Polymerizing the fibre between bacteria and host cells: the biogenesis of functional amyloid fibres

Amyloid fibres are proteinaceous aggregates associated with several human diseases, including Alzheimer's, Huntington's and Creutzfeldt Jakob's. Disease-associated amyloid formation is the result of proteins that misfold and aggregate into β sheet-rich fibre polymers. Cellular toxicity is readily associated with amyloidogenesis, although the molecular mechanism of toxicity remains unknown. Recently, a new class of ‘functional’ amyloid fibres was discovered that demonstrates that amyloids can be utilized as a productive part of cellular biology. These functional amyloids will provide unique insights into how amyloid formation can be controlled and made less cytotoxic. Bacteria produce some of the best-characterized functional amyloids, including a surface amyloid fibre called curli. Assembled by enteric bacteria, curli fibres mediate attachment to surfaces and host tissues. Some bacterial amyloids, like harpins and microcinE492, have exploited amyloid toxicity in a directed and functional manner. Here, we review and discuss the functional amyloids assembled by bacteria. Special emphasis will be paid to the biology of functional amyloid synthesis and the connections between bacterial physiology and pathology.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/75549/1/j.1462-5822.2008.01148.x.pd

Assembly, Spatial Distribution, and Secretion Activity of the Curlin Secretion Lipoprotein, CsgG.

Escherichia coli assembles functional amyloid fibers called curli. Produced by many Enterobacteriaceae spp., curli fibers are associated with biofilm formation, host cell adhesion and invasion, and immune system activation. CsgB nucleates the major curli subunit protein, CsgA, into a self-propagating amyloid fiber on the cell surface. CsgA and CsgB cell surface association and subsequent fiber polymerization are efficient processes, yet little is known about how the subunits reach and become positioned on the surface. I have characterized the role of CsgG in curli subunit secretion across the outer membrane, and found that CsgG is the central molecule of the curli secretion and assembly complex. CsgG formed an oligomeric complex in the outer membrane that interacted with at least three other csg encoded proteins. I found that the CsgG lipoprotein spanned the outer membrane, was sufficient for curlin transport across the outer membrane, and that a specific signal peptide on the N-terminus of the major fiber subunit directed protein secretion via the CsgG secretion apparatus. I also discovered that CsgG was non-uniformly distributed on the surface of curli-producing cells, and that the assembly, spatial organization, and secretion activity of CsgG was modified by other components of the curli assembly machine. One of the other curli assembly proteins, CsgE, was found to specifically function in the secretion pathway and to modulate both the secretion activity of CsgG and to act directly on CsgA to prevent self-polymerization. These results suggest a model where secretion and fiber assembly are tightly coupled and highly ordered processes.Ph.D.Molecular, Cellular, and Developmental BiologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/61667/1/emashman_1.pd

Spatial Clustering of the Curlin Secretion Lipoprotein Requires Curli Fiber Assembly ▿ †

Gram-negative bacteria assemble functional amyloid surface fibers called curli. CsgB nucleates the major curli subunit protein, CsgA, into a self-propagating amyloid fiber on the cell surface. The CsgG lipoprotein is sufficient for curlin transport across the outer membrane and is hypothesized to be the central molecule of the curli fiber secretion and assembly complex. We tested the hypothesis that the curli secretion protein, CsgG, was restricted to certain areas of the cell to promote the interaction of CsgA and CsgB during curli assembly. Here, electron microscopic analysis of curli-producing strains showed that relatively few cells in the population contacted curli fibers and that curli emanated from spatially discrete points on the cell surface. Microscopic analysis revealed that CsgG was surface exposed and spatially clustered around curli fibers. CsgG localization to the outer membrane and exposure of the surface domain were not dependent on any other csg-encoded protein, but the clustering of CsgG required the csg-encoded proteins CsgE, CsgF, CsgA, and CsgB. CsgG formed stable oligomers in all the csg mutant strains, but these oligomers were distinct from the CsgG complexes assembled in wild-type cells. Finally, we found that efficient fiber assembly was required for the spatial clustering of CsgG. These results suggest a new model where curli fiber formation is spatially coordinated with the CsgG assembly apparatus

Atomic Resolution Insights into Curli Fiber Biogenesis

Bacteria produce functional amyloid fibers called curli in a controlled, noncytotoxic manner. These extracellular fimbriae enable biofilm formation and promote pathogenicity. Understanding curli biogenesis is important for appreciating microbial lifestyles and will offer clues as to how disease-associated human amyloid formation might be ameliorated. Proteins encoded by the curli specific genes (csgA-G) are required for curli production. We have determined the structure of CsgC and derived the first structural model of the outer-membrane subunit translocator CsgG. Unexpectedly, CsgC is related to the N-terminal domain of DsbD, both in structure and oxido-reductase capability. Furthermore, we show that CsgG belongs to the nascent class of helical outer-membrane macromolecular exporters. A cysteine in a CsgG transmembrane helix is a potential target of CsgC, and mutation of this residue influences curli assembly. Our study provides the first high-resolution structural insights into curli biogenesis