STRUCTURE AND FUNCTION OF THE ALEUTIAN MINK DISEASE PARVOVIRUS GENOME

这个项目正在进行的目的是比较基因组结构 阿留申水貂病致病株和非致病株的研究 细小病毒(ADV)，并将这些差异与 各种病毒分离株在细胞培养中复制并引起 受感染水貂中的疾病。 通过构建非致病性AdV-G嵌合克隆 (类型1 ADV)和ADV-犹他州1(类型2 ADV)，我们现在已经确定了 影响ADV致病性的片段。白喉的分子克隆 包含复制的ADV-Utah 2的73-88地图单元片段的ADV-G 2log10在细胞培养中的效率低于AdV-G亲本，但 水貂注射病毒诱导产生抗体并致病 从这些克隆人身上。该片段映射到衣壳蛋白编码 序列，类型1和类型之间有4个编码差异 2病毒。此外，该区域与54-63年地图单位不同 衣壳基因片段，控制细胞培养中的通透性。然而， 犬细小病毒的计算机分析及序列比较 (CPV)提示这两个区域与CPV衣壳上的区域对齐 调节寄主范围和致病性。 为了将这些发现与Adv病毒粒子结构联系起来，我们有 开始使ADV具体化的工作。我们正在利用一种重组杆状病毒 来合成衣壳蛋白。衣壳蛋白自组装成 具有物理特性的空腺病毒粒子。 在其他工作中，我们已经开始了分析结构-功能的研究 Adv P36启动子的相互作用。高级动词的顺序和间距 P36分子与其他原型细小病毒显著不同， 这些变化可能解释了低组成和反式激活 发起人的实力。进行了一系列的改动 启动子，其中一些在CAT检测中恢复了启动子的强度，但 当这些变化被引入到ADV的全长克隆中时，所有 克隆是复制缺陷的。

The ongoing purpose of this project is to compare the genome structure of pathogenic and nonpathogenic strains of Aleutian mink disease parvovirus (ADV) and to relate these differences to the ability of the various isolates of virus to replicate in cell culture and to cause disease in infected mink. By constructing clones that are chimeric between the nonpathogenic ADV-G (type 1 ADV) and ADV-Utah 1 (type 2 ADV), we have now identified a segment that influences the pathogenicity of ADV. Molecular clones of ADV-G that contained the 73-88 map unit piece of ADV-Utah 2 replicated 2log10 less efficiently than the ADV-G parent in cell culture, but induced antibody and caused disease in mink injected with virus derived from these clones. This segment maps to the capsid protein coding sequences and there are 4 coding differences between the type 1 and type 2 virus. Furthermore, the region is distinct from the 54-63 map unit capsid gene segment that governs permissivity in cell culture. However, computer analysis and comparison with the sequence of canine parvovirus (CPV) suggests that both regions align with ones on the CPV capsid regulating host range and pathogenicity. In order to relate these findings to the ADV virion structure, we have begun work to crystallize ADV. We are utilizing a recombinant baculovirus that synthesizes capsid proteins. The capsid proteins self-assemble into particles with physical characteristics of empty ADV virions. In other work, we have begun studies to analyze structure-function interactions of the ADV P36 promoter. The sequence and spacing of ADV P36 elements differs significantly from other prototypic parvoviruses and these variations may explain the low constitutive and trans-activated strength of the promoter. A number of alterations were produced in the promoter, some of which "restored" promoter strength in CAT assays, but when the changes were introduced into full-length clones of ADV, all the clones were replication defective.