WOODCHUCK VIRUS--MOLECULAR BIOLOGICAL STUDIES

土拨鼠肝炎病毒（WHV）是分类学和 血清学上与B型肝炎病毒（HBV）相关。 感染 与这些病毒中的每一种都与急性和慢性 肝炎和肝细胞癌; 这些关联似乎是病因学性质的。 因此 WHV/土拨鼠系统提供了一个相关的和方便的模型， 了解人类的HBV感染。 以下实验 成功完成：（1）WHV专用试验 开发了抗原-抗体系统。 (2)重组WHV DNA 被转染到敏感的土拨鼠的肝脏中， 和慢性WHV感染。 (3)基因改变的WHV 在病毒X基因序列中有突变的基因组， 通过重组DNA技术构建。 突变病毒基因组 在土拨鼠中没有感染性，这意味着这些X基因 序列是病毒复制所必需的。 (4)超螺旋 从感染的肝细胞核中提取的WHV， 克隆并确定完整的核苷酸序列。 的 这种复制中间体的序列与 在病毒体中发现的部分双链WHV基因组。 (5)的 另外三个WHV基因组的完整核苷酸序列， 测定 似乎WHV的病毒亚型正在进化， 类似于HBV的表面抗原亚型。 (6)体内 测定WHV聚合酶的保真度。 突变率 对于WHV复制，发现每 基因组，这是低于突变率， 逆转录病毒和其它单链RNA病毒。 (7)两 WHV基因组X基因区特异性转录物， 发现了 一个转录本，长度为0.65 kb，定位于 感染肝细胞的细胞核，并发现缺乏poly（A） 在其3'末端的尾部。 第二个转录本，长度为0.80 kb， 显示为0.65kb转录物的多腺苷酸化形式。

The woodchuck hepatitis virus (WHV) is taxonomically and serologically related to the hepatitis B virus (HBV). Infection with each of these viruses is associated with acute and chronic hepatitis and hepatic cell carcinoma in their respective hosts; these associations appear to be etiological in nature. Thus, the WHV/woodchuck system provides a relevant and convenient model for understanding HBV infections of humans. The following experiments were successfully completed: (1) Tests specific for the WHV antigen-antibody systems were developed. (2) Recombinant WHV DNA was transfected into the liver of susceptible woodchucks and acute and chronic WHV infections produced. (3) Genetically altered WHV genomes, with mutations in the virus X gene sequence, were constructed by recombinant DNA techniques. Mutant virus genomes were non-infectious in woodchucks, which implied that these X gene sequences were essential for virus replication. (4) Supercoiled WHV, from the nucleus of infected liver cells, was successfully cloned and the complete nucleotide sequence determined. The sequence of this replicative intermediate is identical to that of the partially double stranded WHV genome found in virions. (5) The complete nucleotide sequence of three additional WHV genomes was determined. It appears that virus subtypes of WHV are evolving, analogous to the surface antigen subtypes of HBV. (6) The in vivo fidelity of the WHV polymerase was determined. The mutation rate for WHV replication was found to be less than 2 X 10-4 changes per genome per year, which is lower than the mutation rate of retroviruses and other single strand RNA viruses. (7) Two transcripts specific for the X gene region of the WHV genome were discovered. One transcript, 0.65 kb in length, was localized to the nucleus of infected liver cells, and found to lack a poly(A) tail at its 3' end. The second transcript, 0.80 kb in length, was shown to be the poly-adenylated version of the 0.65 kb transcript.