Regulation of the Cyclic AMP Receptor Protein of Yersinia pestisPublic Deposited
Ubiquitous in the environment are bacteria that have evolved to adapt to the environmental niches they colonize. To this end, bacteria sensory and signaling molecules are required for processing these extracellular changes within their environment into changes in gene expression. The plague-causing pathogen, Yersinia pestis, contains a repertoire of two-component systems, transcriptional regulators, and riboregulators that allow Y. pestis to transmit between fleas, mammals, and multiple tissue sites therein. Each of these environments is unique in the available nutrients and host defenses. Its closest evolutionary ancestor, Yersinia pseudotuberculosis, is a gastrointestinal pathogen that is not associated with fatal disease. The close similarity of Y. pestis to Y. pseudotuberculosis allows for studying of the evolutionary changes that permitted Y. pestis to adapt to a new life cycle. The cyclic-AMP receptor protein, Crp, is a transcriptional regulator that is 100% identical between Y. pestis and Y. pseudotuberculosis. However, differences in regulation of the crp gene could change when Crp-regulated genes are activated or repressed. \tOur previous work discovered that the small RNA chaperone, Hfq, is required for translation of Crp in Y. pestis. Hfq binds to messenger RNAs (mRNAs) and small non-coding RNAs (sRNAs) to regulate gene expression. Hfq control of Crp protein levels is temperature- and species-dependent and suggests sRNAs are involved in regulating expression of crp. A transposon screen was employed to identify sRNAs controlling expression of crp at the post-transcriptional level. The screen identified putative sRNAs and the PmrAB two-component system as potential regulators of crp expression. Identifying the regulators of crp expression in Y. pestis may allow for understanding the species-specific differences in crp expression. Y. pestis encounters environments that change as the disease progresses. During pneumonic plague, Y. pestis proliferates within the lungs consuming available glucose by 72 hours post-infection (hpi). Expression of the pla gene, a critical virulence factor required for disease progression and target of Crp, increases as glucose is depleted from the lungs. Crp is necessary for expression of pla, and high-resolution microscopy captured increased expression of crp within large biofilm-like structures in mouse-infected lungs. Deletion of the glucose specific transporter, ptsG, attenuates growth in the lung, prevents glucose depletion, and the induction of crp and pla expression. These findings provide the first, direct association between activation of catabolite-repressed genes in response to changes in available glucose during infection. \tAfter identifying increased expression of crp within biofilms in the absence of glucose, I sought to identify differentially expressed genes within planktonic and biofilm-grown cells in the presence and absence of glucose by RNA-sequencing. RNA was also sequenced from a âˆ†crp mutant of Y. pestis to identify Crp-regulated genes in Y. pestis. Carbon source and Crp had significant impacts on expression of genes in Y. pestis. Crp regulates the expression of hundreds of mRNA-encoding genes and dozens of sRNAs. Crp directly represses the quorum sensing regulator, ypeR, but there is no significant reduction in AHL production in Y. pestis âˆ†crp. In addition, Crp is required for growth of Y. pestis in multiple carbon sources. The information learned from RNA-sequencing will be valuable in understanding how Y. pestis adapts to growth in the biofilm state and identifying Crp-regulated genes.