Neuropeptides and bacterial interactions: The example of the impact of the C-type natriuretic peptide (CNP) on Pseudomonas aeruginosa biofilm formation

International audience; The opening of a new field of research known as microbial endocrinology, has prompted us to investigate the impact of several neuropeptides produced by the host on various bacteria isolated from skin or pathogens in the respiratory tract. We observed that substance P and CGRP two skin nerves neuropeptides are able to modify Staphylococcus epidermidis, S. aureus and Bacillus cereus biofilm formation and virulence regulation through binding to specific bacterial sensors. We also demonstrated that the lung produced C-type Natriuretic Peptide (CNP) enhances Pseudomonas aeruginosa virulence. In the present study, we focus on the CNP action mechanism on P. aeruginosa biofilm formation and on the identification of the bacterial sensor for this human hormone. We observed that exposure of P. aeruginosa to graded concentrations of CNP (10-9 to 10-6 M) induced a dose-dependent inhibition of biofilm formation. This effect was totally abolished by isatin, an antagonist of eukaryotic cells natriuretic peptide receptors. Screening and comparing 3D structures of human natriuretic peptides receptors and P. aeruginosa proteins revealed that the bacterial protein AmiC shows significant homology with the human C-type eukaryotic natriuretic peptide receptor (hNPR-C). Recombinant protein AmiC was obtained and the protein interactions assessed using microscale thermophoresis. The results showed that both CNP and the hNPR-C agonist osteocrin bind AmiC with respective K D of 2 µM and 5 nM confirming the role of AmiC as natriuretic peptides sensor. The amiC gene belongs to the ami operon where amiE encodes the AmiE amidase which hydrolyses short-chain aliphatic amides to their corresponding organic acids. We further observed that over expression of AmiE resulted in altered biofilm formation and strongly reduced bacterial virulence, suggesting that AmiE should be the final effector of the CNP effects on P. aeruginosa. Taken together, these data show that neuropeptides, firstly identified as host signal molecules, could act as bacterial modulator and in certain conditions act as potential antimicrobial peptides.