Structural characterization of nucleoprotein cores of human adenoviruses

人腺病毒核蛋白核心的结构表征

• 批准号: 9807741
• 负责人: VIJAY S REDDY
• 金额: $ 30.5万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-06-01 至 2021-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9807741
• 关键词: 3-Dimensional; Acute; Adenovirus Protein; Adenoviruses; Applications Grants; Bacteriophages; Capsid; Capsid Proteins; Characteristics; Classification; Complex; Core Protein; Cryoelectron Microscopy; Data Set; Disease; Double Stranded DNA Virus; Endosomes; Epidemic Keratoconjunctivitis; Event; Exhibits; Fiber; Genes; Genome; Herpesviridae; Herpesvirus 1; Histones; Human; Image; In Situ; Infection; Knowledge; Location; Lung diseases; Methodology; Methods; Minor; Molecular; Nuclear; Nucleoproteins; Peptide Hydrolases; Proteins; Resolution; Role; Source; Structure; Techniques; Temperature; Vaccines; Viral; Viral Vector; Virion; Virus; base; density; ds-DNA; gastrointestinal infection; image reconstruction; insight; oncolytic virotherapy; organizational structure; particle; penton base; reconstruction; temperature sensitive mutant; thermostability

Human adenoviruses (HAdVs) are large (MW ~150 mDa), complex, non-enveloped dsDNA viruses that cause acute respiratory disease, epidemic keratoconjunctivitis and gastrointestinal infections. Significantly, adenoviruses (AdVs) are being used as viral vectors for gene, vaccine and Oncolytic viral therapies. Although the in situ structures and arrangement of major and minor capsid proteins (CPs) in the assembled (icosahedral) viral capsid have been determined at near-atomic resolution, there is no information currently available on how the nucleoprotein core, comprising linear dsDNA (~36 kbp) and “histone like” core proteins, is organized. The core density shows up as smeared ball in the icosahedral reconstructions of the AdVs. This is in contrast to characteristic spooled organization of the dsDNA seen in bacteriophages and herpesviruses. The details of nuclear core organization provide clues into the mechanism of genome packaging into AdVs, which is still unresolved. Here, we propose to employ emerging cryo-electron microscopy (Cryo-EM) methods to elucidate the structural organization of the AdV nuclear core by subtracting the icosahedral capsid regions from the particle (virion) images and reconstructing the core region(s) independently without imposing icosahedral symmetry. In Aim 1, we will reconstruct the nuclear core organization of mature HAdV5. Additionally, we will elucidate the reorganization of the nuclear core upon heat treatment that is known to mimic the structural changes that occur during partial disassembly and endosome escape. In Aim2 we will image the distinct nuclear core organization observed in hyper-thermostable and noninfectious form (ts1 mutant) of HAdV-C5 as well as elucidate the molecular basis of its greater thermo-stability of ts1 particles. Collectively, these studies provide structural details of nucleoprotein core organization in AdVs with implications for revealing the mechanism of genome packaging into AdVs as well as the changes that occur upon HAdV maturation. In addition, these studies will elucidate the changes that occur during the partial disassembly and infection of HAdV virions, emulated by the heat treatment.

人腺病毒（HADV）是大的（MW〜150 MDA），复杂的，非发育的DSDNA病毒，引起急性呼吸道疾病，流行病角结炎和胃肠道感染。值得注意的是，腺病毒（ADV）被用作基因，疫苗和溶瘤病毒疗法的病毒载体。尽管已在接近原子分辨率下确定了组装（二十面体）病毒衣壳中主要和小衣壳蛋白（CPS）的原位结构和排列，但目前尚无有关如何完成线性DSDNA（〜36 KBP）和“ HASTONE类似”的核蛋白质核心的信息。核心密度显示为ADV的二十面体重建中的涂抹球。这与细菌和疱疹病毒中看到的dsDNA的特征性船长组织形成对比。核心核组织的细节为基因组包装机制的ADV提供了线索，但仍未解决。在这里，我们建议采用新兴的低温电子显微镜（Cryo-EM）方法来阐明ADV核心核的结构组织，通过从粒子图像（视觉）图像中减去二十面体囊状区域并重建核心区域，而无需施加icosahedral Symetry。在AIM 1中，我们将重建成熟的HADV5的核心核组织。此外，我们将在热处理后阐明核核的重组，这是模仿部分拆卸和内体逃生过程中发生的结构变化。在AIM2中，我们将成像HADV-C5的超感染和非感染形式（TS1突变体）观察到的独特的核心核组织，并阐明其TS1颗粒的更大热稳定性的分子基础。总的来说，这些研究在ADV中提供了核蛋白质核心组织的结构细节，对揭示了基因组包装到ADV的机制以及HADV成熟后发生的变化的影响。此外，这些研究将阐明通过热处理模仿HADV病毒的部分拆卸和感染期间发生的变化。