The molecular mechanisms regulating the sexual development of malaria parasites from gametocytes to oocysts in their mosquito vector are still largely unexplored. In other eukaryotes, NIMA-related kinases (Neks) regulate cell cycle progression and have been implicated in the regulation of meiosis. Here, we demonstrate that Nek-4, a new Plasmodium member of the Nek family, is essential for completion of the sexual cycle of the parasite. Recombinant Plasmodium falciparum Nek-4 possesses protein kinase activity and displays substrate preferences similar to those of other Neks. Nek-4 is highly expressed in gametocytes, yet disruption of the nek-4 gene in the rodent malaria parasite P. berghei has no effect on gamete formation and subsequent fertilization. However, further differentiation of zygotes into ookinetes is abolished. Measurements of nuclear DNA content indicate that zygotes lacking Nek-4 fail to undergo the genome replication to the tetraploid level that precedes meiosis. Cell cycle progression in the zygote is identified as a likely precondition for its morphological transition to the ookinete and for the successful establishment of a malaria infection in the mosquito.Malaria remains a devastating disease in most tropical and subtropical regions. The problem has been exacerbated over the last decades of the twentieth century by the emergence and spread of resistance of the causative agents, parasitic protists of the genus Plasmodium, to available antimalarials (Plasmodium falciparum is the species responsible for the vast majority of lethal cases) (1). Infection of the human host is initiated by the bite of an infected Anopheles mosquito, which delivers sporozoites into the bloodstream. The sporozoites rapidly gain the liver, where they invade hepatocytes and undergo a first round of schizogony. The merozoites produced in this process are released into the bloodstream and invade erythrocytes, where they undergo recurrent and synchronized schizogony. This is the phase of the parasite's life cycle that is responsible for malaria pathogenesis. A proportion of merozoites, upon invasion of a new red blood cell, do not enter schizogony, but arrest their cell cycle and develop into male or female gametocytes, the only forms capable of infecting the mosquito vector. Ingestion of gametocytes during a blood meal triggers their further development into gametes, a process, which for the male gametocyte, involves three rounds of genome replication and generation of eight flagellated male gametes. Fertilization is followed by nuclear fusion, one round of genome replication, and meiosis, which occurs within 3 h (2, 3). The nuclear envelope remains intact throughout this process, and meiosis is not followed by nuclear division. As a consequence the ookinete, a motile form that develops from the zygote and exits the mosquito midgut lumen, is tetraploid. The ookinete establishes an oocyst at the basal lamina, which produces several thousand sporozoites. These accumulate in the insect salivary glands and render the mosquito...
