C-di-GMP signalling in Pseudomonas aeruginosa: connecting the dots in a multi modal network

Abstract

Bacteria rely on complex regulatory networks to control their cellular programme in order to respond to extracellular factors in an environment or host setting. Bis-(3′-5′)-cyclic dimeric guanosine monophosphate (c-di-GMP) is a signalling molecule used by several bacterial species to facilitate some of those important lifestyle decisions. One of the most complex and elaborated c-di-GMP signalling networks is found in the human opportunistic pathogen Pseudomonas aeruginosa, which is a major cause of chronic infection in cystic fibrosis patients.

The genome of P. aeruginosa alone encodes 41 proteins harbouring the potential to make or break c-di-GMP. Using a methodical deletion approach, single and multiple c-di-GMP metabolizing genes were knocked out and their role in attachment, biofilm, motility, virulence and antimicrobial resistance systematically assessed, providing a comprehensive pool of data for the c-di-GMP research community. Interestingly, a very large number of c-di-GMP deletion mutants showed an altered biofilm/attachment profile, indicating that the network is of great importance for motile to sessile transition.

We further uncovered that among all c-di-GMP regulating enzymes, PA0285 is one of the most potent phosphodiesterases regulating intracellular c-di-GMP levels leading to the most pronounced biofilm attachment phenotype. It is uniquely controlled by its dual domain structure containing both an EAL domain responsible for breaking c-di-GMP as well as a regulatory GGDEF domain characteristic of c-di-GMP synthesis. Both domains have been shown to be essential for the PA0285 mediated phenotypes in biofilm attachment and intracellular c-di-GMP.

Further, targeted protein-protein interaction analysis revealed the molecular mechanisms of DgcP (PA5487), a c-di-GMP synthesising cyclase. involved in the convergence of c-di-GMP and cyclic adenosine monophosphate (cAMP) signalling pathways facilitating touch-down and attachment behaviour of P. aeruginosa.