A multistage <i>Plasmodium</i> CRL4 ^WIG1 ubiquitin ligase is critical for the formation of functional microtubule organization centers in microgametocytes

Malaria is a mosquito-borne infectious disease caused by unicellular eukaryotic parasites of the <i>Plasmodium</i> genus. Protein ubiquitination by E3 ligases is a critical post-translational modification required for various cellular processes during the lifecycle of <i>Plasmodium</i> parasites. However, little is known about the repertoire and function of these enzymes in <i>Plasmodium</i>. Here, we show that <i>Plasmodium</i> expresses a conserved cullin RING E3 ligase (CRL) complex that is functionally related to CRL4 in other eukaryotes. In <i>P. falciparum</i> asexual 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 include <u>W</u>D-repeat protein <u>i</u>mportant for <u>g</u>ametogenesis 1 (WIG1). This CRL4-related complex is also expressed in <i>P. berghei</i> gametocytes, with WIG1 being the only putative receptor detected in both the schizont and gametocyte stages. <i>WIG1</i> disruption leads to a complete block in microgamete formation. Proteomic analyses indicate that <i>WIG1</i> disruption 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 the formation of the microtubule organization centers that coordinate mitosis with axoneme formation and altered DNA replication during microgametogenesis. This work identifies a CRL4-related ubiquitin ligase in <i>Plasmodium</i> that is critical for the formation of microgametes by regulating proteostasis of ciliary and DNA replication proteins.IMPORTANCE<i>Plasmodium</i> parasites undergo fascinating lifecycles with multiple developmental steps, converting into morphologically distinct forms in both their mammalian and mosquito hosts. Protein ubiquitination by ubiquitin ligases emerges as an important post-translational modification required to control multiple developmental stages in <i>Plasmodium</i>. Here, we identify a cullin RING E3 ubiquitin ligase (CRL) complex expressed in the replicating asexual blood stages and in the gametocyte stages that mediate transmission to the mosquito. WIG1, a putative substrate recognition protein of this ligase complex, is essential for the maturation of microgametocytes into microgametes upon ingestion by a mosquito. More specifically, WIG1 is required for proteostasis of ciliary proteins and components of the DNA replication machinery during gametocytogenesis. This requirement is linked to DNA replication and microtubule organization center formation, both critical to the development of flagellated microgametes.