Parallel evolution of genome structure and transcriptional landscape in the Epsilonproteobacteria

Abstract

BackgroundGene reshuffling, point mutations and horizontal gene transfer contribute to bacterial genome variation, but require the genome to rewire its transcriptional circuitry to ensure that inserted, mutated or reshuffled genes are transcribed at appropriate levels. The genomes of Epsilonproteobacteria display very low synteny, due to high levels of reshuffling and reorganisation of gene order, but still share a significant number of gene orthologs allowing comparison. Here we present the primary transcriptome of the pathogenic Epsilonproteobacterium Campylobacter jejuni, and have used this for comparative and predictive transcriptomics in the Epsilonproteobacteria.ResultsDifferential RNA-sequencing using 454 sequencing technology was used to determine the primary transcriptome of C. jejuni NCTC 11168, which consists of 992 transcription start sites (TSS), which included 29 putative non-coding and stable RNAs, 266 intragenic (internal) TSS, and 206 antisense TSS. Several previously unknown features were identified in the C. jejuni transcriptional landscape, like leaderless mRNAs and potential leader peptides upstream of amino acid biosynthesis genes. A cross-species comparison of the primary transcriptomes of C. jejuni and the related Epsilonproteobacterium Helicobacter pylori highlighted a lack of conservation of operon organisation, position of intragenic and antisense promoters or leaderless mRNAs. Predictive comparisons using 40 other Epsilonproteobacterial genomes suggests that this lack of conservation of transcriptional features is common to all Epsilonproteobacterial genomes, and is associated with the absence of genome synteny in this subdivision of the Proteobacteria.ConclusionsBoth the genomes and transcriptomes of Epsilonproteobacteria are highly variable, both at the genome level by combining and division of multicistronic operons, but also on the gene level by generation or deletion of promoter sequences and 5′ untranslated regions. Regulatory features may have evolved after these species split from a common ancestor, with transcriptome rewiring compensating for changes introduced by genomic reshuffling and horizontal gene transfer.

Transcriptional Landscape Variability within the Bacterial oppA-oppB Intergenic Region as Revealed by Computational Search for the Potential Starts of RNA Synthesis

“…Для регуляторных последовательностей определяющим является достижение баланса в уровне экспрессии различных продуктов, что в большей степени зависит от условий обитания организма и необходимости динамично реагировать на их изменение [29]. Даже у микроорганизмов, относящихся к филогенетически замкнутой группе эпсилон-протеобактерий, Campylobacter jejuni и Helicobacter pylori, наблюдается сильное разнообразие как во взаимном расположении генов-ортологов, так и в позиционировании экспериментально зарегистрированных стартовых точек транскрипции, ассоциированных с этими генами [30]. Поэтому не удивительно, что сходство в распределении предсказанных сигналов транскрипции прослеживается только у близкородственных видов (рис.…”

PerR controls oxidative stress defence and aerotolerance but not motility-associated phenotypes of Campylobacter jejuni

“…Approximately 100 bp of the 39 end of the perR gene region was not removed, to avoid disruption of the cj0323 gene promoter, for which the transcription start site (TSS) is located directly downstream of perR (Fig. S1) (Dugar et al, 2013;Porcelli et al, 2013). In addition, the perR mutation was complemented in trans to ensure that phenotypic changes observed were due to the perR mutation, and not due to secondary mutations or polar effects of the insertion of the antibiotic-resistance cassette.…”

“…RNA was isolated using hot phenol (Mattatall & Sanderson, 1996) to ensure that small RNAs would not be removed by the extraction procedure. The RNA was treated with DNase I to remove genomic DNA, followed by optional treatment with Terminator exonuclease (Epicentre Biotechnology) for enrichment of primary RNAs, and treatment with tobacco acid phosphatase (Cambio) to generate 59-P ends for downstream ligation of 454 adapters (Porcelli et al, 2013;Sharma et al, 2010). After ligation of an RNA oligonucleotide to the phosphorylated 59-ends of RNA, and polyadenylation of RNA, first strand cDNA was generated using an oligo-dT containing 454-B primer.…”

“…The remaining reads were aligned against the C. jejuni NCTC 11168 genome sequence using Segemehl version 0.0.9.3 (Hoffmann et al, 2009), and converted into number of reads per nucleotide position. Graphs representing the number of mapped reads per nucleotide were visualized using the Integrated Genome Browser software from Affymetrix (Nicol et al, 2009) and analysed as described elsewhere (Porcelli et al, 2013;Sharma et al, 2010). Transcript levels of individual genes were expressed as RPKM (reads per kb per million mapped reads) values, calculated after mapping of reads using CLC Genomics Workbench v5 (CLC Bio).…”

“…RNA for RNA-seq analyses was extracted from C. jejuni NCTC 11168 wild-type (Porcelli et al, 2013) and fur perR mutants (this study), grown to an OD 600 of 0.4. RNA was isolated using hot phenol (Mattatall & Sanderson, 1996) to ensure that small RNAs would not be removed by the extraction procedure.…”