STABILITY AND DISEASE TROPISM OF PROVIRAL DNAS

我们正在研究禽类逆转录病毒生物学的两个不同方面： 鸡骨化症的诱导机制及其稳定性研究 前病毒DNA。缺乏癌基因的禽类逆转录病毒通常主要产生 一种非急性疾病。我们把这种偏爱称为疾病偏好 “瞄准目标。”我们感兴趣的是定义病毒中的哪些序列 负责骨化病的基因组以“靶向”为起点 了解诱发疾病的机制。我们的方法一直是 构建“靶向”骨化病病毒的分子克隆 (NTRE-2)，然后通过交换同源基因构建重组病毒 NTRE-2与一种病毒之间的限制性内切酶片段 淋巴瘤的“靶点”(RAV-60NY203)或不是 产生疾病(RAV-O)。这些研究的结果表明，序列 是诱导疾病所必需的，但该序列 在病毒结构基因中，最可能存在于Gag或POL中是必需的 用于生产骨化病。这是第一个关联 病毒结构蛋白与疾病的产生有关。我们会 将这些观察扩展到更小的限制片段，并最终 到DNA序列分析和定点突变的水平 确定导致骨化症的序列。 逆转录病毒前病毒DNA与逆转录病毒DNA结构的相似性 可转座的遗传元件经常被评论。的确有 然而，很少有证据表明，普鲁斯拥有任何 可转座遗传元件的功能特征。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在 相邻的细胞DNA。我们打算用分子生物学的技术 克隆、DNA测序和染色质结构分析确定 特定序列或DNA构象是否可以与前病毒相关 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.