Proteolytic expression in Blastocrithidia culicis: influence of the endosymbiont and similarities with virulence factors of pathogenic trypanosomatids.
Blastocrithidia culicis is an insect trypanosomatid that presents bacterial endosymbionts. The cell-associated and secreted proteinases of the endosymbiont-bearing and aposymbiotic strains were compared through the incorporation of proteinaceous substrates into sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Few qualitative changes could be detected in the proteolytic zymograms in the 2 strains studied when gelatin, casein, haemoglobin or bovine serum albumin (BSA) were tested. However, the level of proteolytic activities was significantly higher in the aposymbiotic strain. Some of the B. culicis proteins reacted in Western blots with antibodies raised against gp63, a zinc-metalloproteinase, and cruzipain, a cysteinyl-proteinase, which are virulence factors of the human pathogenic trypanosomatids, Leishmania spp. and Trypanosoma cruzi, respectively. The anti-cross-reacting determinant (CRD) antibody recognized 2 polypeptides (50 and 58 kDa) in the spent culture media and in the supernatant from glycosylphosphatidylinositol-phospholipase C (GPI-PLC)-treated cells, suggesting that these proteins are GPI-anchored to the plasma membrane. In addition, the anti-gp63 reacted with the 50 kDa protein. The identification of protein homologues in trypanosomatids with distinct life-cycles may help to determine the importance of proteinases in trypanosomatids.
D'Avila-Levy
C M
CM
Departamento de Microbiologia Geral, Instituto de Microbiologia Prof. Paulo de Góes (IMPPG), Centro de Ciências da Saúde (CCS), Universidade Federal do Rio de Janeiro (UFRJ), Rio de Janeiro, RJ 21941-590, Brazil.
Araújo
F M
FM
Vermelho
A B
AB
Soares
R M A
RM
Santos
A L S
AL
Branquinha
M H
MH
D000818:Animals; D015153:Blotting, Western; D002364:Caseins / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005780:Gelatin / Q000378:metabolism; D054800:Glycosylphosphatidylinositol Diacylglycerol-Lyase; D006454:Hemoglobins / Q000378:metabolism; D008666:Metalloendopeptidases / Q000378:metabolism; D010447:Peptide Hydrolases / Q000378:metabolism*; D043265:Phosphatidylinositol Diacylglycerol-Lyase / Q000378:metabolism; D015800:Protozoan Proteins; D012710:Serum Albumin, Bovine / Q000378:metabolism; D013559:Symbiosis / Q000502:physiology; D014351:Trypanosomatina / Q000201:enzymology* / Q000502:physiology
D002364:Caseins; D006454:Hemoglobins; D015800:Protozoan Proteins; D012710:Serum Albumin, Bovine; D005780:Gelatin; D010447:Peptide Hydrolases; D003546:Cysteine Endopeptidases; C063214:cruzipain; D008666:Metalloendopeptidases; C055146:glycoprotein gp63, Leishmania; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054800:Glycosylphosphatidylinositol Diacylglycerol-Lyase