RESTRICTION checkpoint controls bradyzoite development in Toxoplasma gondii. RESTRICTION checkpoint controls bradyzoite development in Toxoplasma gondii

The Toxoplasma gondii G1 RESTRICTION checkpoint operates the switch between parasite growth and differentiation. The Cdk-related G1 kinase TgCrk2 forms alternative complexes with atypical cyclins (TgCycP1, TgCycP2 and TgCyc5) in the rapidly dividing developmentally incompetent RH and slower dividing developmentally competent ME49 tachyzoites and bradyzoites. The TgCycP1 expression interferes with bradyzoite differentiation. The TgCycP2 regulates G1 in the developmentally competent ME49 but not in the developmentally incompetent RH tachyzoites. Examination of TgCycP2 and TgCyc5 in alkaline induced and spontaneous bradyzoite differentiation (rat embryonic brain cells) models confirmed TgCycP2 role in bradyzoite replication and revealed that stress induced TgCyc5 is critical for efficient tissue cyst maturation. Overall design: Total RNA was extracted from the purified tachyzoites and bradyzoites of the ME49DKu80Dhxgprt AtTIR1 strain and derivatives. RNA-Seq Library were prepared and sequenced according to manufacturer’s instructions (Illumina).

刚地弓形虫（Toxoplasma gondii）的G1限制检验点（G1 RESTRICTION checkpoint）调控寄生虫生长与分化的转换。周期蛋白依赖性激酶（Cyclin-dependent kinase, Cdk）相关G1激酶TgCrk2，在快速分裂且发育无能的RH株速殖子，以及分裂较慢、具备发育能力的ME49株速殖子与缓殖子中，可与非典型周期蛋白（TgCycP1、TgCycP2和TgCyc5）形成不同的复合物。TgCycP1的表达会干扰缓殖子分化。TgCycP2在具备发育能力的ME49株中调控G1期，但在发育无能的RH株速殖子中无此调控作用。在碱性诱导及自发缓殖子分化（大鼠胚胎脑细胞）模型中对TgCycP2与TgCyc5的检测，证实了TgCycP2在缓殖子复制中的作用，并揭示应激诱导的TgCyc5对于高效组织囊成熟具有关键意义。实验设计：从ME49DKu80Dhxgprt AtTIR1菌株及其衍生株的纯化速殖子和缓殖子中提取总RNA，按照制造商（Illumina）的操作指南制备RNA测序文库并完成测序。