Project description:Transcriptome analysis of NTHi 86-028NPrpsL, NTHi 86-028NPrpsL∆fur, and NTHi 86-028NPrpsL∆fur(pT-fur) strains Nontypeable Haemophilus influenzae (NTHi) is a commensal microorganism of the normal human nasopharyngeal flora, yet also an opportunistic pathogen of the upper and lower respiratory tracts. Changes in gene expression patterns in response to host microenvironments are likely critical for survival. One such system of gene regulation is the ability to carefully regulate iron uptake. A central regulatory system that controls iron uptake, mediated by the ferric uptake regulator Fur, is present in multiple bacteria, including NTHi. To understand the regulation of iron homeostasis in NTHi, fur was deleted in the NTHi strain 86-028NPrpsL. Using RNA-Seq, we identified both protein-encoding and small RNA genes whose expression was repressed or activated by Fur. Overall design: These data comprise transcriptional anaylses of an rpsL mutant of 86-028NP, an isogenic fur mutant of 86-028NPrpsL and a complemented fur mutant strain. All strains were grown in defined medium containing 10 µg/ml human hemoglobin to mid-log phase. Cells were then harvested and RNA extracted. A total of three biological replicates were generated for these analyses. Analysis of transcriptomes using the Illumina HiSeq 2000 of three strains of nontypeable Haemophilus influenzae which include NTHi 86-028NPrpsL, NTHi 86-028NPrpsL∆fur, and NTHi 86-028NPrpsL∆fur(pT-fur) strains. For each strain three biological replicates were analyzed
Project description:Regulatory elements that dictate how Haemophilus influenzae adapts to the airways of COPD patients are poorly understood. In this work, global fitness profiling of H. influenzae mutants by transposon mutagenesis/sequencing (Tn-seq) within the mouse lung identified Dam methyltransferase as an in vivo requirement, even though dam mutants grow normally during in vitro culture. In turn, methylation sequencing found that genome-wide Dam methylation patterns were undermethylated at GATC motifs with putative regulatory elements. This was confirmed for the htpG heat-shock chaperone gene, making it the first case of phenotypic variation controlled by variable Dam methylation for H. influenzae. Moreover, RNA sequencing revealed a novel multifactorial regulatory network, in which Dam methyltransferase activity is coordinated with the FNR and Fur transcriptional regulators to control the expression anaerobic defences against host-produced reactive species. Our results indicate key bacterial pathways for survival in diseased environments with high nitrosative stress, where damage to the airways reduces the amount of oxygen in the lungs as is encountered in COPD.
Project description:Background: Haemophilus influenzae has an absolute aerobic growth requirement for heme and has developed multiple complex mechanisms to obtain this essential nutrient. Although an understanding of the heme acquisition mechanisms of H. influenzae is emerging, significant gaps remain to be elucidated. In a previous study we utilized H. influenzae strain Rd KW20 to demonstrate the utility of transcriptional profiling in defining the genes exhibiting altered transcription in response to environmental iron and heme levels. The current study expands upon those initial observations by determining the iron/heme regulons of two additional H. influenzae clinical isolates, i.e. the type b isolate 10810 and the nontypeable isolate R2866, to characterize the core iron/heme regulon of the species. Results: A microarray chip was designed to incorporate probes for all of the genes of H. influenzae isolates 10810 and R2866, and microarray studies were performed to compare gene expression under iron/heme-replete and iron/heme-restricted conditions for each isolate. Of 1820 ORFs on the array corresponding to R2866 genes, 363 were significantly differentially expressed. Of these 363 genes, 233 were maximally transcribed under iron/heme-replete conditions and 130 were preferentially transcribed in iron/heme-restricted conditions. Of the 1883 ORFs representing genes of strain10810, 351 were significantly differentially transcribed, 150 of these were preferentially transcribed in iron/heme-replete conditions and 201 were preferentially transcribed in iron/heme-restricted conditions. Comparison of the data sets indicated that 163 genes were similarly regulated in both isolates and that 74 of these also exhibited similar patterns of regulation in strain Rd KW20. Conclusion: This study provides evidence for a core of H. influenzae genes that are regulated by the availability of iron and/or heme in the growth environment. Elucidation of this core regulon provides targets for investigation of genes with an unrecognized role in iron and heme homeostasis, as well as other potential virulence determinants. In addition, the set of core genes potentially provides targets for therapeutic and vaccine designs since these products of these genes are likely to be preferentially expressed during growth in iron/heme restricted sites of the human body. This SuperSeries is composed of the following subset Series: GSE11349: Transcriptional profiling of FeHm effects on Haemophilus influenzae R2866 GSE11354: Transcriptional profiling of FeHm effects on Haemophilus influenzae 10810 Keywords: SuperSeries Refer to individual Series