Research Projects
Assembly of Type IV Pili
Work in our lab was the first to demonstrate clearly that Gram-positive bacteria, including nearly all of the Clostridia, make Type IV pili (TFP). TFP are fibers made of a single protein, called a pilin, that extend out of a cell and provide functions such as motility, adherence to host cells, and DNA uptake (natural transformation). We found that C. perfringens actually has two TFP systems, one for adherence and twitching motility and the other one is potentially involved in secretion of specific proteins, analogous to Type II secretion systems in Gram-negative bacteria. Current studies are focused on the role that individual pilus-associated proteins play in these processes.
References:
Soncini SR*, Hartman AH*, Gallagher TM*, Camper GJ*, Jensen RV, Melville SB. Changes in the expression of genes encoding type IV pili-associated proteins are seen when Clostridium perfringens is grown in liquid or on surfaces. BMC Genomics. 2020 Jan 14;21(1):45. doi: 10.1186/s12864-020-6453-z.
Hendrick*, W.A., M.W. Orr, S.R. Murray*, V.T. Lee & S.B. Melville, (2017) Cyclic Di-GMP Binding by an Assembly ATPase (PilB2) and Control of Type IV Pilin Polymerization in the Gram-Positive Pathogen Clostridium perfringens. J Bacteriol 199.
Melville, S. B. and Craig, L. Type IV pili in Gram-positive bacteria. 2013. Microbiol Mol Biol Rev. 77:323-341.
Varga, J. J., *Nguyen, V., *O'Brien, D. K., *Rodgers, K., Walker, R. A. and Melville, S. B. (2006) Type IV pili-dependent gliding motility in the Gram-positive pathogen Clostridium perfringens and other Clostridia. Mol Microbiol. 62 (3): 680-694.
Gliding motility
Our lab discovered that C. perfringens has the ability to glide on surfaces like agar plates even though it lacks bacterial flagella. It forms long chains of cells connected end-to-end which can move rapidly across a surface, which we thinks helps the bacteria obtain a new source of nutrients.
References:
McMahon, S. G., S. B. Melville, and J. Chen. 2022. Mechanical limitation of bacterial motility mediated by growing cell chains. Biophys J 121:2461-2473.
Liu, H., K. D. McCord*, J. Howarth, D. L. Popham, R. V. Jensen, and S. B. Melville. 2014. Hypermotility in Clostridium perfringens strain SM101 is due to spontaneous mutations in genes linked to cell division. J. Bacteriol. 96:2405-2412.
Liu, H., Bouillaut, L., Sonenshein, A. L. and Melville, S. B. 2013. Use of a mariner-based transposon mutagenesis system to isolate Clostridium perfringens mutants deficient in gliding motility J. Bacteriol. 195: 629-636.
Toxin Secretion
Like other Clostridia such as C. difficile, C. perfringens secretes a large clostridial toxin called TpeL. The secretion mechanism is unusual in that involves the activity of a holin-like protein, which are usually used by bacteriophages to lyse infected bacteria and release the phage. The holin-like protein, TpeE, has a membrane topology identical to that of the bacterial twin-arginine transport (Tat) system which secretes proteins that need to fold in the cytoplasm of the cell. The model shows TpeE oligomerizing to form a pore through which TpeL passes. Current studies in the lab are focusing on the mechanism of TpeE oligomerization and de-oligomerization, since the pore must be removed after TpeL transport to prevent killing the cell.
Reference:
Saadat, A.* and S. B. Melville. 2021. Holin-dependent secretion of the large clostridial toxin TpeL by Clostridium perfringens J Bacteriol Mar 23;203(8):e00580-20. doi: 10.1128/JB.00580-20.
Biofilm Formation and Secretion of Biofilm Matrix Proteins.
C. perfringens forms a biofilm made up of protein, carbohydrates and DNA. We showed that Type IV pili (TFP) are also part of the biofilm matrix and are studying the secretion of biofilm matrix proteins via TFP.
References:
Varga, J. J., *Therit, B. H., and Melville, S. B. 2008. Type IV pili and the CcpA protein are needed for maximal biofilm formation by the gram-positive anaerobic pathogen Clostridium perfringens. Infect Immun 76:4944-4951.
Early stage in biofilm formation by C. perfringens.