Comparative transcriptomics in Yersinia pestis: a global view of environmental modulation of gene expression

Background: Environmental modulation of gene expression in Yersinia pestis is critical for its life style and pathogenesis. Using cDNA microarray technology, we have analyzed the global gene expression of this deadly pathogen when grown under different stress conditions in vitro. Results: To provide us with a comprehensive view of environmental modulation of global gene expression in Y. pestis, we have analyzed the gene expression profiles of 25 different stress conditions. Almost all known virulence genes of Y. pestis were differentially regulated under multiple environmental perturbations. Clustering enabled us to functionally classify co-expressed genes, including some uncharacterized genes. Collections of operons were predicted from the microarray data, and some of these were confirmed by reverse-transcription polymerase chain reaction (RT-PCR). Several regulatory DNA motifs, probably recognized by the regulatory protein Fur, PurR, or Fnr, were predicted from the clustered genes, and a Fur binding site in the corresponding promoter regions was verified by electrophoretic mobility shift assay (EMSA). Conclusion: The comparative transcriptomics analysis we present here not only benefits our understanding of the molecular determinants of pathogenesis and cellular regulatory circuits in Y. pestis, it also serves as a basis for integrating increasing volumes of microarray data using existing methods. Keywords: stress responses The genome-wide transcriptional changes upon the environmental perturbations were monitored by the standard dual-fluorescent hybridization method with a Y. pestis whole-genome DNA microarray spotted with PCR-amplified ORFs. For the stimulon analysis, the gene expression pattern of WT strain grown under a stimulating condition was compared with that of an unperturbed control. For the regulon analysis, we compared the expression profiles of deletion mutant strains of the target regulator to Yersinia pestis strain 201 (wild-type). For microarray hybridization, two independent bacterial cultures for each growth condition were prepared as biological replicates for RNA isolation. Three separate labeled probes were made for each RNA preparation as technical replicates. Pairwise comparisons were made using dye swaps to avoid labeling bias. For microarray hybridization, two or three independent bacterial cultures for each growth condition were prepared as biological replicates for RNA isolation. Three or four separate labeled probes were made for each RNA preparation as technical replicates. Pairwise comparisons were made using dye swaps to avoid labeling bias.