IDENTIFICATION OF HIV PACKAGING SEQUENCES ; GENERATION OF PACKAGING CELL LINES

在这个项目中，我们建议确定的序列， 有效包装人体免疫机能丧失病毒/艾滋病 病毒（HIV）基因组。 在RSV或MLV等肿瘤病毒中，顺式作用 包装所需的序列（但可能不足够） 基因组RNA中的RNA前导序列已被确定 位于负链tRNA引物结合位点之间， gag基因的起始 艾滋病毒的类似部分 基因组，以及其他5'基因组区域将被删除， 以确定对HIV基因组包装的影响。 这些序列也将被工程化到异源细胞中。 表达载体，以确定它们是否赋予 包装特异性。 由于认为肿瘤病毒gag基因 产品与前导序列相互作用以促进基因组 包装，纯化的gag蛋白和突变体， 将分析gag基因以确定是否存在特异性RNA-蛋白 相互作用发生在HIV基因组RNA包装中。 信息 从这些实验中收集的数据将用于 构建有效的、非致细胞病变的HIV包装线， 可以用来将病毒或其他序列递送到 HIV靶细胞。 顺式和反式作用因子的鉴定 艾滋病病毒包装将是有益的建设占主导地位 用于抑制病毒的负突变和反义载体 复制的

In this project we propose to identify the sequences which are required for efficient packaging of the human immunodeficiency virus (HIV) genome. In oncoviruses such as RSV or MLV cis-acting sequences necessary (but perhaps not sufficient) for packaging of genomic RNA have been identified in the RNA leader sequence located between the minus-strand tRNA primer binding site and the start of the gag gene. The analogous portion of the HIV genome, as well as other 5' genomic regions will be deleted in order to determine the effect on packaging of the HIV genome. These sequences will also be engineered into heterologous expression vectors in order to determine whether they confer packaging specificity. Since it is thought that oncoviral gag gene products interact with leader sequences to facilitate genomic packaging, purified gag proteins and mutants engineered in the gag gene will be analyzed to determine if specific RNA-protein interactions occur in HIV genomic RNA packaging. Information gathered from these experiments will be utilized in order to construct efficient, noncytopathic HIV packaging lines which could be utilized in order to deliver viral or other sequences into HIV target cells. Identification of cis-and trans-acting factors in HIV packaging will be useful in the construction of dominant negative mutations and antisense vectors for inhibition of viral replication.