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Abstract:
Pleurotus ostreatus, the oyster mushroom, utilizes lollipop-shaped structures, toxocysts, to rapidly paralyze and kill nematodes under starvation. We have conducted forward genetic screens in P. ostreatus and isolated 22 loss-of-toxicity (lot) mutants that were incapable of paralyzing C. elegans. Whole genome sequencing and genetic mapping were employed to pinpoint the causative mutations in lot1, lot2, and lot3 mutants to be pho85-cyclin 1 (PCL1), HIS1, and a transcription factor for toxocyst development 1 (TTD1), respectively. First, we explored the connection between Pho85, a cyclin-dependent protein kinase, and Pcl1. Through yeast two-hybrid assays, we demonstrated a physical interaction between Pho85 and Pcl1, suggesting that the Pho85-Pcl1 complex may influence the cytoskeleton and septin organization, leading to toxocyst formation. Subsequently, we developed a congenic strain, PC9.15, derived from the original mutagenized PC9 strain, and conducted bulked segregant analysis to map the remaining 19 lot mutants. Our analysis revealed that the lot5, lot9, lot12, lot15, and lot21 mutants have mutations in tetrahydrofolate synthase (MIS1), HIS4, and HIS5. Furthermore, histidine supplementation in the growth medium successfully restored the deficiency in toxocyst development observed in these lot mutants, indicating the histidine biosynthetic pathway is required for toxocyst development.
