Genome-Wide Transcriptional Start Site Mapping and sRNA Identification in the Pathogen Leptospira interrogans

<i>Leptospira</i> are emerging zoonotic pathogens transmitted from animals to humans typically through contaminated environmental sources of water and soil. Regulatory pathways of pathogenic <i>Leptospira</i> spp. underlying the adaptive response to different hosts and environmental conditions remains elusive. In this study, we provide the first global Transcriptional Start Site (TSS) map of a <i>Leptospira</i> species. RNA was obtained from the pathogen <i>Leptospira interrogans</i> grown at 30°C (optimal <i>in vitro</i> temperature) and 37°C (host temperature) and selectively enriched for 5' ends of native transcripts. A total of 2865 and 2866 primary TSS (pTSS) were predicted in the genome of <i>L. interrogans</i> at 30 and 37°C, respectively. The majority of the pTSSs were located between 0 and 10 nucleotides from the translational start site, suggesting that leaderless transcripts are a common feature of the leptospiral translational landscape. Comparative differential RNA-sequencing (dRNA-seq) analysis revealed conservation of most pTSS at 30 and 37°C. Promoter prediction algorithms allow the identification of the binding sites of the alternative sigma factor sigma 54. However, other motifs were not identified indicating that <i>Leptospira</i> consensus promoter sequences are inherently different from the <i>Escherichia coli</i> model. RNA sequencing also identified 277 and 226 putative small regulatory RNAs (sRNAs) at 30 and 37°C, respectively, including eight validated sRNAs by Northern blots. These results provide the first global view of TSS and the repertoire of sRNAs in <i>L. interrogans</i>. These data will establish a foundation for future experimental work on gene regulation under various environmental conditions including those in the host.