title: Cellular localization and expression of gp63 homologous metalloproteases in Leishmania (Viannia) braziliensis strains.
abstract: Leishmania (Viannia) braziliensis is the major causative agent of American tegumentary leishmaniasis, a disease that encompasses a broad spectrum of clinical manifestations. In a previous study, we showed that Brazilian and Colombian L. braziliensis strains, isolated from patients with distinct clinical manifestations, display different pattern of metalloprotease activities. Following these results, we investigated the cellular localization of these molecules and their relation to the major surface protease (gp63) of Leishmania. Comparative analyses of metalloprotease expression among different clinical isolates as well as an evaluation of the effect of long-term in vitro passage on the expression pattern of these metalloproteases were also performed. Western blot analysis, using an anti-gp63 antibody, revealed polypeptide patterns with a similar profile to that observed in zymographic analysis. Flow cytometry and fluorescence microscopy analyses corroborated the presence of metalloproteases with homologous domains to gp63 in the parasites and revealed differences in the expression level of such molecules among the isolates. The cellular distribution of metalloproteases, assessed by confocal analysis, showed the existence of intracellular metalloproteases with homologous domains to gp63, predominantly located near the flagellar pocket. Finally, it was observed that differential zymographic profiles of metalloproteases exhibited by L. (V.) braziliensis isolates remain unaltered during prolonged in vitro culture, suggesting that the proteolytic activity pattern is a stable phenotypic characteristic of these parasites.
authors.affiliation: Laboratório de Pesquisa em Leishmanioses, Instituto Oswaldo Cruz, FIOCRUZ, Rio de Janeiro, Brazil.
authors.forename: Patricia
authors.initials: P
authors.lastname: Cuervo
authors.forename: André L S
authors.initials: AL
authors.lastname: Santos
authors.forename: Carlos R
authors.initials: CR
authors.lastname: Alves
authors.forename: Gustavo C
authors.initials: GC
authors.lastname: Menezes
authors.forename: Bianca A
authors.initials: BA
authors.lastname: Silva
authors.forename: Constança
authors.initials: C
authors.lastname: Britto
authors.forename: Octavio
authors.initials: O
authors.lastname: Fernandes
authors.forename: Elisa
authors.initials: E
authors.lastname: Cupolillo
authors.forename: Jose
authors.initials: J
authors.lastname: Batista De Jesus
mesh_terms: D000818:Animals; D015153:Blotting, Western; D001938:Brazil; D003105:Colombia; D005407:Flagella / Q000737:chemistry; D005434:Flow Cytometry; D006801:Humans; D007892:Leishmania braziliensis / Q000737:chemistry* / Q000302:isolation & purification; D007896:Leishmaniasis / Q000469:parasitology; D008666:Metalloendopeptidases / Q000032:analysis* / Q000276:immunology; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D020543:Proteome / Q000032:analysis; D015800:Protozoan Proteins / Q000032:analysis* / Q000276:immunology; D012692:Serial Passage
chemical_list: D020543:Proteome; D015800:Protozoan Proteins; D008666:Metalloendopeptidases; C055146:glycoprotein gp63, Leishmania