STRUCTURE AND FUNCTION OF THE ALEUTIAN MINK DISEASE PARVOVIRUS GENOME

这个项目的目的是比较基因组结构 水貂阿留申病的致病性和非致病性菌株 细小病毒（ADV），并与这些差异的能力， 各种病毒分离株在细胞培养物中复制， 感染水貂的疾病。 通过构建非致病性ADV-G (type 1 ADV）和ADV-Utah 1（2型ADV），我们现在已经确定了一种 影响ADV致病性的基因片段。 性克隆 ADV-G包含ADV-Utah 2的73-88地图单元片，复制 在细胞培养中比ADV-G亲本低2log 10，但 水貂注射病毒衍生物后产生抗体并致病 这些克隆人。 该片段映射到衣壳蛋白编码 序列之间存在4个编码差异，类型1和类型2之间存在4个编码差异。 2病毒。此外，该区域与54-63地图单元不同 在细胞培养物中控制细胞凋亡的衣壳基因片段。 但是，在这方面， 犬细小病毒基因序列的计算机分析与比较 (CPV)表明这两个区域与CPV衣壳上的区域一致 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 我们开始研究ADV的结晶。我们正在利用重组杆状病毒 合成衣壳蛋白。衣壳蛋白自组装成 具有空ADV病毒粒子的物理特征的颗粒。 在其他工作中，我们已经开始研究分析结构-功能 ADV P36启动子的相互作用。 ADV的序列和间距 P36元件与其他原型细小病毒显著不同， 这些变化可以解释低组成性和反式激活 推动者的力量。 一些改动是在 启动子，其中一些在CAT测定中“恢复”启动子强度，但 当这些变化被引入ADV的全长克隆中时，所有的 克隆是复制缺陷型的。

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.