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Internal and Surface-Localized Major Surface Proteases of Leishmania spp. and Their Differential Release from Promastigotes

Departments of Internal Medicine,¹ Biochemistry,³ Microbiology,⁵ Epidemiology,⁶ Program in Molecular Biology, University of Iowa, Iowa City, Iowa 52242,⁴ VA Medical Center, Iowa City, Iowa 52246²

Received 9 March 2007/ Accepted 20 July 2007

Major surface protease (MSP), also called GP63, is a virulence factor of Leishmania spp. protozoa. There are three pools of MSP, located either internally within the parasite, anchored to the surface membrane, or released into the extracellular environment. The regulation and biological functions of these MSP pools are unknown. We investigated here the trafficking and extrusion of surface versus internal MSPs. Virulent Leishmania chagasi undergo a growth-associated lengthening in the t_1/2 of surface-localized MSP, but this did not occur in the attenuated L5 strain. The release of surface-localized MSP was enhanced in a dose-dependent manner by MßCD, which chelates membrane cholesterol-ergosterol. Furthermore, incubation of promastigotes at 37°C with Matrigel matrix, a soluble basement membrane extract of Engelbreth-Holm-Swarm tumor cells, stimulated the release of internal MSP but not of surface-located MSP. Taken together, these data indicate that MSP subpopulations in distinct cellular locations are released from the parasite under different environmental conditions. We hypothesize that the internal MSP with its lengthy t_1/2 does not serve as a pool for promastigote surface MSP in the sand fly vector but that it instead functions as an MSP pool ready for quick release upon inoculation of metacyclic promastigotes into mammals. We present a model in which these different MSP pools are released under distinct life cycle-specific conditions.

Published ahead of print on 10 August 2007.