Abstract
Bacterial proteins exported to the cell surface play key cellular functions. However, despite the interest to study the localisation of surface proteins such as adhesins, transporters or hydrolases, monitoring their dynamics in live imaging remains challenging, due to the limited availability of fluorescent probes remaining functional after secretion. In this work, we used the Escherichia coli intimin and the Listeria monocytogenes InlB invasin as surface exposed scaffolds fused with the recently developed chemogenetic fluorescent reporter protein FAST. Using both membrane permeant (HBR-3,5DM) and non-permeant (HBRAA-3E) fluorogens that fluoresce upon binding to FAST, we demonstrated that fully functional FAST can be exposed at the cell surface and used to specifically tag the external side of the bacterial envelop in both diderm and monoderm bacteria. Our work opens new avenues to study the organization and dynamics of the bacterial cell surface proteins.
Highlights
Bacterial proteins exported to the cell surface play key cellular functions
We showed that FAST can be exposed on the surface of gram-negative (Escherichia coli) and gram-positive (Listeria monocytogenes) model bacteria
To assess whether FAST can be exported and exposed at the cell surface of E. coli we used an anchoring module based on the scaffold of the intimin protein, an outer membrane protein expressed by Enterohemorrhagic E. coli (EHEC) and Enteropathogenic E. coli (EPEC), and required for intimate attachment to the host cell
Summary
Bacterial proteins exported to the cell surface play key cellular functions. despite the interest to study the localisation of surface proteins such as adhesins, transporters or hydrolases, monitoring their dynamics in live imaging remains challenging, due to the limited availability of fluorescent probes remaining functional after secretion. These results demonstrate the versatility of the FAST fluorescent reporter system to label proteins exposed on the surface of bacteria and to follow their localisation in living cells during colony growth.
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