Transformation of C. elegans with a novel schistosome calcium channel subunit

用新型血吸虫钙通道亚基转化线虫

• 批准号: 7639108
• 负责人: ROBERT M GREENBERG
• 金额: $ 19.69万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-15 至 2011-03-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7639108
• 关键词: Adult; Affect; Biological; Biological Assay; Biological Models; Caenorhabditis elegans; Calcium; Calcium Channel; Cells; Development; Dose; Drug Delivery Systems; Future; Gene Expression; Genome; Goals; Heat-Shock Response; Individual; Laboratories; Measures; Methods; Molecular; Molecular Target; Muscle Cells; Muscle Contraction; Nematoda; Neurons; Paralysed; Parasitic Diseases; Pattern; Pharmaceutical Preparations; Pharyngeal structure; Physiological; Platyhelminths; Play; Praziquantel; Publishing; Role; Schistosoma; Schistosomiasis; Specificity; Staging; System; Testing; Tissues; Toxin; Transgenic Model; Transgenic Organisms; Tropical Disease; Variant; Work; cell type; high risk; innovation; model development; novel; pharynx muscle; promoter; public health relevance; research study; response; voltage

DESCRIPTION (provided by applicant): Trematode flatworms of the genus Schistosoma are the causative agents of schistosomiasis, a tropical parasitic disease with over 200 million people infected worldwide. Praziquantel (PZQ) is the current drug of choice against schistosomiasis, but the mechanism by which PZQ acts remains poorly defined, even several decades following its discovery. Our laboratory has shown that PZQ is likely interacting with a molecular component of schistosome voltage-gated Ca2+ (Cav) channels, specifically a structurally and functionally atypical Cav channel b subunit. This b subunit subtype has been found to date only in platyhelminths; PZQ is not effective against nematodes, and nematode genomes do not contain this variant Cav channel b subunit. This exploratory R21 proposal describes a high-risk/high-potential payoff project to transfer PZQ sensitivity to nematodes by transforming C. elegans with this variant schistosome Cav channel b subunit. We hypothesize that expression of the variant schistosome b subunit in appropriate C. elegans cell types will result in the formation of nematode Cav channels that are sensitive to PZQ. As such, we will place the schistosome b subunit under the control of several different tissue- and cell-specific C. elegans promoters. We will also use conditional promoters to control expression of the gene. If successful, these experiments will offer further compelling evidence that the schistosome variant b subunit is indeed targeted by PZQ, and will also provide the opportunity to acquire important information about the physiological role played by this unique b subunit. Additionally, development of this transgenic model could pave the way for further experiments (as an R01 project) to obtain details of the mode of action of PZQ, including the identification of interacting factors, by exploiting the far more tractable and powerful C. elegans model system. The specific aims of the project are to: 1. Produce transgenic C. elegans lines expressing the schistosome SmCavbvar Cav channel subunit under the control of various promoters. 2. Test transgenic C. elegans lines expressing bvar for sensitivity to PZQ, either in whole worms or in specific tissues. PUBLIC HEALTH RELEVANCE: Schistosomiasis is a major tropical disease caused by parasitic flatworms called schistosomes. Although there is an effective drug against schistosomiasis, the mechanism by which it works is unclear. By understanding that mechanism, it may be possible to obtain drugs that affect the same molecular target, or molecules that interact with that target. In this project, we are using an innovative approach to define the molecular target of this drug with a future goal of identifying other factors that might interact with this drug target.

描述（由申请人提供）：血吸虫属的Trematode扁虫是血吸虫病的病因，这是一种热带寄生虫病，全世界有超过2亿人感染了超过2亿人。 Praziquantel（PZQ）是目前针对血吸虫病的首选药物，但是PZQ作用的机制仍然很差，甚至在发现后几十年。我们的实验室表明，PZQ可能正在与血吸虫电压门控Ca2+（CAV）通道的分子成分相互作用，特别是结构和功能上的非典型CAV通道B亚基。迄今为止，该B亚基亚型仅在铂金分子中被发现。 PZQ对线虫无效，线虫基因组不包含此变体CAV通道B亚基。该探索性R21提案描述了一个高风险/高潜力的回报项目，通过使用这种变体的Schistosoms Cav通道B亚基通过转化秀丽隐杆线虫来将PZQ敏感性传递到线虫。我们假设在适当的秀丽隐杆线虫细胞类型中的变体螺旋体B亚基的表达将导致形成对PZQ敏感的线虫cav通道。因此，我们将将血吸虫B亚基置于几个不同组织和细胞特异性秀丽隐杆线虫启动子的控制之下。我们还将使用条件启动子来控制基因的表达。如果成功的话，这些实验将提供进一步的令人信服的证据，表明PZQ确实是PZQ的目标，并且还将提供机会，以获取有关该独特的B亚基扮演的生理角色的重要信息。此外，这种转基因模型的开发可以为进一步的实验（作为R01项目）铺平道路，以获取PZQ的作用模式（包括识别相互作用因子）的详细信息，通过利用更具可触发和强大的C. C. elegrans Model Model System。该项目的具体目的是：1。产生转基因秀丽隐杆线虫线，在各种启动子的控制下表达smcavbvar cav通道亚基。 2。在整个蠕虫或特定组织中，表达对PZQ敏感的BVAR的转基因秀丽隐杆线虫线。公共卫生相关性：血吸虫病是由称为血吸虫的寄生虫引起的主要热带疾病。尽管有一种有效的针对血吸虫病的药物，但其起作用的机制尚不清楚。通过了解该机制，可以获得影响相同分子靶靶的药物或与该靶标相互作用的分子。在这个项目中，我们正在使用一种创新的方法来定义该药物的分子靶标的未来目标，即确定可能与该药物靶标相互作用的其他因素。