Project description:Comparative analysis of the global gene expression of a Haemophilus ducreyi 35000HP cpxA deletion mutant relative to the wild type strain
Project description:To better understand the role of the (p)ppGpp-mediated stringent response in control of H. ducreyi virulence determinants, here we defined genes potentially regulated by either (p)ppGpp or DksA by using RNA-seq. We were able to show that loss of either (p)ppGpp or DksA resulted in dysregulation of multiple genes including several known virulence determinants. We also show that loss of (p)ppGpp or DksA resulted in differential expression of putative H. ducreyi small RNAs. RNA of Haemophilus ducreyi wildtype, relAspoT and dksA mutants were collected at mid-log, transition, and stationary phases of growth, in quadruplicate using stranded RNA -seq.
Project description:To better understand the role of H. ducreyi CpxRA in controlling virulence determinants, here we defined genes potentially regulated by CpxRA by using RNA-Seq. Activation of CpxR by deletion of cpxA repressed nearly 70% of its targets, including seven established virulence determinants. Inactivation of CpxR by deletion of cpxR differentially regulated few genes and increased the expression of one virulence determinant. We identified a CpxR binding motif that was enriched in downregulated but not upregulated targets. These data reinforce the hypothesis that CpxA phosphatase activity plays a critical role in controlling H. ducreyi virulence in vivo. Characterization of the downregulated genes may offer new insights into pathogenesis. RNA of Haemophilus ducreyi wildtype, cpxA and cpxR mutants were collected at mid-log, transition, and stationary phases of growth, in quadruplicate.
Project description:Comparative analysis of the global gene expression of a Haemophilus ducreyi 35000HP cpxA deletion mutant relative to the wild type strain After the cpxA mutant was generated, both strain were grown in Columbia Broth. After 8 hr of growth RNA wa isolated and processed for DNA microarray analysis. This study includes three biological replicates (paired samples), and all three were subjected to dye swap.
Project description:To potentially understand how H. ducreyi responds to membrane stress, here we defined RpoE-dependent genes using RNA-Seq. We identified 180 RpoE-dependent genes, of which 98% were upregulated; a major set of these genes encodes homologues of envelope maintenance and repair factors. We also identified and validated a putative RpoE promoter consensus sequence, which was enriched in the majority of RpoE-dependent targets. Comparison of RpoE-dependent genes to those controlled by CpxR showed that each transcription factor regulated a distinct set of genes. Given that RpoE activated a large number of genes encoding envelope maintenance and repair factors and CpxRA represses genes encoding envelope-localized proteins, these data suggest that RpoE and CpxRA appear to play distinct yet complementary roles in regulating envelope homeostasis in H. ducreyi. RNA of Haemophilus ducreyi 35000HP wild-type strain containing a rpoE inducible plasmid and wild-type strain containing a control plasmid were collected at 0 minutes, 5 minutes, and 10 minutes after induction in quadruplicate.
Project description:The goal of this study was to compare the global trascription profile of a Haemophilus ducreyi hfq deletion mutant to that of the wild type parental strain.
