The Leishmania chagasi proteasome: role in promastigotes growth and amastigotes survival within murine macrophages.
Proteasomes are multisubunit proteases that exist universally among eukaryotes. They have multiple proteolytic activities and are believed to have important roles in regulating cell cycle, selective intracellular proteolysis, and antigen presentation. Here we have partially purified Leishmania chagasi proteasome. The L. chagasi proteasome rich fraction displayed the typical features of eukaryotic 20S proteasome complexes, being active towards peptidyl substrates with hydrophobic and acidic residues, and sensitive to the proteasome-specific inhibitor lactacystin. We have shown that lactacystin, or its active form clasto-lactacystin beta-lactone, but not E-64, blocks the in vitro growth of L. chagasi promastigotes, demonstrating that the interference with parasite growth is due to the lack of proteasome activity. Furthermore, pre-treatment of L. chagasi promastigotes with lactacystin did not prevent parasite entry in host cells, but markedly restricted its intracellular survival. These results demonstrate that intact parasite proteasome function is required for replication of L. chagasi and for amastigotes survival inside the vertebrate host cell.
Silva-Jardim
Izaltina
I
Departamento de Bioquímica e Imunologia, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, CEP 14049-900, Brazil.
Horta
M F
MF
Ramalho-Pinto
F Juarez
FJ
D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D003374:Coumarins / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000302:isolation & purification / Q000378:metabolism*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004195:Disease Models, Animal; D007891:Leishmania / Q000201:enzymology* / Q000254:growth & development / Q000378:metabolism; D007898:Leishmaniasis, Visceral / Q000469:parasitology*; D007930:Leucine / Q000031:analogs & derivatives* / Q000494:pharmacology; D017737:Macrophages, Peritoneal / Q000469:parasitology*; D008297:Male; D051379:Mice; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D009842:Oligopeptides / Q000378:metabolism; D010664:Phenylmethylsulfonyl Fluoride / Q000494:pharmacology; D046988:Proteasome Endopeptidase Complex
D003374:Coumarins; D015853:Cysteine Proteinase Inhibitors; D009097:Multienzyme Complexes; D009842:Oligopeptides; C488643:lactacystin beta-lactone; C067713:lactacystin; D010664:Phenylmethylsulfonyl Fluoride; C046582:succinyl-leucyl-leucyl-valyl-tyrosyl-methylcoumarinamide; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D007930:Leucine; C024974:E 64; D000111:Acetylcysteine