ADENOASSOCIATED VIRUS (AAV) REP PROTEINS MEDIATE COMPLEX-FORMATION BETWEEN AAV DNA AND ITS INTEGRATION SITE IN HUMAN DNA

AAV is unique among eukaryotic viruses in the ability of its DNA to in tegrate preferentially into a specific region of the human genome. Und erstanding AAV integration may aid in developing gene therapy systems with predictable integration sites. Using a gel mobility-shift assay, we have identified a DNA sequence within the AAV integration locus on human chromosome 19 which is specifically bound by the AAV Rep78 and R ep68 proteins. This Rep recognition sequence is a GCTC repeating moth very similar to sequences within the inverted terminal repeats of the AAV genome which are also bound by Rep78 and Rep68. Cloned oligonucleo tides containing the recognition sequence can direct specific binding by Rep proteins. Binding assays with mutant Rep proteins show that the amino-terminal portion of Rep78 and Rep68 can direct binding to eithe r the AAV terminal repeat hairpin DNA or chromosome 19. This human gen omic DNA can be complexed with AAV DNA by Rep proteins as demonstrated by a dual-label (P-32/biotin) assay. These results suggest a role for Rep in targeting viral integration.