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A label-free approach for relative spatial quantitation of c-di-GMP in microbial biofilms.

ABSTRACT: Microbial biofilms represent an important lifestyle for bacteria and are dynamic three dimensional structures. Cyclic dimeric guanosine monophosphate (c-di-GMP) is a ubiquitous signaling molecule that is known to be tightly regulated with biofilm processes. While measurements of global levels of c-di-GMP have proven valuable towards understanding the genetic control of c-di-GMP production, there is a need for tools to observe the local changes of c-di-GMP production in biofilm processes. We have developed a label-free method for the direct detection of c-di-GMP in microbial colony biofilms using matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI). We applied this method to the enteric pathogen Vibrio cholerae, the marine symbiont V. fischeri, and the opportunistic pathogen Pseudomonas aeruginosa PA14 and detected spatial and temporal changes in c-di-GMP signal that accompanied genetic alterations in factors that synthesize and degrade the compound. We further demonstrated how this method can be simultaneously applied to detect additional metabolites of interest in a single experiment.

SUBMITTER: McCaughey CS

PROVIDER: S-EPMC10592747 | biostudies-literature | 2023 Oct

REPOSITORIES: biostudies-literature

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A label-free approach for relative spatial quantitation of c-di-GMP in microbial biofilms.

McCaughey Catherine S CS Trebino Michael A MA McAtamney Allyson A Isenberg Ruth R Mandel Mark J MJ Yildiz Fitnat H FH Sanchez Laura M LM

bioRxiv : the preprint server for biology 20231010

Microbial biofilms represent an important lifestyle for bacteria and are dynamic three dimensional structures. Cyclic dimeric guanosine monophosphate (c-di-GMP) is a ubiquitous signaling molecule that is known to be tightly regulated with biofilm processes. While measurements of global levels of c-di-GMP have proven valuable towards understanding the genetic control of c-di-GMP production, there is a need for tools to observe the local changes of c-di-GMP production in biofilm processes. We have ...[more]

PMID: 37873360

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Project description:Light is known to trigger regulatory responses in diverse organisms, including slime molds, animals, plants, and phototrophic bacteria. However, light-dependent processes in nonphototrophic bacteria, and those of pathogens in particular, have received comparatively little research attention. In this study, we examined the impact of light on multicellular development in Pseudomonas aeruginosa, a leading cause of biofilm-based bacterial infections. We grew P. aeruginosa strain PA14 in a colony morphology assay and found that growth under prolonged exposure to low-intensity blue light inhibited biofilm matrix production and thereby the formation of vertical biofilm structures (i.e., "wrinkles"). Light-dependent inhibition of biofilm wrinkling was correlated with low levels of cyclic di-GMP (c-di-GMP), consistent with the role of this signal in stimulating matrix production. A screen of enzymes with the potential to catalyze c-di-GMP synthesis or degradation identified c-di-GMP phosphodiesterases that contribute to light-dependent inhibition of biofilm wrinkling. One of these, RmcA, was previously characterized by our group for its role in mediating the effect of redox-active P. aeruginosa metabolites called phenazines on biofilm wrinkle formation. Our results suggest that an RmcA sensory domain that is predicted to bind a flavin cofactor is involved in light-dependent inhibition of wrinkling. Together, these findings indicate that P. aeruginosa integrates information about light exposure and redox state in its regulation of biofilm development.IMPORTANCE Light exposure tunes circadian rhythms, which modulate the immune response and affect susceptibility to infection in plants and animals. Though molecular responses to light are defined for model plant and animal hosts, analogous pathways that function in bacterial pathogens are understudied. We examined the response to light exposure in biofilms (matrix-encased multicellular assemblages) of the nonphotosynthetic bacterium Pseudomonas aeruginosa We found that light at intensities that are not harmful to human cells inhibited biofilm maturation via effects on cellular signals. Because biofilm formation is a critical factor in many types of P. aeruginosa infections, including burn wound infections that may be exposed to light, these effects could be relevant for pathogenicity.

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A label-free approach for relative spatial quantitation of c-di-GMP in microbial biofilms.

Publications

A label-free approach for relative spatial quantitation of c-di-GMP in microbial biofilms.

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