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.

[1]  Alane Beatriz Vermelho,et al.  A novel extracellular calcium-dependent cysteine proteinase from Crithidia deanei. , 2003, Archives of biochemistry and biophysics.

[2]  J. Donelson,et al.  The major surface protease (MSP or GP63) of Leishmania sp. Biosynthesis, regulation of expression, and function. , 2003, Molecular and biochemical parasitology.

[3]  Alane Beatriz Vermelho,et al.  A metalloproteinase extracellularly released by Crithidia deanei. , 2003, Canadian journal of microbiology.

[4]  D. Sánchez,et al.  gp63 Homologues in Trypanosoma cruzi: Surface Antigens with Metalloprotease Activity and a Possible Role in Host Cell Infection , 2003, Infection and Immunity.

[5]  C. Jaffe,et al.  Extracellular release of the surface metalloprotease, gp63, from Leishmania and insect trypanosomatids , 2003, Parasitology Research.

[6]  J. Donelson,et al.  Expression and Function of the Trypanosoma brucei Major Surface Protease (GP63) Genes* , 2003, Journal of Biological Chemistry.

[7]  N. Hooper,et al.  Surface Coat Remodeling during Differentiation of Trypanosoma brucei* , 2003, Journal of Biological Chemistry.

[8]  S. Goldenberg,et al.  Leishmania (Leishmania) amazonensis: differential expression of proteinases and cell-surface polypeptides in avirulent and virulent promastigotes. , 2003, Experimental parasitology.

[9]  Alane Beatriz Vermelho,et al.  Extracellular metalloproteinases in Phytomonas serpens. , 2003, Canadian journal of microbiology.

[10]  Alane Beatriz Vermelho,et al.  Extracellular metalloproteinase activity in Phytomonas françai , 2003, Parasitology Research.

[11]  Alane Beatriz Vermelho,et al.  Crithidia guilhermei: gelatin- and haemoglobin-degrading extracellular metalloproteinases. , 2002, Experimental parasitology.

[12]  C. Abreu,et al.  Activation of the Glycosylphosphatidylinositol-Anchored Membrane Proteinase upon Release from Herpetomonas samuelpessoai by Phospholipase C , 2002, Current Microbiology.

[13]  M. Sajid,et al.  Cysteine proteases of parasitic organisms. , 2002, Molecular and biochemical parasitology.

[14]  Alane Beatriz Vermelho,et al.  Differential expression of proteolytic enzymes in endosymbiont-harboring Crithidia species. , 2001, FEMS microbiology letters.

[15]  V. Turk,et al.  The major cysteine proteinase of Trypanosoma cruzi: a valid target for chemotherapy of Chagas disease. , 2001, Current pharmaceutical design.

[16]  C. Alviano,et al.  Cell-Associated and Extracellular Proteinases in Blastocrithidia culicis: Influence of Growth Conditions , 2001, Current Microbiology.

[17]  Alane Beatriz Vermelho,et al.  Crithidia guilhermei: purification and partial characterization of a 62-kDa extracellular metalloproteinase. , 2001, Experimental parasitology.

[18]  M. Ferguson,et al.  The structure, biosynthesis and functions of glycosylphosphatidylinositol anchors, and the contributions of trypanosome research. , 1999, Journal of cell science.

[19]  André Souza dos Santos,et al.  Characterization of Proteinases in Herpetomonas anglusteri and Herpetomonas roitmani , 1999, Current Microbiology.

[20]  W. de Souza,et al.  Endosymbiosis in protozoa of the Trypanosomatidae family. , 1999, FEMS microbiology letters.

[21]  M. Jarvinen,et al.  Simplified endoproteinase assays using gelatin or azogelatin. , 1998, Analytical biochemistry.

[22]  S. Goldenberg,et al.  Ubiquity of Cysteine‐ and Metalloproteinase Activities in a Wide Range of Trypanosomatids , 1996, The Journal of eukaryotic microbiology.

[23]  M. A. de Sousa.  Trypomastigotes in cultures of Blastocrithidia culicis (Novy, MacNeal & Torrey, 1907) (Kinetoplastida: Trypanosomatidae). , 1994, Memorias do Instituto Oswaldo Cruz.

[24]  T. Glaser,et al.  Characterization of a surface metalloprotease from Herpetomonas samuelpessoai and comparison with Leishmania major promastigote surface protease. , 1993, Molecular and biochemical parasitology.

[25]  R. Etges.  Identification of a surface metalloproteinase on 13 species of Leishmania isolated from humans, Crithidia fasciculata, and Herpetomonas samuelpessoai. , 1992, Acta tropica.

[26]  F. Ashall,et al.  Electrophoretic detection of Trypanosoma cruzi peptidases. , 1990, Molecular and biochemical parasitology.

[27]  C. Barbiéri,et al.  Cross-Reactivity between Trypanosoma Cruzi and Insect Trypanosomatids as a Basis for the Diagnosis of Chagas' Disease* , 1981 .

[28]  I. Roitman,et al.  Growth of an insect Trypanosomatid at 37 C in a defined medium. , 1972, The Journal of protozoology.

[29]  F. G. Wallace.  The trypanosomatid parasites of insects and arachnids. , 1966, Experimental parasitology.