CHARACTERIZATION OF LATE GENE PROMOTERS OF CHLAMYDIA-TRACHOMATIS

Chlamydiae possess an intracellular developmental cycle defined by the orderly interconversion of infectious, metabolically inactive element ary bodies and noninfectious, dividing reticulate bodies. Only a few s tage-specific genes have been cloned and sequenced, including the late -stage cysteine-rich protein operon and two late-stage genes encoding histone-like proteins. The aims of this study were to identify additio nal late-stage genes of Chlamydia trachomatis, analyze the upstream DN A sequence of late genes, and determine the sigma factor requirement o f late genes. Stage-specific RNA, made by chlamydiae isolated from hos t cells, was used to probe C. trachomatis genomic libraries. Two new l ate genes, designated ltuA and ltuB, were identified, cloned, and sequ enced. The predicted peptides encoded by ltuA and ltuB do not bear str ong homology to known proteins, and the function of the new late genes is not known. The 5' ends of the transcripts of ltuA, ltuB, the cyste ine-rich protein operon, and the two histone-like genes (hctA and hctB ) were mapped, and a consensus -10 promoter region of TATAAT was deriv ed from their upstream DNA sequences. In vitro transcription from temp lates encoding the promoter regions of ltuA, ltuB, and hctA cloned int o the transcription assay vector pUC19-spf' was found to be strongly s timulated by the addition of recombinant chlamydial sigma(66), while t ranscription from the putative hctB promoter region cloned in pUC19-sp f' was not detected in either the presence or absence of added sigma(6 6). These results suggest that the transcription of at least some chla mydial late-stage genes is dependent on sigma(66), which is homologous to the major sigma factors of other eubacteria.