C-di-GMP signalling in Pseudomonas aeruginosa: connecting the dots in a multi modal network
File(s)
Author(s)
Glatzel, Kira
Type
Thesis
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.
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.
Version
Open Access
Date Issued
2022-04
Date Awarded
2023-06
Copyright Statement
Creative Commons Attribution NonCommercial Licence
License URL
Advisor
Filloux, Alain
Sponsor
Wellcome Trust (London, England)
Grant Number
WMNM_P67006
Publisher Department
Life Sciences
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)