Molecular signaling mechanisms controlling Cryptosporidium proliferation and development

控制隐孢子虫增殖和发育的分子信号机制

• 批准号: 10548206
• 负责人: Sumiti Vinayak Alam
• 金额: $ 38.45万
• 依托单位: UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-01 至 2026-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10548206
• 关键词: Biochemical Genetics; Biological; Biological Assay; Biological Models; Biological Process; Biology; Calcium; Candidate Disease Gene; Cessation of life; Child; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Cryptosporidiosis; Cryptosporidium; Cryptosporidium parvum; Dependence; Development; Diarrhea; Disease; Disease Outbreaks; Drug Targeting; Epithelial Cells; Epitopes; FDA approved; Fertilization; Future; Gametogenesis; Genetic; Genome; Germ Cells; Goals; Homologous Gene; Human; Immunocompromised Host; In Vitro; Individual; Infant; Infection; Intestines; Knowledge; Laboratories; Life Cycle Stages; Light; Link; Mediating; Meiosis; Methods; Modeling; Molecular; Molecular Genetics; Mus; Nature; Oocysts; Parasite Control; Parasites; Pharmaceutical Preparations; Phenotype; Phosphorylation; Phosphotransferases; Plants; Plasmodium; Play; Proliferating; Protein-Serine-Threonine Kinases; Recreation; Resource-limited setting; Role; Rotavirus; Sexual Development; Signal Pathway; Signal Transduction; Signaling Protein; System; Technology; Testing; Toddler; Toxoplasma; Transgenic Organisms; Vaccines; Water; asexual; calcium-dependent protein kinase; candidate identification; chemical genetics; diarrheal disease; drug development; effective therapy; enteric infection; genetic approach; genetic manipulation; genome editing; immunological status; in vivo; insight; intestinal epithelium; kinase inhibitor; male; mouse model; mutant; nitazoxanide; novel; novel therapeutics; novel vaccines; parasite genome; pathogen; prevent; programs; protein degradation; reverse genetics; stem; superresolution microscopy; transmission process; vaccine development; waterborne outbreak

Project Abstract Cryptosporidium is a leading cause of diarrheal disease (cryptosporidiosis) and death among young children living in resource-poor settings. In the US, Cryptosporidium is the major cause of waterborne outbreaks linked to recreational water use. Currently, there is no fully effective drug and no vaccine to treat or prevent cryptosporidiosis. The only available US FDA approved drug, nitazoxanide has no proven efficacy in young children with weak immune status and immunocompromised individuals. Therefore, there is an urgent need to develop new drugs and vaccine to reduce the burden of cryptosporidiosis. Progress in anti-cryptosporidial drug and vaccine development has been hampered due to our limited understanding of parasite biology. The underlying reasons for this slow progress have been the unavailability of a robust method to continuously propagate Cryptosporidium, and the absence of molecular genetics to manipulate the parasite genome. We have overcome these hurdles by developing a powerful technology to manipulate the Cryptosporidium genome and propagate these genetically modified parasites in an immunocompromised mouse model system. The key advantage of this genetic system is that the entire life cycle of Cryptosporidium (both asexual and sexual stages) is completed in the mouse intestine, allowing us to unravel parasite biology (Vinayak et al 2015, Nature 523:477). We lack an understanding of the molecular signaling mechanisms that control development of parasite stages for successful completion of the complex life cycle. Signaling pathway components such as the plant-like calcium-dependent protein kinases (CDPKs) have emerged as attractive drug targets in Cryptosporidium and related parasites, due to the absence of their homologues in human host. Taking advantage of our genetic system, we have demonstrated the efficacy of selective bumped kinase inhibitors against calcium-dependent protein kinase-1 (CDPK1), thus indicating a critical role of this signaling kinase in C. parvum. Utilizing the conditional protein degradation system recently developed in our laboratory, we have demonstrated the essential role of CDPK1 in asexual proliferation and parasite survival. Moreover, we have compelling preliminary evidence that sheds light on the role of two signaling kinases in sexual developmental stages. The goal of this project is to elucidate the mechanistic role of these signaling proteins in regulating development of asexual and sexual stages in C. parvum required for parasite proliferation and transmission. Elucidation of these mechanisms will provide novel insights into the fundamental biology of Cryptosporidium, and open new avenues for development of effective therapies.

项目摘要 隐孢子虫是幼儿腹泻病（隐孢子虫病）和死亡的主要原因 生活在资源匮乏的环境中。在美国，隐孢子虫是水传播疾病爆发的主要原因 用于娱乐用水。目前，尚无完全有效的药物和疫苗可以治疗或预防 隐孢子虫病。硝唑尼特是美国 FDA 批准的唯一可用药物，尚未证实对年轻人有效 免疫状态较弱的儿童和免疫功能低下的个体。因此，迫切需要 开发新药和疫苗以减轻隐孢子虫病的负担。抗隐孢子虫药物研究进展 由于我们对寄生虫生物学的了解有限，疫苗的开发受到了阻碍。这 进展缓慢的根本原因是缺乏稳健的方法来持续 传播隐孢子虫，并且缺乏分子遗传学来操纵寄生虫基因组。我们有 通过开发强大的技术来操纵隐孢子虫基因组来克服这些障碍 在免疫功能低下的小鼠模型系统中繁殖这些转基因寄生虫。关键 该遗传系统的优点是隐孢子虫的整个生命周期（无性和有性阶段） 是在小鼠肠道中完成的，使我们能够解开寄生虫生物学的谜团（Vinayak et al 2015, Nature 523:477）。 我们对控制寄生虫阶段发育的分子信号机制缺乏了解 成功完成复杂的生命周期。信号通路成分，如植物样 钙依赖性蛋白激酶（CDPK）已成为隐孢子虫和隐孢子虫中有吸引力的药物靶点 相关寄生虫，由于在人类宿主中缺乏同源物。利用我们的基因优势 系统，我们已经证明了选择性碰撞激酶抑制剂对抗钙依赖性 蛋白激酶-1 (CDPK1)，从而表明该信号激酶在 C. parvum 中发挥着关键作用。利用 我们实验室最近开发了条件蛋白质降解系统，我们已经证明了必要的条件 CDPK1 在无性增殖和寄生虫存活中的作用。此外，我们还有令人信服的初步证据 这揭示了两种信号激酶在性发育阶段的作用。该项目的目标是 阐明这些信号蛋白在调节无性和有性发育中的机制作用 寄生虫增殖和传播所需的 C. parvum 阶段。对这些机制的阐明将 为隐孢子虫的基础生物学提供新颖的见解，并开辟新的发展途径 的有效疗法。