BldD-based bimolecular fluorescence complementation for in vivo detection of the second messenger cyclic di-GMP.
BldD
EAL
GGDEF
bimolecular fluorescence complementation
biosensor
c-di-GMP
diguanylate cyclase
phosphodiesterase
Journal
Molecular microbiology
ISSN: 1365-2958
Titre abrégé: Mol Microbiol
Pays: England
ID NLM: 8712028
Informations de publication
Date de publication:
03 2022
03 2022
Historique:
revised:
21
12
2021
received:
06
12
2021
accepted:
23
12
2021
pubmed:
29
12
2021
medline:
28
4
2022
entrez:
28
12
2021
Statut:
ppublish
Résumé
The widespread bacterial second messenger bis-(3'-5')-cyclic diguanosine monophosphate (c-di-GMP) is an important regulator of biofilm formation, virulence and cell differentiation. C-di-GMP-specific biosensors that allow detection and visualization of c-di-GMP levels in living cells are key to our understanding of how c-di-GMP fluctuations drive cellular responses. Here, we describe a novel c-di-GMP biosensor, CensYBL, that is based on c-di-GMP-induced dimerization of the effector protein BldD from Streptomyces resulting in bimolecular fluorescence complementation of split-YPet fusion proteins. As a proof-of-principle, we demonstrate that CensYBL is functional in detecting fluctuations in intracellular c-di-GMP levels in the Gram-negative model bacteria Escherichia coli and Salmonella enterica serovar Typhimurium. Using deletion mutants of c-di-GMP diguanylate cyclases and phosphodiesterases, we show that c-di-GMP dependent dimerization of CBldD-YPet results in fluorescence complementation reflecting intracellular c-di-GMP levels. Overall, we demonstrate that the CensYBL biosensor is a user-friendly and versatile tool that allows to investigate c-di-GMP variations using single-cell and population-wide experimental set-ups.
Substances chimiques
bis(3',5')-cyclic diguanylic acid
61093-23-0
Cyclic GMP
H2D2X058MU
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
705-713Informations de copyright
© 2021 The Authors. Molecular Microbiology published by John Wiley & Sons Ltd.
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