Browsing by Subject "C-di-GMP"

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A diguanylate cyclase acts as a cell division inhibitor in a two-step response to reductive and envelope stresses
(2016-12) Kim, Hyo Kyung; Harshey, Rasika M.; Georgiou, George; Jayaram, Makkuni; O’Halloran, Theresa; Whiteley, Marvin
Bacteria use diverse nucleotide-based small molecules as second messengers to transduce various signals in their extra- and intracellular conditions, and to elicit appropriate cellular responses. The signaling molecule cyclic diguanylate (c-di-GMP) hasemerged as a ubiquitous nucleotide that controls a variety of cellular processes including motility, biofilm formation, virulence and cell differentiation. The intracellular levels of c-di-GMP are determined by the balance between its synthesis by diguanylate cyclases (DGCs) and degradation by phosphodiesterases (PDEs). Single bacterial species encode multiple DGC/PDEs harboring different sensory domains, surmised to integrate various input signals to regulate a common pool of c-di-GMP, which in turn regulates wideranging output processes. However, some c-di-GMP pathways appear to relay information selectively by spatial sequestration of particular DGC/PDEs. Here I discovered a new signaling pathway for YfiN, one of multiple DGCs found in E. coli and Salmonella. I show that YfiN interacts directly with components of the cell division machinery to inhibit division and growth, rather than acting through its product c-di-GMP. The DGC function of YfiN was known previously to be activated by a redox stress signal. My studies have revealed a second function, where redox-activated YfiN responds further to envelope stress by dynamically localizing to the division site and halting division. The unexpected bifunctionality of YfiN provides evidence that protein-protein interactions between c-di-GMP signaling components and their targets also confer signaling specificity, and reveals a new pathway for simultaneously inhibiting both cell division and cell growth in response to two stresses applied in a sequence.
Mechanosensing and early biofilm development in Pseudomonas aeruginosa
(2017-05) Rodesney, Christopher Anania; Gordon, Vernita Diane; Florin, Ernst-Ludwig; Marder, Michael; Fink, Manfred; Whiteley, Marvin
Biofilms are communities of sessile microbes that are phenotypically distinct from their genetically-identical, free-swimming counterparts. Biofilms initiate when bacteria attach to a solid surface, as this attachment triggers intracellular signaling to change gene expression of individual bacteria from the planktonic to the biofilm phenotype. However the initial cues leading allowing bacteria to sense a surface, as well as the role of spatial structure in biofilm development, are not well known. This dissertation has two main parts, the first presenting a method for growing biofilms from initiating cells whose positions are controlled with single-cell precision using laser trapping. Biofilm infections are notoriously intractable, in part due to changes in the bacterial phenotype that result from spatial structure. Understanding the role of structure in biofilm development requires methods to control the spatial structure of biofilms. The native growth, motility, and surface adhesion of positioned microbes are preserved, as we show for model organisms Pseudomonas aeruginosa and Staphylococcus aureus. We demonstrate that laser-trapping and placing bacteria on surfaces can reveal the e↵ects of spatial structure on bacterial growth in early biofilm development. In the second part we show that mechanical shear acts as a cue for surface adhesion in P. aeruginosa. For P. aeruginosa, it has long been known that intracellular levels of the signaling molecule cyclic-di-GMP increase upon surface adhesion and that increased cyclic-di-GMP is required to begin biofilm development. The magnitude of the shear force, and thereby the corresponding activation of cyclic-di-GMP signaling, can be adjusted both by varying the strength of the adhesion that binds bacteria to the surface and by varying the rate of fluid flow over surface-bound bacteria. We show that the envelope protein PilY1 and functional Type IV pili are required mechanosensory elements. Finally, we propose an analytic model that accounts for the feedback between mechanosensors, cyclic-di-GMP signaling, and production of adhesive polysaccharides, describing our data