Biochemical and genetic analyses of integrase-interacting proteins lens epithelium-derived growth factor growth factor related protein 2 function and (LEDGF)/p75 and hepatoma-derived (HRP2) in preintegration complex HTV-1 replication

Human immunodeficiency virus type 1 (HIV-1) integrase (IN) functions in cells within the context of high molecular weight preintegration complexes (PICs). Lens epithelium-derived growth factor (LEDGF) transcriptional coactivator/p75 and hepatoma-derived growth factor related protein 2 (HRP2) tightly bind to HIV-1 IN and stimulate its integration activity in vitro. Here, we show that each recombinant host cell factor efficiently reconstitutes the in vitro activity of HIV-1 PTCs disrupted for functional integration by pre-treatment with high concentrations of salt. Mutational analysis reveals that both the IN-binding and I)NA-binding activities of LEDGF/p75 contribute to functional PIC reconstitution. We also investigate a role(s) for these proteins in HIV-1 infection by using short-interfering RNA. HIV-1 infection was essentially unaffected in HeLa-P4 cells depleted for LEDGF/p75, HRP2, or both proteins. We conclude that cells knocked-out for LEDGF/p75 and/or HRP2 will be useful genetic tools to address the roles of these host cell factors in HIV-1 replication. (C) 2005 Elsevier Inc. All rights reserved.