Discovery and functional analysis of small RNAs and RNA binding proteins originating from phage

In recent years, evidence is mounting that small RNAs (sRNA) play an important regulatory role in eukaryotes as well as in bacteria. In model pathogens like Escherichia coli and Salmonella, these regulatory molecules can impact every stage of transcription and translation by base-pairing to their target. In the opportunistic pathogen Pseudomonas aeruginosa small RNAs have been shown to be involved in iron acquisition, quorum-sensing, carbon catabolism as well as the regulation of many virulence factors. For bacteriophages, our knowledge about regulation on the sRNA level is limited to just a few examples in (cryptic) prophages while nothing is known for sRNAs in lytic viruses. This project aims to identify small RNAs and RNA-binding proteins originating from 4 different clades of P. aeruginosa-infecting bacteriophages. For this, predicted sRNAs will be validated by Northern blot analysis and a global interactome technique, called Gradient sequencing, will be performed. Subsequently, the predicted interaction partners (originating from phage or host) will be confirmed using RT-qPCR. By determining the specific binding site and their interaction with an RNA-binding protein, these sRNAs will be further analysed. Finally, this project will investigate the signalling pathways which are affected by the phage-encoded sRNAs. This will offer new insights from a fundamental biological perspective and will inspire their potential use as biotechnological tools.