S-adenosylmethionine synthetase (SAMS) is a key enzyme for the synthesis of the lone methyl donor S-adenosyl methionine (SAM), which is involved in transmethylation reactions and hence required for cellular processes such as DNA, RNA, and histone methylation, but also polyamine biosynthesis and proteostasis. In the human malaria parasite Plasmodium falciparum, PfSAMS is encoded by a single gene and has been suggested to be crucial for malaria pathogenesis and transmission; however, to date, PfSAMS has not been fully characterized. To gain deeper insight into the function of PfSAMS, we generated a conditional gene knockdown (KD) using the glmS ribozyme system. We show that PfSAMS localizes to the cytoplasm and the nucleus of blood-stage parasites. PfSAMS-KD results in reduced histone methylation and leads to impaired intraerythrocytic growth and gametocyte development. To further determine the interaction network of PfSAMS, we performed a proximity-dependent biotin identification analysis. We identified a complex network of 1114 proteins involved in biological processes such as cell cycle control and DNA replication, or transcription, but also in phosphatidylcholine and polyamine biosynthesis and proteasome regulation. Our findings highlight the diverse roles of PfSAMS during intraerythrocytic growth and sexual stage development and emphasize that PfSAMS is a potential drug target.

Perforins are secreted proteins of eukaryotes, which possess a membrane attack complex/perforin (MACPF) domain enabling them to form pores in the membranes of target cells. In higher eukaryotes, they are assigned to immune defense mechanisms required to kill invading microbes or infected cells. Perforin-like proteins (PLPs) are also found in apicomplexan parasites. Here they play diverse roles during lifecycle progression of the intracellularly replicating protozoans. The apicomplexan PLPs are best studied in Plasmodium and Toxoplasma, the causative agents of malaria and toxoplasmosis, respectively. The PLPs are expressed in the different lifecycle stages of the pathogens and can target and lyse a variety of cell membranes of the invertebrate and mammalian hosts. The PLPs thereby either function in host cell destruction during exit or in overcoming epithelial barriers during tissue passage. In this review, we summarize the various PLPs known for apicomplexan parasites and highlight their roles in Plasmodium and Toxoplasma lifecycle progression.

Transmission of malaria parasites to the mosquito is mediated by sexual precursor cells, the gametocytes. Upon entering the mosquito midgut, the gametocytes egress from the enveloping erythrocyte while passing through gametogenesis. Egress follows an inside-out mode during which the membrane of the parasitophorous vacuole ruptures prior to the erythrocyte membrane. Membrane rupture requires the exocytosis of specialized secretory vesicles of the parasites; i.e. the osmiophilic bodies (OBs) involved in rupturing the parasitophorous vacuole membrane, and vesicles (here termed g-exonemes) that harbour the perforin-like protein PPLP2 required for erythrocyte lysis. While several OB proteins are known, like G377 and MDV1/Peg3, the protein composition of the g-exonemes remains unclear. Here, we used highresolution imaging and BioID methods to study the two types of egress vesicles in Plasmodium falciparum gametocytes. We show that OB exocytosis precedes discharge of the g-exonemes and that exocytosis of the g-exonemes, but not of the OBs, is calcium-sensitive. Further, the two types of vesicles exhibit distinct proteomes. In addition to known egress-related proteins, our analyses revealed novel components of OBs and g-exonemes, including proteins involved in vesicle trafficking. Our data provide insight into the immense molecular machinery required for the inside-out egress of P. falciparum gametocytes.

Transmission of malaria parasites to the mosquito is mediated by sexual precursor cells, the gametocytes. Upon entering the mosquito midgut, the gametocytes egress from the enveloping erythrocyte while passing through gametogenesis. Egress follows an inside‐out mode during which the membrane of the parasitophorous vacuole (PV) ruptures prior to the erythrocyte membrane. Membrane rupture requires exocytosis of specialized egress vesicles of the parasites; that is, osmiophilic bodies (OBs) involved in rupturing the PV membrane, and vesicles that harbor the perforin‐like protein PPLP2 (here termed P‐EVs) required for erythrocyte lysis. While some OB proteins have been identified, like G377 and MDV1/Peg3, the majority of egress vesicle‐resident proteins is yet unknown. Here, we used high‐resolution imaging and BioID methods to study the two egress vesicle types in Plasmodium falciparum gametocytes. We show that OB exocytosis precedes discharge of the P‐EVs and that exocytosis of the P‐EVs, but not of the OBs, is calcium sensitive. Both vesicle types exhibit distinct proteomes with the majority of proteins located in the OBs. In addition to known egress‐related proteins, we identified novel components of OBs and P‐EVs, including vesicle‐trafficking proteins. Our data provide insight into the immense molecular machinery required for the inside‐out egress of P. falciparum gametocytes.