Plasmodium falciparum Calcium-Dependent Protein Kinase 4 is Critical for Male Gametogenesis and Transmission to the Mosquito Vector

Abstract

Transmission of the malaria parasite to the mosquito vector is critical for the completion of the sexual stage of the parasite life cycle and is dependent on the release of male gametes from the gametocyte body inside the mosquito midgut. In the present study, we demonstrate that PfCDPK4 is critical for male gametogenesis and is involved in phosphorylation of proteins essential for male gamete emergence.

Adapt or Die: Targeting Unique Transmission-Stage Biology for Malaria Elimination

“…The PKG mediated cascade (including involvement of Pf PI4K and phosphatidylinositol 4-phosphate 5-kinase, PIP5K) results ultimately in opening of IP 3 -gated calcium channels on the ER and a resultant increase in intracellular Ca 2+ ( Bennink et al., 2016 ). Activation of several sex-specific CDPKs ensues ( Brochet et al., 2021 ), including CDPK1-mediated release of DOZI, CITH and Pf Puf2 for translation in female gametes and CDPK4 to activate cyclins and cyclin-dependent kinases for cell division in male gametes ( Kumar et al., 2021 ) ( Figure 4 ). The latter is mediated by successive S and M cell cycle phases ( Raabe et al., 2009 ; Meibalan and Marti, 2017 ).…”

“…Cysteine and serine proteases (falcipain-1, plasmepsin IX, plasmalepsin X, SERA-6, SERA-7, and PfSUB3), MTRAP and PfDPAP2 are involved in PVM and erythrocyte membrane rupture, and aspartic proteases in gamete round-up ( Munsamy et al., 2018 ). Egress is followed by exflagellation, mediated by the male-specific axoneme assembly (flagellum formation) proteins (e.g., the armadillo repeat protein PF16, Kinesin 8B, SAS-6 ( Straschil et al., 2010 ; Depoix et al., 2020 ); MAP2 and CDC20 ( Kumar et al., 2021 ); cytoskeletal proteins α-tubulin II and actin II ( Elena et al., 2011 ) and Pf MR5 ( Eksi et al., 2006 ). Mutual recognition and male and female gamete attachment are mediated by P230, Pfs48/45, Pfs47 ( Van Dijk et al., 2010 ), complement control CCp proteins ( Baker, 2010 ) and Pb115 ( Liu et al., 2019 ) ( Figure 4 ).…”

A Putative Plasmodium RNA-Binding Protein Plays a Critical Role in Female Gamete Fertility and Parasite Transmission to the Mosquito Vector

“…To ascertain the gender-specific function of Pf MaCFET, we assessed the fertility of microgametes and macrogametes of Pfmacfet¯ using an additional genetic cross. The cross was arranged between Pfmacfet¯ parasites and a gene deletion parasite line of the CDPK4 locus, which we have shown generates only fertile macrogametes ( Pfcdpk4¯ ) ( Kumar et al, 2021 ). WT Pf NF54, Pfcdpk4¯ and Pfmacfet¯ gametocytes were generated in vitro and first fed individually to female A. stephensi mosquitoes on day 15 of culture.…”

“…This demonstrated that one gender of Pfmacfet¯ gametes must be fertile. An additional genetic crosse, utilizing a male sterile line ( Pfcdpk4¯ ) ( Kumar et al, 2021 ) and Pfmacfet¯ , revealed that Pfmacfet¯ male gametes are fertile but female gametes are sterile.…”

“…When fully mature transmissible forms of stage V gametocytes are taken up by the mosquito vector in an infectious blood meal, they become activated to form gametes by a combination of factors including a drop in temperature ( Sinden and Croll, 1975 ), an increase in pH ( Sinden, 1983 ), and/or exposure to xanthurenic acid (XA), a metabolite of tryptophan ( Billker et al, 1998 ). Gametogenesis is regulated by mobilization of intracellular calcium (Ca 2+ ) stores, which control Ca 2+ dependent protein function ( Billker et al, 2004 ; Kumar et al, 2021 ) and is a rapid process with the male (micro) gametocyte undergoing three rounds of DNA replication and assembly of axonemes to form eight flagellar microgametes. The female (macro) gametocyte only undergoes a marked reduction in cytoplasmic density and nuclear changes to form a single macrogamete ( Andreadaki et al, 2018 ).…”