Structure and function of the Plasmodium myosin XIV-actin glideosome.

疟原虫肌球蛋白 XIV-肌动蛋白滑胶体的结构和功能。

• 批准号: 10650841
• 负责人: KATHLEEN M TRYBUS
• 金额: $ 66.16万
• 依托单位: UNIVERSITY OF VERMONT & ST AGRIC COLLEGE
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-05-11 至 2027-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10650841
• 关键词: ATP phosphohydrolase; Actins; Actomyosin; Africa; Age; Artemisinins; Binding; Binding Sites; Biological; Biological Assay; Cell division; Cell physiology; Cells; Cellular biology; Cessation of life; Child; Complex; Cryoelectron Microscopy; Crystallization; Culicidae; Development; Drug resistance; Erythrocytes; Foundations; Future; Genetic; Genetic study; Goals; Heart; Human; In Vitro; Invaded; Investigation; Kinetics; Licensure; Life Cycle Stages; Light; Malaria; Malaria Vaccines; Molecular; Motion; Motor; Myosin ATPase; Myosin Type V; N-terminal; Organelles; Parasites; Parasitic infection; Pathogenesis; Pharmaceutical Preparations; Phosphorylation; Phosphorylation Site; Plasmodium; Plasmodium falciparum; Play; Positioning Attribute; Power stroke; Process; Property; Reporting; Research; Role; Sexual Development; Sporozoites; Structure; Tissues; Vaccines; Virulent; Work; X-Ray Crystallography; cell motility; conditional knockout; druggable target; global health; imaging study; in vivo; inhibitor; insight; live cell imaging; malaria infection; mutant; myosin VI; new therapeutic target; novel; optic trap; optical traps; small molecule inhibitor; superresolution imaging; tool; trafficking; virtual

Malaria infection in humans, caused by single-celled parasites from the genus Plasmodium, is a major global health challenge. Despite marked progress in the last 15 years, more than 400 million deaths occur worldwide annually, the majority being children under age 5. Recent licensure of the first ever malaria vaccine heralds a new era in efforts to control malaria, but the relatively modest efficacy of the RTS,S vaccine means that complementary approaches will be essential if the WHO's goal of a 90% reduction in rates by 2030 is to be realized. Malaria parasites are motile throughout their complex human and mosquito lifecycle. They move by a process called gliding motility, which underpins their ability to reach, cross, and enter host tissues and cells. Gliding is powered by a parasite actomyosin motor the disruption of which kills the infectious parasite. Towards development of the parasite actomyosin motor as a druggable target, our collaborative team has worked to characterize the essential class XIV single-headed myosin motor PfMyoA, the core of gliding motility. We were the first to characterize and crystallize PfMyoA, demonstrating that its function is uniquely tuned by N-terminal heavy chain phosphorylation. We were the first to show the essential role of PfMyoA and its essential light chain in powering red blood cell (RBC) invasion, the stage responsible for all malaria pathogenesis. We have since used PfMyoA mutants to reveal the energetic barriers necessary for RBC invasion using live cell imaging. These foundations expertly position our team to extend investigation of gliding motility across the malaria lifecycle and explore additional Plasmodium myosins and their cellular roles, which are the combined aims of this competitive renewal. We propose (Aim 1) to define the cellular roles of PfMyoB versus PfMyoA by comparing structures, functional properties, and the role of heavy chain phosphorylation in vitro and in vivo. Aim 2 proposes to determine the binding pocket, mechanism of action, and impact on the parasite of two first-in-class small molecule inhibitors of PfMyoA ATPase activity. Aim 3 investigates two other essential Plasmodium myosins (PfMyoF and K), which are virtually unstudied both as motors and potential future druggable targets. PfMyoF likely plays a role as a processive transporter, while PfMyoK likely functions during sexual development, with motor domain inserts typical of reverse-directionality in eukaryotic class VI motors. We will use an integrative approach highlighting in vitro functional assays (motility and ensemble force assays, optical trap assays, steady- state and transient kinetics) and structural studies (X-ray crystallography and cryo-EM) together with live cell approaches (including super resolution imaging) and genetic investigation of motors (conditional knockouts/substitutions) in several stages of the Plasmodium parasite lifecycle. At completion we will have developed a previously unattainable depth of understanding into the function of the essential Plasmodium myosins as druggable targets, revealed profound insights into the structural basis by which myosins produce force and motion, and discovered fundamental insights into malaria parasite cell biology.

