RIBOBASE

Welcome to Campylobacter jejuni

Introduction

The food-borne pathogen Campylobacter jejuni is a microaerophilic Epsilonproteobacterium and currently the leading cause of bacterial gastroenteritis worldwide. Because its annotated genome lacks homologs of key virulence factors used by other enteric pathogens, little is known about how it causes disease. In its small genome of ~1.6 Mbp, 54 small proteins of less than 70 aa are annotated. So far almost nothing is known about their involvement in C. jejuni physiology and verification of translation is lacking for most of them. Furthermore, the C. jejuni small proteome is likely larger than what is currently annotated.

The lab of Prof. Dr. Cynthia Sharma (University of Würzburg, Germany), in collaboration with the group of Prof. Dr. Rolf Backofen (University of Freiburg, Germany) in the framework of the DFG funded SPP2002 priority program, has applied an integrative translatomics approach to map the coding capacity of this major human pathogen.

CampyBrowse includes data from our three translatomics experiments (Exp1, Exp2, Exp3; Froschauer, Svensson et al. submitted) and dRNA-seq (Dugar et al. 2013).

For quick browsing, we provide four pre-set JBrowse sessions for 1) dRNA-seq & Ribo-seq, 2) Ribo-seq & TIS profiling, 3) Ribo-seq & TIS/TTS profiling, and 4) all translatomics experiments as described in the JBrowse2 section.

All data was generated in standard growth conditions (log phase (OD₆₀₀~0.5), rich medium (Brucella broth), microaerobic atmosphere) in the wild-type (WT) strain, unless otherwise noted.

Publication

The publication describing this project can be accessed via the following DOI: 10.1101/2022.11.09.515450

Documentation

For detailed instructions on how to use CampyBrowse, check out our README.

Data availability

Our custom annotation used for the analysis can be downloaded here.

An annotation updated based on translatomics can be downloaded here.

The raw sequencing data used in this study can be accessed via the GEO accession number GSE208756.

The processed files used in this JBrowse2 genome browser can be downloaded here.

JBrowse2

Access Area

Preset 1: dRNA-seq & Ribo-seq

Differential RNA-seq (dRNA-seq) was used to map transcription start sites based on enrichment in +TEX vs. -TEX libraries. In addition, full-read coverage for Ribo-seq (ribosome footprints) and parallel RNA-seq (total RNA) is provided for identification of translated genes.

Access Preset 1

Preset 2: Ribo-seq & TIS (translation initiation site)

TIS profiling (Weaver et al., 2019; Meydan et al., 2019) and parallel Ribo-seq data identify ORFs and start codons. To stall ribosomes at TIS, cultures were treated with Ret (retapamulin; Exp2) or Onc (oncocin; Exp3). Tracks display coverage for 3'-end positions of reads only; TIS peaks are expected at ~16/17 nt (Ret/Onc, respectively) downstream of start codons. Peaks are generally enriched in the TIS vs. Ribo-seq (no drug) library. RNA- and Ribo-seq with full-read mapping is also displayed. For Exp2 (TIS(Ret)), a ΔcmeB strain (efflux pump mutant) was used.

Access Preset 2

Preset 3: Ribo-seq, TIS & TTS (translation termination site) profiling

This environment provides data from all translatomics approaches to reveal ORFs and their boundaries (start and stop codons). For TTS, ribosomes were stalled at stop codons with apidaecin (Api) (Mangano et al. 2020) and includes libraries generated from monosomes [TTS(Mono)] and disome [TTS(Di)] footprints from collisions at stop codons. A parallel TIS (Onc) library can be used to identify start codons, where Api might also enrich ribosomes. Tracks show either 5' or 3'-end mapping. For 3' ends, a peak (for TTS(Mono), enriched in TTS vs. Ribo-seq/TIS) is expected ~13 nt downstream of stop codons. For 5' ends ([TTS(Di)] only), a peak is expected ~45 nt upstream of the stop codons. RNA- and Ribo-seq with full-read mapping is also displayed.

Access Preset 3

Preset 4: All libraries

This environment provides data from all translatomics experiments to reveal ORFs and their boundaries (start and stop codons).

