Protein Export Marks the Early Phase of Gametocytogenesis of the Human Malaria Parasite Plasmodium falciparum

Despite over a century of study of malaria parasites, parts of the Plasmodium falciparum life cycle remain virtually unknown. One of these is the early gametocyte stage, a round shaped cell morphologically similar to an asexual trophozoite in which major cellular transformations ensure subsequent development of the elongated gametocyte. We developed a protocol to obtain for the first time highly purified preparations of early gametocytes using a transgenic line expressing a green fluorescent protein from the onset of gametocytogenesis. We determined the cellular proteome (1427 proteins) of this parasite stage by high accuracy tandem mass spectrometry and newly determined the proteomes of asexual trophozoites and mature gametocytes, identifying altogether 1090 previously undetected parasite proteins. Quantitative label-free comparative proteomics analysis determined enriched protein clusters for the three parasite developmental stages. Gene set enrichment analysis on the 251 proteins enriched in the early gametocyte proteome revealed that proteins putatively exported and involved in erythrocyte remodeling are the most overrepresented protein set in these stages. One-tenth of the early gametocyte-enriched proteome is constituted of putatively exported proteins, here named PfGEXPs (P. falciparum gametocyte-exported proteins). N-terminal processing and N-acetylation at a conserved leucine residue within the Plasmodium export element pentamotif were detected by mass spectrometry for three such proteins in the early but not in the mature gametocyte sample, further supporting a specific role in protein export in early gametocytogenesis. Previous reports and results of our experiments confirm that the three proteins are indeed exported in the erythrocyte cytoplasm. This work indicates that protein export profoundly marks early sexual differentiation in P. falciparum, probably contributing to host cell remodeling in this phase of the life cycle, and that gametocyte-enriched molecules are recruited to modulate this process in gametocytogenesis. Despite over a century of study of malaria parasites, parts of the Plasmodium falciparum life cycle remain virtually unknown. One of these is the early gametocyte stage, a round shaped cell morphologically similar to an asexual trophozoite in which major cellular transformations ensure subsequent development of the elongated gametocyte. We developed a protocol to obtain for the first time highly purified preparations of early gametocytes using a transgenic line expressing a green fluorescent protein from the onset of gametocytogenesis. We determined the cellular proteome (1427 proteins) of this parasite stage by high accuracy tandem mass spectrometry and newly determined the proteomes of asexual trophozoites and mature gametocytes, identifying altogether 1090 previously undetected parasite proteins. Quantitative label-free comparative proteomics analysis determined enriched protein clusters for the three parasite developmental stages. Gene set enrichment analysis on the 251 proteins enriched in the early gametocyte proteome revealed that proteins putatively exported and involved in erythrocyte remodeling are the most overrepresented protein set in these stages. One-tenth of the early gametocyte-enriched proteome is constituted of putatively exported proteins, here named PfGEXPs (P. falciparum gametocyte-exported proteins). N-terminal processing and N-acetylation at a conserved leucine residue within the Plasmodium export element pentamotif were detected by mass spectrometry for three such proteins in the early but not in the mature gametocyte sample, further supporting a specific role in protein export in early gametocytogenesis. Previous reports and results of our experiments confirm that the three proteins are indeed exported in the erythrocyte cytoplasm. This work indicates that protein export profoundly marks early sexual differentiation in P. falciparum, probably contributing to host cell remodeling in this phase of the life cycle, and that gametocyte-enriched molecules are recruited to modulate this process in gametocytogenesis. The burden imposed by malaria protozoan parasites to human populations worldwide is due to the severity of the disease, particularly when caused by the species Plasmodium falciparum, and to the efficient transmission of the parasite between humans and the Anopheles mosquito vectors. Gametocytes, the precursors of male and female gametes, are the Plasmodium developmental stages critical to the transmission of the parasites from the human blood stream to the mosquito gut when the insect bites an infected individual. In the human host and in in vitro cultures, P. falciparum gametocytogenesis proceeds in about 10 days through the five classically described morphological stages I to V (1.Hawking F. Wilson M.E. Gammage K. Evidence for cyclic development and short-lived maturity in the gametocytes of Plasmodium falciparum.Trans. R. Soc. Trop. Med. Hyg. 1971; 65: 549-559Abstract Full Text PDF PubMed Scopus (189) Google Scholar). The typical elongated form of gametocytes in this species appears around day 2 of maturation in the crescent-shaped stage II gametocyte, which progressively further elongates during maturation. In P. falciparum infections, mature stage V gametocytes are in the stages are in P. Plasmodium falciparum of a PubMed Scopus Google Scholar). malaria parasites were over a century on parts of the Plasmodium life cycle is One such stage is the early stage I gametocyte, which is morphologically similar to a asexual The of this stage for to the of proteins, and from and K. R. and of the Plasmodium falciparum sexual stage 65: PubMed Scopus Google R. K. P. Plasmodium an protein during early gametocyte PubMed Scopus Google Scholar). parasites and to gametocytes at the onset of gametocytogenesis F. P. of at the onset of gametocytogenesis in Plasmodium PubMed Scopus Google of a during the stage Scopus Google and proteins in the first of sexual F. 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