Identification and functional analysis of CT069 as a novel transcriptional regulator in Chlamydia

Only a small number of transcription factors have been predicted in Chlamydia spp., which are obligate intracellular bacteria that include a number of important human pathogens. We used a bioinformatics strategy to identify novel transcriptional regulators from the Chlamydia trachomatis genome by predicting proteins with the general structure and characteristic functional domains of a bacterial transcription factor. With this approach, we identified CT069 as a candidate transcription factor with sequence similarity at its C terminus to Treponema pallidum TroR. Like TroR, the gene for CT069 belongs to an operon that encodes components of a putative ABC transporter for importing divalent metal cations. However, CT069 has been annotated as YtgC because of sequence similarity at its N terminus to TroC, a transmembrane component of this metal ion transporter. Instead, CT069 appears to be a fusion protein composed of YtgC and a TroR ortholog that we have called YtgR. Although it has not been previously reported, a similar YtgC-YtgR fusion protein is predicted to be encoded by other Chlamydia spp. and several other bacteria, including Bacillus subtilis. We show that recombinant YtgR polypeptide bound specifically to an operator sequence upstream of the ytg operon and that binding was enhanced by Zn(2+). We also demonstrate that YtgR repressed transcription from the ytg promoter in a heterologous in vivo reporter assay. These results provide evidence that CT069 is a negative regulator of the ytg operon, which encodes a putative metal ion transporter in C. trachomatis.