To identify HDAC3 occupancy in the genomes of ME49 and ΔXII-5 strains. To identify HDAC3 occupancy in the genomes of ME49 and ΔXII-5 strains

Toxoplasma gondii is a ubiquitous protozoan parasite with a complex life cycle containing multiple developmental stages. The parasites have distinct gene expression patterns at different stages to enable stage specific life activities, but the underlying regulatory mechanisms are largely unknown. In this study, we identified a nuclear complex that controls the expression of developmentally regulated genes, especially merozoite specific genes. This complex consists of the AP2 family transcription factor AP2Ⅻ-5, the epigenetic factors MORC and HDAC3, as well as a novel AP2Ⅻ-5 interacting protein 1 (AIP1) that stabilizes this complex. At the tachyzoite stage when the parasites proliferate rapidly by asexual endodyogeny, AP2Ⅻ-5 binds to the promoter regions of developmentally activated genes and recruits MORC and HDAC3 to suppress their expression. When sexual commitment and merozoite development is triggered, the abundance of AP2Ⅻ-5 decreases and its suppression on target genes is relieved. In contrast to MORC and HDAC3, which regulate Toxoplasma development but are also essential for tachyzoite growth, AP2Ⅻ-5 and AIP1 are dispensable for tachyzoite proliferation in vitro. These data suggest that while MORC and HDAC3 have broad regulatory activities, forming a complex with AP2Ⅻ-5 and AIP1 enables them to specifically regulate gene expression during development. Overall design: To determine the HDAC3 binding sites in the genomes of ME49 and ΔⅫ-5 strains using CUT&Tag.

刚地弓形虫（Toxoplasma gondii）是一种分布广泛的原生动物寄生虫，其生活史复杂且包含多个发育阶段。该寄生虫在不同发育阶段具备独特的基因表达模式，以支撑阶段特异性的生命活动，但其背后的调控机制在很大程度上仍未明确。本研究鉴定到一种可调控发育相关基因（尤其是裂殖子特异性基因）表达的核复合物。该复合物由AP2家族转录因子AP2Ⅻ-5、表观遗传因子MORC与HDAC3，以及一种能够稳定该复合物的新型AP2Ⅻ-5互作蛋白1（AIP1）组成。在寄生虫通过无性内二芽殖快速增殖的速殖子阶段，AP2Ⅻ-5结合至发育激活基因的启动子区域，并招募MORC与HDAC3以抑制靶基因的表达。当触发性分化启动与裂殖子发育进程时，AP2Ⅻ-5的蛋白丰度会下降，其对靶基因的抑制作用随之解除。与调控弓形虫发育且对速殖子生长不可或缺的MORC和HDAC3不同，AP2Ⅻ-5与AIP1对于体外培养条件下的速殖子增殖并非必需。上述实验数据表明，尽管MORC与HDAC3具备广泛的调控活性，但与AP2Ⅻ-5和AIP1形成复合物后，可使其在寄生虫发育过程中特异性调控基因表达。实验整体设计：利用CUT&Tag技术鉴定ME49株与ΔⅫ-5株基因组中HDAC3的结合位点。