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MS-based identification of small proteins in Campylobacter jejuni

ABSTRACT: The Gram-negative bacterium Campylobacter jejuni is a widespread food-borne pathogen. Knowing the proteins encoded by the bacterial pathogen and how their expression is regulated is essential to understand how it survives, colonizes, and causes diseases. The present study focusses on small proteins (≤ 50-100 amino acids) translated from small open reading frames (sORFs). These poorly annotated components of the genome show emerging roles in bacterial physiology and virulence. Here, the proteome of C. jejuni during exponential growth in a complex medium was analyzed with the proteogenomics workflow presented in Fuchs et al., 2021 with the special focus on small proteins . In combination with different Ribo-Seq approaches new insights into the coding potential of the genome of C. jejuni are provided.

INSTRUMENT(S): Orbitrap Fusion

ORGANISM(S): Campylobacter Jejuni Subsp. Jejuni Nctc 11168 = Atcc 700819

SUBMITTER: Martin Kucklick

LAB HEAD: Susanne Engelmann

PROVIDER: PXD036790 | Pride | 2024-05-23

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	Campylobacter_jejuni_NCTC11168_ATCC700819_NCBI_salt_TRDB_20200117.fasta	Fasta
	Campylobacter_jejunu_NCTC11168_ATCC700819_NCBI_HIRI-novelORFS_aus_Wuerzburg-20200109.fasta	Fasta
	E7913_ude19_Cj_R1_Tryp_F1.raw	Raw
	E7914_ude19_Cj_R1_Tryp_F2.raw	Raw
	E7915_ude19_Cj_R1_Tryp_F3.raw	Raw

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Project description:Campylobacter jejuni is currently the leading cause of bacterial gastroenteritis in humans. Comparison of multiple Campylobacter strains revealed a high genetic and phenotypic diversity. However, little is known about differences in transcriptome organization, gene expression, and small RNA (sRNA) repertoires. Here we present the first comparative primary transcriptome analysis based on the differential RNAM-bM-^@M-^Sseq (dRNAM-bM-^@M-^Sseq) of four C. jejuni isolates. Our approach includes a novel, generic method for the automated annotation of transcriptional start sites (TSS), which allowed us to provide genome-wide promoter maps in the analyzed strains. These global TSS maps are refined through the integration of a SuperGenome approach that allows for a comparative TSS annotation by mapping RNAM-bM-^@M-^Sseq data of multiple strains into a common coordinate system derived from a whole-genome alignment. Considering the steadily increasing amount of RNAM-bM-^@M-^Sseq studies, our automated TSS annotation will not only facilitate transcriptome annotation for a wider range of pro- and eukaryotes but can also be adapted for the analysis among different growth or stress conditions. Our comparative dRNAM-bM-^@M-^Sseq analysis revealed conservation of most TSS, but also single-nucleotide-polymorphisms (SNP) in promoter regions, which lead to strain-specific transcriptional output. Furthermore, we identified strain-specific sRNA repertoires that could contribute to differential gene regulation among strains. In addition, we identified a novel minimal CRISPR-system in Campylobacter of the type-II CRISPR subtype, which relies on the host factor RNase III and a trans-encoded sRNA for maturation of crRNAs. This minimal system of Campylobacter, which seems active in only some strains, employs a unique maturation pathway, since the crRNAs are transcribed from individual promoters in the upstream repeats and thereby minimize the requirements for the maturation machinery. Overall, our study provides new insights into strain-specific transcriptome organization and sRNAs, and reveals genes that could modulate phenotypic variation among strains despite high conservation at the DNA level. Our dRNA-seq study of multiple C. jejuni strains represents the first comparative analysis of the primary transcriptomes of multiple strains and provides new insights into riboregulation in this bacterial pathogen.

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MS-based identification of small proteins in Campylobacter jejuni

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