Proteolytic activation of leishporin: evidence that Leishmania amazonensis and Leishmania guyanensis have distinct inactive forms.
Crude extracts of Leishmania amazonensis, but not of L. guyanensis, are lytic to erythrocytes and nucleated cells, including macrophages. L. amazonensis-mediated lysis is caused by a membrane-associated pore-forming protein, named a-leishporin. Here we show that L. amazonensis, but not L. guyanensis, promastigote extracts increase their hemolytic activity when kept at 4 degrees C for a few days or at 37 degrees C for a few hours. We show that the activation in the extracts is mediated by a cytosolic serine-protease. Although L. guyanensis extracts are hemolytically inactive and unable to generate hemolytic activity, their membrane fraction becomes hemolytic in the presence of the cytosolic fraction of L. amazonensis, also by the action of a serine-protease. This suggests that L. guyanensis contains a potential lytic molecule, named here g-leishporin. The cytosolic fraction of L. guyanensis is unable to activate either a- or g-leishporin, indicating that this species does not possess the protease(s) that activate(s) the cytolysin. Trypsin, chymotrypsin, collagenase, Pronase and proteinase K, are also effective in activating a-leishporin but not g-leishporin. This suggests that the inactive forms of a-leishporin and g-leishporin are distinct in structure and/or are activated by different mechanisms. We are considering two hypotheses for the activation of leishporins: (1) proteolysis of an inactive precursor and (2) dissociation and/or proteolytic degradation of an inhibitory oligopeptide. The present data and preliminary results argue for the second hypothesis. We speculate that leishporin could be activated in the protease-rich, low pH, and dissociating environment of parasitophorous vacuole contributing for the release of the parasites from the macrophage.
Almeida-Campos
F R
FR
Departamento de Bioquímica-Imunologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais, Brazil.
Horta
M F
MF
D000818:Animals; D003600:Cytosol / Q000378:metabolism; D003603:Cytotoxins / Q000378:metabolism*; D004789:Enzyme Activation; D004912:Erythrocytes / Q000502:physiology*; D006461:Hemolysis; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007891:Leishmania / Q000378:metabolism*; D018334:Leishmania guyanensis / Q000378:metabolism; D015800:Protozoan Proteins / Q000378:metabolism*; D012697:Serine Endopeptidases / Q000378:metabolism*; D013696:Temperature
D003603:Cytotoxins; D015800:Protozoan Proteins; C412371:leishporin protein, Leishmania; D012697:Serine Endopeptidases