STABILITY AND DISEASE TROPISM OF PROVIRAL DNAS

我们正在研究鸟类逆转录病毒生物学的两个独立方面： 鸡石骨症的诱导机制及稳定性 前病毒DNA 缺乏致癌基因的禽逆转录病毒通常主要产生 一种非急性疾病。 我们称这种疾病偏好为 “瞄准。“我们有兴趣确定病毒中的哪些序列 基因组负责骨硬化症“靶向”作为一个开始， 了解疾病诱发的机制。 我们的方法是 构建一种“靶向”骨硬化症的病毒的分子克隆 （NTRE-2），然后通过交换同源重组构建重组病毒。 NTRE-2和病毒之间的限制性内切酶片段， “靶向”淋巴瘤的产生（RAV-60 NY 203）或不 产生疾病（RAV-O）。 这些研究结果表明， LTR内的序列是疾病诱导所必需的， 在病毒结构基因中，最可能在gag或pol中的基因是必需的， 导致骨硬化症。 这代表了第一个关联 病毒结构蛋白与疾病的关系 我们将 将这些观察结果扩展到更小的限制性片段， 到DNA序列分析和位点特异性诱变的水平， 定义负责骨硬化症诱导的序列。 逆转录病毒前病毒DNA与 转座遗传因子经常被评论。 有 不管有多少证据表明， 转座基因元件的功能特征。 我们有 在几个克隆中观察到前病毒DNA模式的“不稳定性”， td-RSV感染QT-6细胞。 观察到的“不稳定”模式包括 前病毒序列的丢失和获得以及缺失向 相邻的细胞DNA 我们打算使用分子技术 克隆，DNA测序和染色质结构分析，以确定 特定的序列或DNA构象是否可以与前病毒 不稳定

We are studying two separate aspects of the biology of avian retroviruses: the mechanism of induction of avian osteopetrosis and the stability of proviral DNA. Avian retroviruses lacking oncogenes often produce primarily one form of non-acute disease. We have termed this disease preference "targeting." We are interested in defining which sequences in the viral genome are responsible for osteopetrosis "targeting" as a start to understanding the mechanism of disease induction. Our approach has been to construct a molecular clone of a virus which "targets" toward osteopetrosis (NTRE-2) and then to construct recombinant viruses by exchanging homologous restriction endonulease frangments between NTRE-2 and a virus which "targets" toward the production of lymphoma (RAV-60NY203) or which does not produce disease (RAV-O). Results of these studies suggest that sequences within the LTR are necessary for disease induction but that sequences within the viral structural genes most probably in gag or pol are required for the production of osteopetrosis. This represents the first association of a viral structural protein with the production of disease. We will extend these observations to smaller restriction fragments and ultimately to the level of DNA sequence analysis and site specific mutagenesis to define the sequences responsible for osteopetrosis induction. The similarity in structure between retrovirus proviral DNA and transposable genetic elements has been frequently commented upon. There is however little if any eveidence that proviruses possess any of the functional characteristics of transposable genetic elements. We have observed "instability" in the pattern of proviral DNA in several clones of td-RSV infected QT-6 cells. The patterns of "instability" observed include both loss and gain of proviral sequences and propagation of deletions into adjacent cellular DNA. We intend to use the techniques ol molecular cloning, DNA sequencing and chromatin structure analysis to determine whether specific sequences or DNA conformations can be correlated proviral instability.