The formation of biofilms provides structural and adaptive bacterial response to the environment. In Bacillus species, the biofilm extracellular matrix is composed of exopolysaccharides, hydrophobins, and several functional amyloid proteins. We report, using multiscale approaches such as solid‐state NMR (SSNMR), electron microscopy, X‐ray diffraction, dynamic light scattering, attenuated total reflection Fourier transform infrared (FTIR), and immune‐gold labeling, the molecular architecture of B. subtilis and pathogenic B. cereus functional amyloids. SSNMR data reveal that the major amyloid component TasA in its fibrillar amyloid form contain β‐sheet and α‐helical secondary structure, suggesting a nontypical amyloid architecture in B. subtilis. Proteinase K digestion experiments indicate the amyloid moiety is ~100 aa long, and subsequent SSNMR and FTIR signatures for B. subtilis and B. cereus TasA filaments highlight a conserved amyloid fold, albeit with substantial differences in structural polymorphism and secondary structure composition. Structural analysis and coassembly data on the accessory protein TapA in B. subtilis and its counterpart camelysin in B. cereus reveal a catalyzing effect between the functional amyloid proteins and a common structural architecture, suggesting a coassembly in the context of biofilm formation. Our findings highlight nontypical amyloid behavior of these bacterial functional amyloids, underlining structural variations between biofilms even in closely related bacterial species.—El Mammeri, N., Hierrezuelo, J., Tolchard, J., Cámara‐Almirón, J., Caro‐Astorga, J., Álvarez‐Mena, A., Dutour, A., Berbon, M., Shenoy, J., Morvan, E., Grélard, A., Kauffmann, B., Lecomte, S., de Vicente, A., Habenstein, B., Romero, D., Loquet, A. Molecular architecture of bacterial amyloids in Bacillus biofilms. FASEB J. 33, 12146‐12163 (2019). http://www.fasebj.org
Bacteria can form biofilms that consist of multicellular communities embedded in an extracellular matrix (ECM). In Bacillus subtilis, the main protein component of the ECM is the functional amyloid TasA. Here, we study further the roles played by TasA in B. subtilis physiology and biofilm formation on plant leaves and in vitro. We show that ΔtasA cells exhibit a range of cytological symptoms indicative of excessive cellular stress leading to increased cell death. TasA associates to the detergent-resistant fraction of the cell membrane, and the distribution of the flotillin-like protein FloT is altered in ΔtasA cells. We propose that, in addition to a structural function during ECM assembly and interactions with plants, TasA contributes to the stabilization of membrane dynamics as cells enter stationary phase.
In nature, bacteria form biofilms—differentiated multicellular communities attached to surfaces. Within these generally sessile biofilms, a subset of cells continues to express motility genes. We found that this subpopulation enabled Bacillus subtilis biofilms to expand on high-friction surfaces. The extracellular matrix (ECM) protein TasA was required for the expression of flagellar genes. In addition to its structural role as an adhesive fiber for cell attachment, TasA acted as a developmental signal stimulating a subset of biofilm cells to revert to a motile phenotype. Transcriptomic analysis revealed that TasA stimulated the expression of a specific subset of genes whose products promote motility and repress ECM production. Spontaneous suppressor mutations that restored motility in the absence of TasA revealed that activation of the biofilm-motility switch by the two-component system CssR/CssS antagonized the TasA-mediated reversion to motility in biofilm cells. Our results suggest that although mostly sessile, biofilms retain a degree of motility by actively maintaining a motile subpopulation.
Bacillus spp. MBI 600 is a gram-positive bacterium and is characterized as a PGPR strain involved in plant growth promotion and control of various plant pathogens which has recently been introduced into the agricultural practice. In this study we performed a Next Generation Sequencing analysis, to analyze the full genome of this microorganism and to characterize it taxonomically. Results showed that MBI 600 strain was phylogenetically close to other Bacillus spp. strains used as biocontrol agents and identified as B. subtilis. GOG analysis showed clusters contributed to secondary metabolites production such as fengycin and surfactin. In addition, various genes which annotated according to other plant-associated strains, showed that play a main role in nutrient availability from soil. The root colonization ability of MBI 600 strain was analyzed in vivo with a yellow fluorescence protein (yfp) tag. Confocal laser scanning microscopy of cucumber roots treated with yfp-tagged MBI 600 cells, revealed that the strain exhibits a strong colonization ability of cucumber roots, although it is affected significantly by the growth substrate of the roots. In vitro and in planta experiments with MBI 600 strain and F. oxysporum f.sp. radicis cucumerinum and P. aphanidernatum, showed a high control ability against these soilborne pathogens. Overall, our study demonstrates the effectiveness of MBI 600 in plant growth promotion and antagonism against different pathogens, highlighting the use of this microorganism as a biocontrol agent.
Intense research has confirmed the formerly theoretical distribution of amyloids in nature, and studies on different systems have illustrated the role of these proteins in microbial adaptation and in interactions with the environment. Two lines of research are expanding our knowledge on functional amyloids: (i) structural studies providing insights into the molecular machineries responsible for the transition from monomer to fibers and (ii) studies showing the way in which these proteins might participate in the microbial fitness in natural settings. Much is known about how amyloids play a role in the social behavior of bacteria, or biofilm formation, and in the adhesion of bacteria to surfaces; however, we are still in the initial stages of understanding a complementary involvement of amyloids in bacteria-host interactions. This review will cover the following two topics: first, the key aspects of the microbial platforms dedicated to the assembly of the fibers, and second, the mechanisms by which bacteria utilize the morphological and biochemical variability of amyloids to modulate the immunological response of the host, plants and humans, contributing to (i) infection, in the case of pathogenic bacteria or (ii) promotion of the health of the host, in the case of beneficial bacteria.
Since they were discovered, amyloids have proven to be versatile proteins able to participate in a variety of cellular functions across all kingdoms of life. This multitask trait seems to reside in their ability to coexist as monomers, aggregates or fibrillar entities, with morphological and biochemical peculiarities. It is precisely this common molecular behaviour that allows amyloids to cross react with one another, triggering heterologous aggregation. In bacteria, many of these functional amyloids are devoted to the assembly of biofilms by organizing the matrix scaffold that keeps cells together. However, consistent with their notion of multifunctional proteins, functional amyloids participate in other biological roles within the same organisms, and emerging unprecedented functions are being discovered. In this review, we focus on functional amyloids reported in gram-positive bacteria, which are diverse in their assembly mechanisms and remarkably specific in their biological functions that they perform. Finally, we consider cross-seeding between functional amyloids as an emerging theme in interspecies interactions that contributes to the diversification of bacterial biology.
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