Structural basis of the polar tube invasion machinery from microsporidia parasites

微孢子虫寄生虫极管入侵机制的结构基础

• 批准号: 10563182
• 负责人: Gira Bhabha
• 金额: $ 68.34万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-03-01 至 2025-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10563182
• 关键词: 3-Dimensional; Acquired Immunodeficiency Syndrome; Address; Animals; Architecture; Biochemical; Biochemistry; Biological Assay; Biology; Biophysics; Bombyx; Categories; Cells; Cellular biology; Cilia; Classification; Complex; Cryo-electron tomography; Cryoelectron Microscopy; Data; Disease; Encephalitozoon cuniculi; Encephalitozoon hellem; Environment; Face; Farm; Fishes; Fluorescent Dyes; Freezing; Growth; Human; Image; Immunocompromised Host; In Situ; In Vitro; Individual; Infection; Insecta; Invaded; Ions; Light; Mass Spectrum Analysis; Microscopy; Microsporidia; Microsporidiosis; Microtubules; Modernization; Molecular; Molecular Conformation; Movement; Names; National Institute of Allergy and Infectious Disease; Optics; Organelles; Parasites; Patients; Process; Protein Subunits; Proteins; Proteome; Reproduction spores; Resolution; Sampling; Scanning Electron Microscopy; Sequence Homology; Side; Speed; Structure; Techniques; Thinness; Tube; Visualization; Work; X-Ray Crystallography; burden of illness; electron tomography; emerging pathogen; experimental study; genetic manipulation; human pathogen; in vivo; insight; light microscopy; lightspeed; mortality; opportunistic pathogen; organ transplant recipient; particle; pathogen; protein complex; protein protein interaction; protein purification; reconstruction; structural biology

Project Summary/Abstract Microsporidia are unicellular, ​fungal​ parasites with a wide host-range, from insects to humans. ​They are emerging pathogens, classified as NIAID Category B opportunistic pathogens, and cause microsporidiosis in immunocompromised patients.​ To gain entry into a target cell, microsporidia employ a remarkably unique and specialized harpoon-like invasion machinery called the polar tube, which is conserved among microsporidial species. While initially coiled neatly within the spore of the parasite, infection of a new cell begins with the rapid extrusion of the polar tube from the spore on a fast timescale (< 2s), which anchors the spore to the host cell. After it has been fired, the polar tube is thought to act as a conduit for the transfer of the infectious “sporoplasm” into the target cell, where replication can begin. Early work has yielded global insights into this process, and the molecular and structural underpinnings of the invasion process are ripe for exploration with modern techniques, such as cryo electron microscopy. This work aims to address fundamental questions and paradoxes in our understanding of the microsporidial polar tube machinery and how it drives invasion into host cells. We will use a combined bottom-up (structural biology, biochemistry and other ​in vitro​ techniques on purified proteins) and top-down (​in vivo​ light microscopy, electron tomography) approach; the intersection of these approaches will allow us to unravel the mechanistic biology of this unique invasion process. ​Here we focus on three human pathogens: ​Anncaliia algerae, Encephalitozoon cuniculi​ and ​Encephalitozoon hellem​. The specific aims are 1) To characterize the dynamics of polar tube firing and movement of sporoplasm through the tube using high-speed optical microscopy, and to comprehensively define the composition of the polar tube​ using mass spectrometry; 2) To biochemically and structurally characterize the individual protein components of the polar tube organelle using X-ray crystallography, single particle cryo electron microscopy and protein-protein interaction assays; 3) To elucidate the overall architecture and packing of the polar tube in the spore using structural cell biology techniques such as serial block face scanning electron microscopy (SBFSEM) and cryo focused ion beam scanning electron microscopy (cryo FIB-SEM) followed by cryo electron tomography (cryo ET).

项目总结/文摘