Plasmodium falciparum contains functional SCF and CRL4 ubiquitin E3 ligases, and CRL4 is critical for cell division and membrane integrity

Apicomplexan parasites, including Plasmodium falciparum and Toxoplasma gondii, contain specialized secretory organelles like micronemes, rhoptries, and dense granules, whose secretions are essential for various parasite activities. As lipid mobilization is key for organelle biogenesis and function, we investigated if the DedA superfamily lipid scramblase vacuole membrane protein 1 (VMP1) has a role in the biogenesis and function of apicomplexan secretory organelles. Our study demonstrates that VMP1 of P. falciparum (PfVMP1) and T. gondii (TgVMP1) localize to ER. Depletion of TgVMP1 caused reduced microneme secretion, loss of rhoptries and their decreased secretion, and accumulation of dense granules in ER, indicating a key role of TgVMP1 in the biogenesis and function of these organelles. Consistently, TgVMP1 depletion adversely affected parasite development, motility, host cell invasion, and egress. TgVMP depletion decreased the number of lipid droplets, which is similar to the phenotype of ER lipid scramblase-deficient cells and suggests that TgVMP1 is a lipid scramblase. Importantly, both human VMP1 and PfVMP1 localized to ER of TgVMP1-depleted parasites and rescued the depleted parasites, indicating a crucial role of an ER-resident lipid scramblase in the biogenesis and function of secretory organelles. The essentiality of TgVMP1 for parasite development and likely functional conservation of apicomplexan VMP1 proteins highlight their drug-target potential for developing pan-apicomplexan drugs.

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

Rashpa,

Smith,

Artavanis-Tsakonas

et al. 2024

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Malaria is a mosquito-borne infectious disease caused by unicellular eukaryotic parasites of the Plasmodium genus. Protein ubiquitination by E3 ligases is a critical post-translational modification required for various cellular processes during the lifecycle of Plasmodium parasites. However, little is known about the repertoire and function of these enzymes in Plasmodium . Here, we show that Plasmodium expresses a conserved cullin RING E3 ligase (CRL) complex that is functionally related to CRL4 in other eukaryotes. In P. falciparum 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 W D-repeat protein i mportant for g ametogenesis 1 (WIG1). This CRL4-related complex is also expressed in P. berghei gametocytes, with WIG1 being the only putative receptor detected in both the schizont and gametocyte stages. WIG1 disruption leads to a complete block in microgamete formation. Proteomic analyses indicate that WIG1 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 Plasmodium that is critical for the formation of microgametes by regulating proteostasis of ciliary and DNA replication proteins. IMPORTANCE Plasmodium 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 Plasmodium . 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.

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Plasmodium falciparum contains functional SCF and CRL4 ubiquitin E3 ligases, and CRL4 is critical for cell division and membrane integrity

Cited by 3 publications

References 92 publications

The Role of Neddylation in Malaria Parasites

The Role of Neddylation in Malaria Parasites

The apicomplexan DedA superfamily protein VMP1 is crucial for the biogenesis and function of secretory organelles

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

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Plasmodium falciparum contains functional SCF and CRL4 ubiquitin E3 ligases, and CRL4 is critical for cell division and membrane integrity

Cited by 3 publications

References 92 publications

The Role of Neddylation in Malaria Parasites

The Role of Neddylation in Malaria Parasites

The apicomplexan DedA superfamily protein VMP1 is crucial for the biogenesis and function of secretory organelles

A multistage Plasmodium CRL4 WIG1 ubiquitin ligase is critical for the formation of functional microtubule organization centers in microgametocytes

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