A multistagePlasmodiumCRL4WIG1ubiquitin ligase is critical for the formation of functional microtubule organisation centres in microgametocytes

Abstract

AbstractMalaria is a mosquito-borne infectious disease caused by unicellular eukaryotic parasites of thePlasmodiumgenus. Protein ubiquitination by E3 ligases is a critical post-translational modification required for various cellular processes during the lifecycle ofPlasmodiumparasites. However, little is known about the repertoire and function of these enzymes inPlasmodium.Here we show thatPlasmodiumexpresses a conserved cullin RING E3 ligase (CRL) complex that is functionally related to the eukaryotic CRL4. InP. falciparumasexual blood stages, a cullin-4 scaffold interacts with the RING protein RBX1, the adaptor protein DDB1 and a set of putative receptor proteins that may determine substrate specificity for ubiquitination. These receptor proteins contain WD40-repeat domains and includeWD-repeat protein Important forGametogenesis 1 (WIG1). This CRL4-related complex is also expressed inP. bergheigametocytes, with WIG1 being the only putative receptor detected in both schizont and gametocyte stages. While WIG1 is not required for the proliferation ofP. bergheiasexual blood stages, its disruption leads to a complete block in microgamete formation. Proteomic analyses indicate thatWIG1disruption alters proteostasis of ciliary proteins and components of the DNA replication machinery during gametocytogenesis. Further analysis by ultrastructure expansion microscopy (U-ExM) indicates that WIG1-dependent depletion of ciliary proteins is associated with impaired formation of the microtubule organisation centres that coordinate mitosis with axoneme formation and altered DNA replication during microgametogenesis. This work identifies a CRL4-related ubiquitin ligase inPlasmodiumthat is critical for the transmission of malaria parasites by regulating proteostasis of ciliary and DNA replication proteins.