Apical Surface Expression of Aspartic Protease Plasmepsin 4, a Potential Transmission-blocking Target of the Plasmodium Ookinete

Fengwu Li,Kailash P Patra,Charles A Yowell,John B Dame,Karen Chin,Joseph M Vinetz

doi:10.1074/jbc.m109.063388

Fengwu Li, Kailash P Patra + Show 4 more

Open Access

https://doi.org/10.1074/jbc.m109.063388

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Abstract

To invade its definitive host, the mosquito, the malaria parasite must cross the midgut peritrophic matrix that is composed of chitin cross-linked by chitin-binding proteins and then develop into an oocyst on the midgut basal lamina. Previous evidence indicates that Plasmodium ookinete-secreted chitinase is important in midgut invasion. The mechanistic role of other ookinete-secreted enzymes in midgut invasion has not been previously examined. De novo mass spectrometry sequencing of a protein obtained by benzamidine affinity column of Plasmodium gallinaceum ookinete axenic culture supernatant demonstrated the presence of an ookinete-secreted plasmepsin, an aspartic protease previously only known to be present in the digestive vacuole of asexual stage malaria parasites. This plasmepsin, the ortholog of Plasmodium falciparum plasmepsin 4, was designated PgPM4. PgPM4 and PgCHT2 (the P. gallinaceum ortholog of P. falciparum chitinase PfCHT1) are both localized on the ookinete apical surface, and both are present in micronemes. Aspartic protease inhibitors (peptidomimetic and natural product), calpain inhibitors, and anti-PgPM4 monoclonal antibodies significantly reduced parasite infectivity for mosquitoes. These results suggest that plasmepsin 4, previously known only to function in the digestive vacuole of asexual blood stage Plasmodium, plays a role in how the ookinete interacts with the mosquito midgut interactions as it becomes an oocyst. These data are the first to delineate a role for an aspartic protease in mediating Plasmodium invasion of the mosquito and demonstrate the potential for plasmepsin 4 as a malaria transmission-blocking vaccine target.

Highlights

Through the bite of infected Anopheles mosquitoes
We demonstrate that the P. gallinaceum plasmepsin 4 (PgPM4)2 synergizes with the P. gallinaceum chitinase PgCHT2 (the ortholog of the P. falciparum chitinase [18]) to facilitate malaria parasite invasion of the mosquito midgut and/or may be involved in the development of ookinete to oocyst
By searching the recently completed 3-fold coverage of P. gallinaceum genome, the four peptides were found to be encoded by a single gene, designated PgPM4 based on the highest degree of similarity to plasmepsin 4 of P. falciparum (PfPM4), P. vivax (PvPM4), Plasmodium ovale (PoPM4), and Plasmodium malariae (PmPM4)

Summary

Introduction

Through the bite of infected Anopheles mosquitoes. Preventing transmission from the human reservoir to the definitive host, the mosquito, is one approach to malaria control [2]. We demonstrate that the P. gallinaceum plasmepsin 4 (PgPM4) synergizes with the P. gallinaceum chitinase PgCHT2 (the ortholog of the P. falciparum chitinase [18]) to facilitate malaria parasite invasion of the mosquito midgut and/or may be involved in the development of ookinete to oocyst These data are the first to indicate a specific mechanistic function for a plasmepsin in any stage of the malaria parasite other than the asexual blood stage. This aspartic protease plays an important role in ookinete invasion of the mosquito midgut and/or parasite development, and may be a novel target of blocking malaria transmission

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