      utilizes multiple mechanisms to regulate late gene expression and uses alternative forms of RNA polymerases for specialized control of specific late genes that likely have important roles in reticulate body to elementary body conversion. This genome-wide binding approach can be applied to identify target genes of alternative sigma factors in other pathogenic bacteria.\n                    <\/jats:p>\n                  <\/jats:sec>","DOI":"10.1128\/msystems.00292-25","type":"journal-article","created":{"date-parts":[[2025,6,12]],"date-time":"2025-06-12T09:00:23Z","timestamp":1749718823000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":1,"title":["Late gene regulation by the alternative sigma factors of\n                    <i>Chlamydia trachomatis<\/i>"],"prefix":"10.1128","volume":"10","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-0867-0589","authenticated-orcid":false,"given":"Syed M. 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