Analysis of the roles of cysteine proteinases of Leishmania mexicana in the host-parasite interaction

Promastigotes of Leishmania mexicana mutants lacking the multicopy CPB cysteine proteinase genes (Delta CPB) are markedly less able than wild-type parasites to infect macrophages in vitro. Delta CPB promastigotes invade macrophages in large numbers but are unable to survive in the majority of the cells. In contrast, Delta CPB amastigotes invade and survive within macrophages in vitro. This extreme in vitro stage-specific difference was not mimicked in vivo; both promastigotes and amastigotes of Delta CPB produced lesions in BALB/c mice, but in each case the lesions grew considerably more slowly than those caused by wild-type parasites and only small lesions resulted. Inhibition of CPB in situ using cell-permeant peptidyl-diazomethylketones had no measurable effect on parasite growth or differentiation axenically in vitro. In contrast, N-benzoyloxycarbonyl-phe-ala-diazomethylketone reduced the infectivity of wild-type parasites to macrophages by 80%. Time-course experiments demonstrated that application of the inhibitor caused effects not seen with Delta CPB, suggesting that CPB may not be the prime target of this inhibitor. The data show that the CPB genes of L. mexicana encode enzymes that have important roles in intracellular survival of the parasite and more generally in its interaction with its mammalian host.