A transcriptional switch underlies commitment to sexual development in malaria parasites

The life cycles of many parasites involve transitions between disparate host species, requiring these parasites to go through multiple developmental stages adapted to each of these specialized niches. Transmission of malaria parasites (Plasmodium spp.) from humans to the mosquito vector requires differentiation from asexual stages replicating within red blood cells into non-dividing male and female gametocytes. Although gametocytes were first described in 1880, our understanding of the molecular mechanisms involved in commitment to gametocyte formation is extremely limited, and disrupting this critical developmental transition remains a long-standing goal. Here we show that expression levels of the DNA-binding protein PfAP2-G correlate strongly with levels of gametocyte formation. Using independent forward and reverse genetics approaches, we demonstrate that PfAP2-G function is essential for parasite sexual differentiation. By combining genome-wide PfAP2-G cognate motif occurrence with global transcriptional changes resulting from PfAP2G ablation, we identify early gametocyte genes as probable targets of PfAP2-G and show that their regulation by PfAP2-G is critical for their wild-type level expression. In the asexual blood-stage parasites pfap2-g appears to be among a set of epigenetically silenced loci prone to spontaneous activation. Stochastic activation presents a simple mechanism for a low baseline of gametocyte production. Overall, these findings identify PfAP2-G as a master regulator of sexual-stage development in malaria parasites and mark the first discovery of a transcriptional switch controlling a differentiation decision in protozoan parasites. From its uptake in a mosquito blood meal to initial infection of red blood cells in the subsequent host, the malaria parasite Plasmodium falciparum goes through at least seven key developmental changes (asexual red cell stage R gametocyte R gamete R ookinete R oocyst R sporozoite R liver stage R asexual red cell stage). In all but one case, as the parasite reaches its subsequent niche within the host, differentiation into the appropriate developmental stage is a necessity for continuation of the life cycle. The lone exception occurs once the parasite has started replicating in red blood cells. During the 48-h intraerythrocytic developmental cycle following each new red blood cell invasion, a developmental decision is made that determines whether daughter parasites will continue replicating asexually and maintain the infection of the current host or differentiate into non-dividing male or female gametocytes. Although the latter decision is a dead-end for replication within the current host it is essential for infection of mosquitoes and thus transmission to the next host. A recent study on transcriptional variation identified differentially expressed genes linked to early gametocyte development in two stocks (3D7-A and 3D7-B) of the common 3D7 P. falciparum parasite line. Within this expression cluster of early gametocyte markers, we noted the presence of a potential transcriptional regulator, PfAP2-G (PFL1085w/ PF3D7_1222600; http://www.plasmodb.org), which belongs to the apicomplexan AP2 (ApiAP2) family of DNA-binding proteins (Supplementary Fig. 1) and is conserved among most members of the phylum (Supplementary Fig. 2). ApiAP2 proteins represent the main family of transcriptional regulators in malaria parasites and have thus far been found to regulate several of the parasite’s developmental transitions, including ookinete formation and oocyst sporozoite maturation within the mosquito, and development in the mammalian liver. Follow-up quantitative PCR with reverse transcription (qRT–PCR) analysis in blood-stage parasites confirmed higher pfap2-g transcript abundance in 3D7-B compared to 3D7-A and also revealed significant variation in expression levels between individual 3D7-B subclones (Fig. 1a). Notably, when gametocyte formation was measured in these lines, pfap2-g transcript levels were highly predictive (R . 0.99) of relative gametocyte production (Fig. 1b).