title: Cleavage of human fibronectin and other basement membrane-associated proteins by a Cryptococcus neoformans serine proteinase.
abstract: The interaction between the fungal pathogen Cryptococcus neoformans and human fibronectin (HFN) was examined in this study. Polypeptides from cryptococcal whole homogenates and cell wall with molecular masses of 25 and 35 kDa, respectively reacted with HFN. The relevance of the occurrence of these proteins in intact cells was uncertain, since yeast cells from different strains and serotypes of C. neoformans did not significantly adhere to soluble or solid-phased HFN. In contrast, an exocellular proteolytic activity that cleaves HFN was suggested. Degradation of HFN by culture supernatant fluids was demonstrated by Western blotting using a monoclonal anti-HFN antibody. Several fragments of lower molecular weights were observed which reacted with the antibody. Proteolysis was mediated by a serine protease activity, since HFN cleavage was completely inhibited by phenylmethylsulfonyl fluoride (PMSF), aprotinin, and N-tosyl-L-phenylalanyl chloromethylketone (TPCK), but not by inhibitors of metalo, cysteine, or aspartyl proteases. Similar results were obtained when the fluorogenic peptide carbobenzoxy-phenylalanyl-arginyl-7-amido-4-methylcoumarin (CBZ-Phe-Arg-NHmet-C) was used as substrate. The cryptococcal supernatant also cleaved laminin and type IV collagen, as demonstrated by polyacrylamide gel electrophoresis with co-polymerized proteins. The hydrolysis of these proteins was mediated by a single cryptococcal protease with a molecular mass of 75 kDa. The cleavage of key host components of the basement membrane and extracellular matrix by C. neoformans may be a relevant factor in the process of fungal invasion.
authors.affiliation: Departamento de Microbiologia Geral, Instituto de Microbiologia Professor Paulo de Góes, Universidade Federal do Rio de Janeiro, Cidade Universitária, CCS, Bloco G-Ilha do Fundão, Rio de Janeiro, RJ 21941-590, Brazil. marcio@micro.ufrj.br
authors.forename: Marcio L
authors.initials: ML
authors.lastname: Rodrigues
authors.forename: Flavia C G
authors.initials: FC
authors.lastname: dos Reis
authors.forename: Rosana
authors.initials: R
authors.lastname: Puccia
authors.forename: Luiz R
authors.initials: LR
authors.lastname: Travassos
authors.forename: Celuta S
authors.initials: CS
authors.lastname: Alviano
mesh_terms: D007611:Aprotinin / Q000494:pharmacology; D001422:Bacterial Adhesion / Q000502:physiology; D001485:Basement Membrane / Q000378:metabolism; D024141:Collagen Type IV / Q000378:metabolism; D003453:Cryptococcosis / Q000473:pathology; D003455:Cryptococcus neoformans / Q000201:enzymology*; D003470:Culture Media / Q000378:metabolism; D004151:Dipeptides / Q000378:metabolism; D016326:Extracellular Matrix Proteins / Q000378:metabolism*; D005353:Fibronectins / Q000378:metabolism*; D006801:Humans; D007797:Laminin / Q000378:metabolism; D012697:Serine Endopeptidases / Q000378:metabolism*
chemical_list: D024141:Collagen Type IV; D003470:Culture Media; D004151:Dipeptides; D016326:Extracellular Matrix Proteins; D005353:Fibronectins; D007797:Laminin; D007611:Aprotinin; D012697:Serine Endopeptidases