Abstract
Apicomplexan parasites rely on tubulin structures throughout their cell and life cycles, particularly in the polymerization of spindle microtubules to separate the replicated nucleus into daughter cells. Additionally, tubulin structures, including conoid and subpellicular microtubules, provide the necessary rigidity and structure for dissemination and host cell invasion. However, it is unclear whether these tubulin structures are nucleated via a highly conserved γ-tubulin complex or through a specific process unique to apicomplexans. This study demonstrates that Toxoplasma γ-tubulin is responsible for nucleating spindle microtubules, akin to higher eukaryotes, facilitating nucleus division in newly formed parasites. Interestingly, γ-tubulin colocalizes with nascent conoid and subpellicular microtubules during division, potentially nucleating these structures as well. Loss of γ-tubulin results in significant morphological defects due to impaired nucleus scission and the loss of conoid and subpellicular microtubule nucleation, crucial for parasite shape and rigidity. Additionally, the nucleation process of tubulin structures involves a concerted action of γ-tubulin and Gamma Tubulin Complex proteins (GCPs), recapitulating the localization and phenotype of γ-tubulin. This study also introduces new molecular markers for cytoskeletal structures and applies iterative expansion microscopy to reveal microtubule-based architecture in Cryptosporidium parvum sporozoites, further demonstrating the conserved localization and probable function of γ-tubulin in Cryptosporidium.