Molecular determinants for Rous sarcoma virus intasome assemblies

Integration of retroviral DNA into the host genome involves formation of integrase (IN)-DNA complexes termed intasomes. Here, we report the single-particle cryo-EM structure of the Rous sarcoma virus (RSV) strand transfer complex (STC) intasome produced with IN and a preassembled viral/target DNA substrate. The STC structure had an overall resolution of 3.36 Å and 3 Å in the conserved intasome core (CIC) region. Our structure demonstrated the flexibility of the distal IN subunits relative to the IN subunits in the CIC, similar to previously shown with the RSV octameric cleaved synaptic complex (CSC) intasome produced with IN and viral DNA only. An extensive analysis of higher-resolution STC structure helped in identification of nucleoprotein interactions important for intasome assembly. Using structure-function studies, we determined the mechanisms of several IN-DNA interactions critical for assembly of both RSV intasomes. We determined the role of IN residues R244, Y246 and S124 in CSC and STC intasome assemblies and their catalytic activities, demonstrating differential effects. Taken together, these studies advance our understanding of different RSV intasome structures and molecular determinants involved in their assembly.