IDENTIFICATION OF HIV PACKAGING SEQUENCES; GENERATION OF PACKAGING CELL LINES

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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.