ABSTRACT
In Pseudomonas aeruginosa the RNA chaperone Hfq and the catabolite repression control protein (Crc) act as post-transcriptional regulators during carbon catabolite repression (CCR). In this regard Crc is required for full-fledged Hfq-mediated translational repression of catabolic genes. RNAseq based transcriptome analyses revealed a significant overlap between the Crc and Hfq regulons, which in conjunction with genetic data supported a concerted action of both proteins. Biochemical and biophysical approaches further suggest that Crc and Hfq form an assembly in the presence of RNAs containing A-rich motifs, and that Crc interacts with both, Hfq and RNA. Through these interactions, Crc enhances the stability of Hfq/Crc/RNA complexes, which can explain its facilitating role in Hfq-mediated translational repression. Hence, these studies revealed for the first time insights into how an interacting protein can modulate Hfq function. Moreover, Crc is shown to interfere with binding of a regulatory RNA to Hfq, which bears implications for riboregulation. These results are discussed in terms of a working model, wherein Crc prioritizes the function of Hfq toward utilization of favored carbon sources.
Subject(s)
Bacterial Proteins/genetics , Catabolite Repression , Host Factor 1 Protein/genetics , Protein Biosynthesis , Pseudomonas aeruginosa/genetics , RNA, Bacterial/genetics , Repressor Proteins/genetics , Bacterial Proteins/chemistry , Bacterial Proteins/metabolism , Binding Sites , Bordetella pertussis/genetics , Bordetella pertussis/metabolism , Carbohydrate Metabolism/genetics , Escherichia coli/genetics , Escherichia coli/metabolism , Gene Expression Regulation, Bacterial , Host Factor 1 Protein/chemistry , Host Factor 1 Protein/metabolism , Kinetics , Models, Molecular , Nucleotide Motifs , Protein Binding , Protein Interaction Domains and Motifs , Protein Structure, Secondary , Pseudomonas aeruginosa/metabolism , RNA, Bacterial/chemistry , RNA, Bacterial/metabolism , Regulon , Repressor Proteins/chemistry , Repressor Proteins/metabolism , TranscriptomeABSTRACT
The RNA chaperone Hfq regulates virulence and metabolism in the opportunistic pathogen Pseudomonas aeruginosa. During carbon catabolite repression (CCR) Hfq together with the catabolite repression control protein Crc can act as a translational repressor of catabolic genes. Upon relief of CCR, the level of the Hfq-titrating RNA CrcZ is increasing, which in turn abrogates Hfq-mediated translational repression. As the interdependence of Hfq-mediated and RNA based control mechanisms is poorly understood, we explored the possibility whether the regulatory RNA CrcZ can interfere with riboregulation. We first substantiate that the P. aeruginosa Hfq is proficient and required for riboregulation of the transcriptional activator gene antR by the small RNA PrrF1-2. Our studies further revealed that CrcZ can interfere with PrrF1-2/Hfq-mediated regulation of antR. The competition for Hfq can be rationalized by the higher affinity of Hfq for CrcZ than for antR mRNA.
