ANALYSIS OF RETROVIRUS ASSEMBLY BY IN VITRO MUTAGENESIS

Project: Research project

Project Details

Description

RNA tumor viruses are important etiological agents for human disease, most notably causing human t-cell leukemia-lymphoma and acquired immune deficiency syndrome (AIDS). In spite of their importance, surprisingly little is known about the molecular mechanisms by which the components of these viruses are assembled. An understanding of this assembly process may be essential to devising novel approaches for the control retroviral diseases. The long-range goal of the work proposed here is to elucidate the roles of the gag gene products in the assembly of Rous sarcoma virus (RSV), the prototype retrovirus. The gag gene is known to encode the sole viral products needed for budding of this avian virus. These products do not function properly in mammalian cells, and their functions are not well understood even in avian cells. Attempts will be made to release the block to RSV replication that occurs in mammalian cells by constructing a myristylation site on the N-terminus of the gag product. The altered products will be expressed using an SV40 vector. A cis-acting element of gag that interferes with the replication of SV40-gag DNA will also be located. To investigate RSV assembly in avian cells, an in depth study of p27, the major capsid protein, will be undertaken. This gag product is probably essential for assembly as well as infection. The number, location, and function of the domains of p27 will be sought using the method of suppressor-linker mutagenesis. This method will allow the coding sequence of p27 to be saturated with small insertions, and the effects of these mutations on virus assembly will be determined after introducing them into avian cells. Nonsense mutations will also be produced by the mutagenesis scheme, and these will be used to determine the minimal length of the gag product that will still permit budding. For this purpose, a vaccinia virus vector will be developed to express the wild-type and truncated gag products.
StatusFinished
Effective start/end date8/1/877/31/91

Funding

  • National Cancer Institute

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