Access Preset 4

Annotation tracks

We provide multiple annotation tracks for the Campylobacter jejuni genome.

All annotation files can be loaded and unloaded using their respective check box in the track selector.

Annotation files are indexed and specific genes can be searched by gene name, locus tag or gene ID using the search box in the top center of the JBrowse2 instance.

This JBrowse2 instance includes:

Track	Description
The Reference Annotation	The reference annotation for Campylobacter jejuni
Re-annotation	ORF modifications and additions based on Ribo-seq/TIS/TTS
Novel CJsORFs	New ORFs ≤70 aa based on Ribo-seq/TIS/TTS
RBS Motif	All AAGG motifs for the reference genome

Coverage tracks

Coverage tracks show the normalized read counts per nucleotide position. There are multiple mapping types used (threeprime/fiveprime/global). The mapping method of each file is marked in their description.

Shareable Links

JBrowse2 allows you to easily share observations with your colleagues and collaborators.
Using the "share"-button next to the session name, you can automatically generate a link. Anyone that uses this link will see the exact session you are currently looking at. This includes colors, zoom level, scale, loaded tracks etc...
This is a very powerful tool for sharing potential discoveries.

JBrowse2 user guide

For more information on how to use JBrowse2, please have a look at the comprehensive user guide provided by the JBrowse team. The chapter on basic usage is highly recommended to get familiar with the JBrowse2 interface.

Contributors

This project is a collaboration between the universities Würzburg and Freiburg.

AG Sharma

Prof. Dr. Cynthia Sharma
Dr. Sarah Svensson
Dr. Kathrin Froschauer
Dr. Elisabetta Fiore
Philipp Kible

AG Engelmann

Prof. Dr. Susanne Engelmann
Dr. Martin Kucklick
Alicia Klaude

Robert Koch Institut

Dr. Stephan Fuchs

Shanghai Institute of Immunity and Infection

Prof. Dr. Daniel Falush
Dr. Chao Yang

AG Backofen

Prof. Dr. Rolf Backofen
Dr. Florian Eggenhofer
Rick Gelhausen

Acknowledgments

We thank the DFG funded Z2 project SPP2002 "Small Proteins in Prokaryotes, an Unexplored World" for enabling this project and de.NBI for providing computational resources.

Z2 project SPP2002
Small Proteins in prokaryotes, an Unexplored World

DFG

de.NBI

References

Froschauer, K^*., Svensson. SL^*., et al. Complementary Ribo-seq approaches refine the translatome and provide a small protein census in Campylobacter jejuni. submitted
Dugar, G., Herbig, A., Förstner, KU., Heidrich, N., Reinhardt. R., et al. (2013) High-Resolution Transcriptome Maps Reveal Strain-Specific Regulatory Features of Multiple Campylobacter jejuni Isolates. PLOS Genetics 9(5): e1003495.
https://doi.org/10.1371/journal.pgen.1003495
JBrowse 2: A modular genome browser with views of synteny and structural variation (2022). bioRxiv.
https://doi.org/10.1101/2022.07.28.501447
Weaver, J., Mohammad, F., Buskirk, AR., Storz, G. Identifying Small Proteins by Ribosome Profiling with Stalled Initiation Complexes. mBio.
https://doi.org/10.1128/mbio.02819-18
Meydan S, Marks J, Klepacki D, Sharma V, Baranov PV, Firth AE, Margus T, Kefi A, Vázquez-Laslop N, Mankin AS. Retapamulin-Assisted Ribosome Profiling Reveals the Alternative Bacterial Proteome. Mol Cell. 74: 481-493.e6.
https://doi.org/10.1016/j.molcel.2019.02.017
Mangano, K., Florin, T., Shao, X., Klepacki, D., Chelysheva, I., Ignatova, Z., Gao, Y., Mankin, AS., Vázquez-Laslop, N. Genome-wide effects of the antimicrobial peptide apidaecin on translation termination in bacteria. Elife. 2020 Oct 8;9:e62655.
https://doi.org/10.7554/elife.62655

Welcome to Campylobacter jejuni