Organization of Toxoplasma invasion and cell division by EF-hand proteins

EF-hand 蛋白组织弓形虫入侵和细胞分裂

• 批准号: 8661114
• 负责人: Marc-Jan Gubbels
• 金额: $ 7.83万
• 依托单位: BOSTON COLLEGE
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-05-14 至 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8661114
• 关键词: Address; Affect; Affinity; Amino Acids; Binding; Binding Proteins; Calcineurin; Calmodulin; Calmodulin-Binding Proteins; Cell Cycle; Cell Division Process; Cell division; Cells; Centrosome; Complement; Complex; Congenital Abnormality; Contracts; Cre-LoxP; Cytoskeleton; DNA Repair; Data; Daughter; Disease; Dissection; EF Hand Motifs; EF-Hand Domain; Encephalitis; Essential Genes; Event; Family; Family member; Future; Gene Family; Genes; Genome; Goals; Helix-Loop-Helix Motifs; Immune response; Immunocompromised Host; Infection; Ionophores; Ions; Knock-out; Lead; Life; Life Cycle Stages; Lifting; Light; Lytic; Lytic Phase; Mammalian Cell; Mediating; Mitosis; Names; Nonmuscle Myosin Type IIA; Organelles; Organism; Orthologous Gene; Parasites; Pathogenesis; Pathology; Pattern; Play; Process; Protein Dynamics; Protein Family; Proteins; Research Personnel; Role; Signal Pathway; Signal Transduction Pathway; Structure; Time; Tissues; Toxoplasma; Toxoplasma gondii; Translating; Trimethoprim-Sulfamethoxazole; Work; calcineurin phosphatase; calcium-dependent protein kinase; cell motility; extracellular; follow-up; insight; member; mutant; protein function; public health relevance; research study; segregation

DESCRIPTION (provided by applicant): The apicomplexan parasite Toxoplasma gondii is the causative agent of life-threatening encephalitis in immunocompromised patients and in addition can cause a variety of birth defects if the infection is contracted congenitally. The pathology associated with disease originates in fast rounds of lytic intracellular replication cycles, causin extensive tissue damage. Since parasite replication only occurs within a cell from the mammalian host, the related processes of host cell invasion and egress mark essential steps in the pathogenesis. It has been firmly established that the signaling pathways underlying invasion and egress revolve around the cytoplasmic rise of Ca2+ concentration. Two small, Ca2+-binding proteins, calmodulin (CaM; 4 EF-hands) and the calcineurin phosphatase (the regulatory ¿-subunit has 4 EF hands), have been associated with activation of egress/invasion through pharmacological studies. Furthermore, a related group of small, 4-EF-hand containing proteins consisting of three centrins (Cen1-3) are critical components to the cell division machinery: Cen1 and Cen3 in centrosome duplication, required for mitosis and coordination of the unusual cell division process; Cen2 in contraction of the basal complex required for daughter parasite segregation. In essence, this family of EF-hand proteins plays pivotal roles in the defining steps of pathogenesis: invasion/egress and cell division. However this protein family has never been comprehensively studied, despite their critical roles in pathogenesis. To fill this void, the researchers identified a total of 16 such proteins in the genome, most of which have not been studied at all but likely execute additional functions in these processes. Preliminary sub-cellular localization dynamics of these proteins throughout the lytic cycle support this hypothesis and permits putative assignment to roles in cell division or invasion/egress. It is proposed to complete these preliminary studies for the whole family followed by conditional gene knock-outs of the ten prioritized candidates. To illustrate the power of this approach, these experiments will for the first time directly address whether CaM or calcineurin have a role in egress/invasion, for which only indirect pharmacological data is currently available. Furthermore, using the Cen2 and CaM as a pilot, the direct dissection of the role of Ca2+-binding in the family members with critical roles in invasion or cell division will be pioneered. It is anticipated that these studieswill lift the veil on the enigmatic role of Cen2 in Toxoplasma cell division and/or cytoskeleton structure. In conclusion, this study of a poorly studied gene family is expected to provide key insights into two key aspects in apicomplexan parasite pathogenesis.

描述（由申请方提供）：顶复门寄生虫弓形虫是免疫功能低下患者危及生命的脑炎的病原体，此外，如果先天感染，还可导致各种出生缺陷。与疾病相关的病理学起源于快速的细胞内溶解性复制循环，导致广泛的组织损伤。由于寄生虫复制只发生在哺乳动物宿主的细胞内，宿主细胞侵入和排出的相关过程标志着发病机制中的重要步骤。它已被牢固地建立，信号转导通路的基础上的入侵和出口围绕细胞质的Ca 2+浓度上升。两种小的Ca 2+结合蛋白，钙调蛋白（CaM; 4 EF-手）和钙调磷酸酶（调节亚基有4 EF手），已通过药理学研究与外出/侵入的激活相关。此外，一组相关的小的，4-EF-手包含蛋白质组成的三个中心蛋白（Cen 1 -3）是细胞分裂机制的关键组成部分：Cen 1和Cen 3在中心体复制，有丝分裂和协调不寻常的细胞分裂过程中所需的; Cen 2在收缩的基础复合物所需的子寄生虫分离。从本质上讲，EF-手蛋白家族在发病机制的定义步骤中起着关键作用：侵入/外出和细胞分裂。然而，这个蛋白质家族从未被全面研究过，尽管它们在发病机制中起着关键作用。为了填补这一空白，研究人员在基因组中总共确定了16种这样的蛋白质，其中大多数尚未被研究过，但可能在这些过程中执行额外的功能。初步亚细胞 这些蛋白质在整个裂解周期中的定位动力学支持这一假设，并允许假定的在细胞分裂或侵入/外出中的作用。建议完成整个家族的这些初步研究，然后对10个优先候选者进行条件基因敲除。为了说明这种方法的威力，这些实验将 第一次直接解决钙调素或钙调神经磷酸酶是否在外出/入侵中发挥作用，目前只有间接的药理学数据。此外，使用Cen 2和CaM作为试点，直接解剖的作用，Ca 2+结合在入侵或细胞分裂的关键作用的家庭成员将是开创性的。预计这些研究将揭开Cen 2在弓形虫细胞分裂和/或细胞骨架结构中神秘作用的面纱。总之，这项研究的一个研究不足的基因家族，预计将提供关键的见解两个关键方面的顶复门寄生虫的发病机制。