A grad-seq view of RNA and protein complexes in Pseudomonas aeruginosa under standard and bacteriophage predation conditions

Please always quote using this URN: urn:nbn:de:bvb:20-opus-259054
  • The Gram-negative rod-shaped bacterium Pseudomonas aeruginosa is not only a major cause of nosocomial infections but also serves as a model species of bacterial RNA biology. While its transcriptome architecture and posttranscriptional regulation through the RNA-binding proteins Hfq, RsmA, and RsmN have been studied in detail, global information about stable RNA-protein complexes in this human pathogen is currently lacking. Here, we implement gradient profiling by sequencing (Grad-seq) in exponentially growing P. aeruginosa cells toThe Gram-negative rod-shaped bacterium Pseudomonas aeruginosa is not only a major cause of nosocomial infections but also serves as a model species of bacterial RNA biology. While its transcriptome architecture and posttranscriptional regulation through the RNA-binding proteins Hfq, RsmA, and RsmN have been studied in detail, global information about stable RNA-protein complexes in this human pathogen is currently lacking. Here, we implement gradient profiling by sequencing (Grad-seq) in exponentially growing P. aeruginosa cells to comprehensively predict RNA and protein complexes, based on glycerol gradient sedimentation profiles of >73% of all transcripts and ∼40% of all proteins. As to benchmarking, our global profiles readily reported complexes of stable RNAs of P. aeruginosa, including 6S RNA with RNA polymerase and associated product RNAs (pRNAs). We observe specific clusters of noncoding RNAs, which correlate with Hfq and RsmA/N, and provide a first hint that P. aeruginosa expresses a ProQ-like FinO domain-containing RNA-binding protein. To understand how biological stress may perturb cellular RNA/protein complexes, we performed Grad-seq after infection by the bacteriophage ΦKZ. This model phage, which has a well-defined transcription profile during host takeover, displayed efficient translational utilization of phage mRNAs and tRNAs, as evident from their increased cosedimentation with ribosomal subunits. Additionally, Grad-seq experimentally determines previously overlooked phage-encoded noncoding RNAs. Taken together, the Pseudomonas protein and RNA complex data provided here will pave the way to a better understanding of RNA-protein interactions during viral predation of the bacterial cell. IMPORTANCE Stable complexes by cellular proteins and RNA molecules lie at the heart of gene regulation and physiology in any bacterium of interest. It is therefore crucial to globally determine these complexes in order to identify and characterize new molecular players and regulation mechanisms. Pseudomonads harbor some of the largest genomes known in bacteria, encoding ∼5,500 different proteins. Here, we provide a first glimpse on which proteins and cellular transcripts form stable complexes in the human pathogen Pseudomonas aeruginosa. We additionally performed this analysis with bacteria subjected to the important and frequently encountered biological stress of a bacteriophage infection. We identified several molecules with established roles in a variety of cellular pathways, which were affected by the phage and can now be explored for their role during phage infection. Most importantly, we observed strong colocalization of phage transcripts and host ribosomes, indicating the existence of specialized translation mechanisms during phage infection. All data are publicly available in an interactive and easy to use browser.show moreshow less

Download full text files

Export metadata

Additional Services

Share in Twitter Search Google Scholar Statistics
Metadaten
Author: Milan GerovaORCiD, Laura WickeORCiD, Kotaro Chihara, Cornelius Schneider, Rob LavigneORCiD, Jörg VogelORCiD
URN:urn:nbn:de:bvb:20-opus-259054
Document Type:Journal article
Faculties:Medizinische Fakultät / Institut für Molekulare Infektionsbiologie
Fakultät für Chemie und Pharmazie / Lehrstuhl für Biochemie
Language:English
Parent Title (English):mbio
Year of Completion:2021
Volume:12
Issue:1
Pagenumber:e03454-20
Source:mBio (2021) 12:1, e03454-20. https://doi.org/10.1128/mBio.03454-20
DOI:https://doi.org/10.1128/mBio.03454-20
Dewey Decimal Classification:6 Technik, Medizin, angewandte Wissenschaften / 61 Medizin und Gesundheit / 610 Medizin und Gesundheit
Tag:Grad-seq; Pseudomonas; Pseudomonas aeruginosa; RNA-binding proteins; UKZ; bacteriophage; infection; noncoding RNA; phage
Release Date:2022/05/04
EU-Project number / Contract (GA) number:819800
OpenAIRE:OpenAIRE
Collections:Open-Access-Publikationsfonds / Förderzeitraum 2021
Licence (German):License LogoCC BY: Creative-Commons-Lizenz: Namensnennung 4.0 International