由疟原虫属的单细胞寄生虫引起的人类疟疾感染是全球主要的疟疾流行病， 健康挑战。尽管过去15年取得了显著进展，但全世界仍有4亿多人死亡 大多数是5岁以下的儿童。最近首次疟疾疫苗获得许可， 新时代的努力，以控制疟疾，但相对温和的效力RTS，S疫苗意味着， 如果要实现世界卫生组织到2030年将发病率降低90%的目标， 实现了疟疾寄生虫在其复杂的人类和蚊子生命周期中是能动的。它们移动的速度 这一过程被称为滑动运动，它支撑着它们到达、穿过和进入宿主组织和细胞的能力。 滑翔是由寄生虫的肌动球蛋白发动机驱动的，这种发动机的破坏会杀死传染性寄生虫。朝向 寄生虫肌动球蛋白马达作为药物靶点的发展，我们的合作团队一直致力于 描述了必需的XIV类单头肌球蛋白马达PfMyoA，滑行运动的核心。我们 第一个表征和结晶PfMyoA，证明其功能是独特的N-末端调节， 重链磷酸化。我们是第一个显示PfMyoA及其必需轻链的重要作用的人。 在推动红细胞（RBC）入侵，负责所有疟疾发病机制的阶段。我们已经 使用PfMyoA突变体，使用活细胞成像揭示RBC侵入所必需的能量屏障。这些 基金会专业地定位我们的团队，以扩大整个疟疾生命周期的滑行运动的调查， 探索更多的疟原虫肌球蛋白和它们的细胞作用，这是这项竞争性研究的综合目标。 退款我们提出（目的1）通过比较结构来定义PfMyoB与PfMyoA的细胞作用， 功能特性，以及体外和体内重链磷酸化的作用。目标2建议， 确定结合口袋，作用机制，并对寄生虫的影响，两个一流的小 PfMyoA ATP酶活性的分子抑制剂。目的3研究其他两种必需的疟原虫肌球蛋白 （PfMyoF和K），这是几乎未经研究的电机和潜在的未来药物的目标。PfMyoF PfMyoK可能在性发育过程中发挥作用，而PfMyoK可能在性发育过程中发挥作用， 马达域插入物在真核VI类马达中具有典型的反向性。我们将使用一个综合 方法强调体外功能测定（运动性和系综力测定，光阱测定，稳态- 状态和瞬态动力学）和结构研究（X射线晶体学和冷冻电镜）以及活细胞 方法（包括超分辨率成像）和电机的遗传研究（条件 基因敲除/置换）在疟原虫寄生虫生命周期的几个阶段。完成后，我们将有 对疟原虫的基本功能有了前所未有的深入了解 肌球蛋白作为药物靶点，揭示了肌球蛋白产生的结构基础的深刻见解 力和运动，并发现了疟疾寄生虫细胞生物学的基本见解。

项目成果

Rapid tool for cell nanoarchitecture integrity assessment.

• DOI: 10.1016/j.jsb.2021.107801
• 发表时间: 2021-12
• 期刊: Journal of structural biology
• 影响因子: 3
• 作者: Gaietta G;Swift MF;Volkmann N;Hanein D
• 通讯作者: Hanein D

Mechanism of small molecule inhibition of Plasmodium falciparum myosin A informs antimalarial drug design.

• DOI: 10.1038/s41467-023-38976-7
• 发表时间: 2023-06-12
• 期刊: NATURE COMMUNICATIONS
• 影响因子: 16.6
• 作者: Moussaoui, Dihia;Robblee, James P.;Robert-Paganin, Julien;Auguin, Daniel;Fisher, Fabio;Fagnant, Patricia M.;Macfarlane, Jill E.;Schaletzky, Julia;Wehri, Eddie;Mueller-Dieckmann, Christoph;Baum, Jake;Trybus, Kathleen M.;Houdusse, Anne
• 通讯作者: Houdusse, Anne

The actomyosin interface contains an evolutionary conserved core and an ancillary interface involved in specificity.

• DOI: 10.1038/s41467-021-22093-4
• 发表时间: 2021-03-25
• 期刊: Nature communications
• 影响因子: 16.6
• 作者: Robert-Paganin J;Xu XP;Swift MF;Auguin D;Robblee JP;Lu H;Fagnant PM;Krementsova EB;Trybus KM;Houdusse A;Volkmann N;Hanein D
• 通讯作者: Hanein D

Plasmodium myosin A drives parasite invasion by an atypical force generating mechanism.

疟原虫肌球蛋白 A 通过非典型的力产生机制驱动寄生虫入侵。

• DOI: 10.1038/s41467-019-11120-0
• 发表时间: 2019
• 期刊: Nature communications
• 影响因子: 16.6
• 作者: Robert-Paganin,Julien;Robblee,JamesP;Auguin,Daniel;Blake,ThomasCA;Bookwalter,CarolS;Krementsova,ElenaB;Moussaoui,Dihia;Previs,MichaelJ;Jousset,Guillaume;Baum,Jake;Trybus,KathleenM;Houdusse,Anne
• 通讯作者: Houdusse,Anne

Full-length Plasmodium falciparum myosin A and essential light chain PfELC structures provide new anti-malarial targets.

• DOI: 10.7554/elife.60581
• 发表时间: 2020-10-13
• 期刊: eLife
• 影响因子: 7.7
• 作者: Moussaoui D;Robblee JP;Auguin D;Krementsova EB;Haase S;Blake TCA;Baum J;Robert-Paganin J;Trybus KM;Houdusse A
• 通讯作者: Houdusse A