Dual amyloid domains promote differential functioning of the chaplin proteins duringStreptomycesaerial morphogenesis

Abstract

The chaplin proteins are functional amyloids found in the filamentous Streptomyces bacteria. These secreted proteins are required for the aerial development of Streptomyces coelicolor, and contribute to an intricate rodlet ultrastructure that decorates the surfaces of aerial hyphae and spores. S. coelicolor encodes eight chaplin proteins. Previous studies have revealed that only three of these proteins (ChpC, ChpE, and ChpH) are necessary for promoting aerial development, and of these three, ChpH is the primary developmental determinant. Here, we show that the model chaplin, ChpH, contains two amyloidogenic domains: one in the N terminus and one in the C terminus of the mature protein. These domains have different polymerization properties as determined using fluorescence spectroscopy, secondary structure analyses, and electron microscopy. We coupled these in vitro assays with in vivo genetic studies to probe the connection between ChpH amyloidogenesis and its biological function. Using mutational analyses, we demonstrated that both N-and C-terminal amyloid domains of ChpH were required for promoting aerial hypha formation, while the N-terminal domain was dispensable for assembly of the rodlet ultrastructure. These results suggest that there is a functional differentiation of the dual amyloid domains in the chaplin proteins.Streptomyces development | sporulation | surface ultrastructure

Protein aggregation in bacteria: the thin boundary between functionality and toxicity

“…It was shown that ChpH chaplin was the major contributor to aerial hypha development, and structural analysis of the chaplin proved that ChpH indeed possess two amyloid domains, an Nand a C-terminal domain. The C-terminal ClpH domain contains a sequence which is critically important for effective amyloid polymerization and promotion of aerial hyphae (Capstick et al, 2011).…”

Dimethyl Sulfoxide and Ethanol Elicit Increased Amyloid Biogenesis and Amyloid-Integrated Biofilm Formation in Escherichia coli

“…Since the discovery of curli as adhesive fimbriae in 1989 (58) and the identification of curli as amyloid in 2002 (15), we now appreciate that functional amyloids are prevalent among microorganisms, e.g., Salmonella and Pseudomonas species, Streptomyces coelicolor, Mycobacterium tuberculosis, and Bacillus subtilis (1,4,11,15,21,58,64). The assembly of functional amyloids in microbes is regulated in order to direct polymerization at the right time and place and to prevent toxicity (4, 24).…”

Exploiting amyloid: how and why bacteria use cross-β fibrils

“…ChpH has two amyloid domains, one near the N-terminus and the other near the C-terminus. The two domains have different assembly kinetics and both domains are necessary for aerial hyphae formation, but the C-terminal domain appears to be more important for rodlet assembly than the N-terminal domain [38].…”

“…Results indicate that expression of both long and short chaplins, with at least one short chaplin containing the conserved cysteine motif, is vital for the development of a robust aerial mycelium, but that there is otherwise some degree of redundancy among the chaplins [37]. Two recent biophysical studies have shed further light on the role of the chaplins in the formation of the hydrophobic coat [36,38].…”

“…A recent study by Capstick et al [38] identified two amyloidogenic regions in ChpH that contribute differentially to the assembly of chaplin fibrils. Computational analysis led to the identification of a five residue SVIGL motif in the C-terminus of ChpH that was predicted to be highly aggregation-prone.…”