FOOTPRINTS ON THE VIRAL-DNA ENDS IN MOLONEY MURINE LEUKEMIA-VIRUS PREINTEGRATION COMPLEXES REFLECT A SPECIFIC ASSOCIATION WITH INTEGRASE

Retroviral DNA integration is mediated by the preintegration complex, a large nucleoprotein complex derived from the core of the infecting v irion. We previously have used Mu-mediated PCR to probe the nucleoprot ein organization of Moloney murine leukemia virus preintegration compl exes. A region of protection spans several hundred base pairs at each end of the viral DNA, and strong enhancement are present near the term ini, Here, we show that these footprints reflect a specific associatio n between integrase and the viral DNA ends in functional preintegratio n complexes. Barrier-to-autointegration factor, a cellular protein tha t blocks autointegration of Moloney murine leukemia virus DNA, also pl ays an indirect role in generating the footprints at the ends of the v iral DNA. We have exploited Mu-mediated PCR to examine the effect of m utations at the viral DNA termini on complex formation. We find that a replication competent mutant with a deletion at one end of the viral DNA still exhibits a strong enhancement about 20 hp from the terminus of the mutant DNA end. The site of the enhancement therefore appears t o be at a fixed distance from the ends of the viral DNA. We also find that a mutation at one end of the viral DNA, which renders the virus i ncompetent for replication, abolishes the enhancements and protection at both the U3 and U5 ends. A pair of functional viral DNA ends theref ore are required to interact before the chemical step of 3' end proces sing.