TgATAT-Mediated α-Tubulin Acetylation Is Required for Division of the Protozoan Parasite Toxoplasma gondii.
TgATAT-Mediated α-Tubulin Acetylation Is Required for Division of the Protozoan Parasite Toxoplasma gondii.
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DOI：
10.1128/msphere.00088-15
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发表时间：
2016-01
期刊：
影响因子：
4.8
通讯作者：
Sullivan WJ Jr
中科院分区：
文献类型：
--
作者：
Varberg JM;Padgett LR;Arrizabalaga G;Sullivan WJ Jr
弓形虫是一种机会性寄生虫，感染至少三分之一的世界人口。由于目前的药物对患者有毒，因此需要针对这种疾病（弓形体病）的新疗法。微管是由微管蛋白构建的重要细胞结构，有望成为抗菌药物靶点。微管可以通过化学修饰来调节，例如赖氨酸 40 (K40) 上的乙酰化。为了确定 K40 乙酰化在弓形虫中的作用以及它是否是寄生虫的负担，我们对 α-微管蛋白基因进行了突变分析。我们的结果表明，如果没有 K40 乙酰化，寄生虫就无法生存，除非微管通过二次突变稳定下来。此外，我们还鉴定了乙酰化 α-微管蛋白 (TgATAT) 的寄生虫酶。 TgATAT 的基因破坏导致寄生虫复制的严重缺陷，进一步凸显了弓形虫中 α-微管蛋白 K40 乙酰化的重要性及其作为潜在新药物靶点的前景。弓形虫是一种广泛分布的原生动物寄生虫，可引起潜在危及生命的机会性疾病。迫切需要新的寄生虫复制抑制剂，因为目前的抗叶酸治疗对患者也有毒害作用。微管是重要的细胞骨架成分，已被选择性地靶向微生物病原体；对弓形虫中微管蛋白的进一步研究可能会揭示新的治疗机会。人们注意到，赖氨酸 40 (K40) 处的 α-微管蛋白乙酰化在子代寄生虫形成过程中得到富集，但这种修饰对弓形虫分裂和介导其传递的酶的影响尚未确定。我们进行了突变分析，以证明 K40 乙酰化可以稳定弓形虫微管，并且是寄生虫复制所必需的。我们还表明，一种不寻常的弓形虫α-微管蛋白乙酰转移酶同源物（TgATAT）以细胞周期调节的方式表达，并且其表达在分裂期间达到峰值。使用 CRISPR/Cas9 破坏 TgATAT 可消除 K40 乙酰化并诱导复制缺陷；寄生虫似乎启动有丝分裂，但表现出不完整或不正确的核分裂。总之，这些发现确立了微管蛋白乙酰化的重要性，揭示了弓形虫的一个新的脆弱性，可以作为药理学目标。重要性 弓形虫是一种机会性寄生虫，感染至少三分之一的世界人口。由于目前的药物对患者有毒，因此需要针对这种疾病（弓形体病）的新疗法。微管是由微管蛋白构建的重要细胞结构，有望成为抗菌药物靶点。微管可以通过化学修饰来调节，例如赖氨酸 40 (K40) 上的乙酰化。为了确定 K40 乙酰化在弓形虫中的作用以及它是否是寄生虫的负担，我们对 α-微管蛋白基因进行了突变分析。我们的结果表明，如果没有 K40 乙酰化，寄生虫就无法生存，除非微管通过二次突变稳定下来。此外，我们还鉴定了乙酰化 α-微管蛋白 (TgATAT) 的寄生虫酶。 TgATAT 的基因破坏导致寄生虫复制的严重缺陷，进一步凸显了弓形虫中 α-微管蛋白 K40 乙酰化的重要性及其作为潜在新药物靶点的前景。
Toxoplasma gondii is an opportunistic parasite that infects at least one-third of the world population. New treatments for the disease (toxoplasmosis) are needed since current drugs are toxic to patients. Microtubules are essential cellular structures built from tubulin that show promise as antimicrobial drug targets. Microtubules can be regulated by chemical modification, such as acetylation on lysine 40 (K40). To determine the role of K40 acetylation in Toxoplasma and whether it is a liability to the parasite, we performed mutational analyses of the α-tubulin gene. Our results indicate that parasites cannot survive without K40 acetylation unless microtubules are stabilized with a secondary mutation. Additionally, we identified the parasite enzyme that acetylates α-tubulin (TgATAT). Genetic disruption of TgATAT caused severe defects in parasite replication, further highlighting the importance of α-tubulin K40 acetylation in Toxoplasma and its promise as a potential new drug target. Toxoplasma gondii is a widespread protozoan parasite that causes potentially life-threatening opportunistic disease. New inhibitors of parasite replication are urgently needed, as the current antifolate treatment is also toxic to patients. Microtubules are essential cytoskeletal components that have been selectively targeted in microbial pathogens; further study of tubulin in Toxoplasma may reveal novel therapeutic opportunities. It has been noted that α-tubulin acetylation at lysine 40 (K40) is enriched during daughter parasite formation, but the impact of this modification on Toxoplasma division and the enzyme mediating its delivery have not been identified. We performed mutational analyses to provide evidence that K40 acetylation stabilizes Toxoplasma microtubules and is required for parasite replication. We also show that an unusual Toxoplasma homologue of α-tubulin acetyltransferase (TgATAT) is expressed in a cell cycle-regulated manner and that its expression peaks during division. Disruption of TgATAT with CRISPR/Cas9 ablates K40 acetylation and induces replication defects; parasites appear to initiate mitosis yet exhibit incomplete or improper nuclear division. Together, these findings establish the importance of tubulin acetylation, exposing a new vulnerability in Toxoplasma that could be pharmacologically targeted. IMPORTANCE Toxoplasma gondii is an opportunistic parasite that infects at least one-third of the world population. New treatments for the disease (toxoplasmosis) are needed since current drugs are toxic to patients. Microtubules are essential cellular structures built from tubulin that show promise as antimicrobial drug targets. Microtubules can be regulated by chemical modification, such as acetylation on lysine 40 (K40). To determine the role of K40 acetylation in Toxoplasma and whether it is a liability to the parasite, we performed mutational analyses of the α-tubulin gene. Our results indicate that parasites cannot survive without K40 acetylation unless microtubules are stabilized with a secondary mutation. Additionally, we identified the parasite enzyme that acetylates α-tubulin (TgATAT). Genetic disruption of TgATAT caused severe defects in parasite replication, further highlighting the importance of α-tubulin K40 acetylation in Toxoplasma and its promise as a potential new drug target.