STUDIES OF PROVIRAL-DNA SYNTHESIS DURING FV-1 MEDIATED HOST RESTRICTION OF FRIEND MURINE LEUKEMIA VIRUS

Abstract

The basis for the restriction of retroviral replication encoded by the murine locus, Fv-1, was investigated. Previous studies had indicated that Fv-1 host restriction of viral replication occurred early during the replication cycle, following adsorption and penetration but prior to or during the integration of proviral DNA. Therefore, this thesis examined the synthesis of proviral DNA in Fv-1 restrictive hosts relative to permissive hosts. In this way, it was hoped that the events surrounding Fv-1 mediated host restriction of proviral DNA integration could be elucidated.;The technique of Southern blot hybridization was used to examine the various forms of proviral DNA synthesized in permissive and restrictive cells infected with Friend MuLV. Following infection of nonpermissive cells, the accumulation of unintegrated viral DNA was inhibited at the level of form I DNA, form III DNA, or not at either level compared to permissive infection. Quantitative measurements of the difference observed in nonpermissive infections indicated that the decrease in form I DNA did not directly correlate with the degree of biological restriction observed in tissue culture assays. Furthermore, structural studies indicated that the viral DNA accumulated in Fv-1 restrictive hosts was not grossly different in size or sequence compared to unintegrated viral DNA synthesized in permissive hosts. Experiments involving the DNA polymerase inhibitor aphidicolin indicated that the accumulation of the normally observed large amounts of form I DNA need not occur in order for the permissively infected cell to produce viral progeny. In addition, the form I DNA obtained from a Fv-1 restrictive infection was infectious when microinjected into either permissive or nonpermissive cells.;The results of this thesis suggest that Fv-1 restriction of viral replication occurs during the events surrounding the integration of proviral DNA. The inability to accumulate form I DNA in certain restrictive infections may not fully explain the restriction of viral replication. A variety of factors, including viral and host products, may be involved in the integration of proviral DNA and represent potential sites for the Fv-1 restrictive event(s).