In the metabolically versatile bacterium Pseudomonas aeruginosa, the RNA-binding protein Crc is involved in catabolite repression of a range of degradative genes, such as amiE (encoding aliphatic amidase). We found that a CA-rich sequence (termed CA motif) in the amiE translation initiation region was important for Crc binding. The small RNA CrcZ (407 nt) containing 5 CA motifs was able to bind the Crc protein with high affinity and to remove it from amiE mRNA in vitro. Overexpression of crcZ relieved catabolite repression in vivo, whereas a crcZ mutation pleiotropically prevented the utilization of several carbon sources. The sigma factor RpoN and the CbrA/CbrB two-component system, which is known to maintain a healthy carbon-nitrogen balance, were necessary for crcZ expression. During growth on succinate, a preferred carbon source, CrcZ expression was low, resulting in catabolite repression of amiE and other genes under Crc control. By contrast, during growth on mannitol, a poor carbon source, elevated CrcZ levels correlated with relief of catabolite repression. During growth on glucose, an intermediate carbon source, CrcZ levels and amiE expression were intermediate between those observed in succinate and mannitol media. Thus, the CbrA-CbrB-CrcZ-Crc system allows the bacterium to adapt differentially to various carbon sources. This cascade also regulated the expression of the xylS (benR) gene, which encodes a transcriptional regulator involved in benzoate degradation, in an analogous way, confirming this cascade's global role.

Carbon Catabolite repression (CCR) allows a fast adaptation of Bacteria to changing nutrient supplies. The Pseudomonas aeruginosa (PAO1) catabolite repression control protein (Crc) was deemed to act as a translational regulator, repressing functions involved in uptake and utilization of carbon sources. However, Crc of PAO1 was recently shown to be devoid of RNA binding activity. In this study the RNA chaperone Hfq was identified as the principle post-transcriptional regulator of CCR in PAO1. Hfq is shown to bind to A-rich sequences within the ribosome binding site of the model mRNA amiE, and to repress translation in vitro and in vivo. We further report that Crc plays an unknown ancillary role, as full-fledged repression of amiE and other CCR-regulated mRNAs in vivo required its presence. Moreover, we show that the regulatory RNA CrcZ, transcription of which is augmented when CCR is alleviated, binds to Hfq with high affinity. This study on CCR in PAO1 revealed a novel concept for Hfq function, wherein the regulatory RNA CrcZ acts as a decoy to abrogate Hfq-mediated translational repression of catabolic genes and thus highlights the central role of RNA based regulation in CCR of PAO1.

SummaryThe Pseudomonas aeruginosa quorum-sensing (QS) systems, Las and Rhl, control the production of several virulence factors and other proteins, which are important to sustain adverse conditions. A comparative transcriptome analysis of a rpoS -and a rpoS

No abstract