LEDGF/p75 in retroviral DNA integration: from structure to function to potential applications

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Akin to other retroviruses, HIV must insert a DNA replica of its genome into a human chromosome. Retroviral DNA integration defines a step of no return in the life cycle of the virus and the infected cell, the cell becoming a permanent carrier of the provirus (i.e. integrated copy of the viral genome). This feature contributes to the virus? persistency and later progression to AIDS. However, it also makes HIV and other retroviruses natural gene delivery vehicles for permanent repair of genetic disorders. Unfortunately, uncontrolled integration of modified retroviruses or ?vectors? can result in disruption or untuning of important genes and lead to cancer. Thus, insertion of a retroviral vector in proximity of the LMO2 gene is thought to be the cause of leukemias that affected three out of eleven children that participated in an X-SCID (X chromosome-linked severe combined immunodeficiency, also known as ?Bubble Boy Disease?) gene therapy trial. Although the essential enzymatic steps associated with integration are orchestrated by virally-encoded enzyme integrase, many cellular factors are recruited by the virus to aid the intracellular transport, protection and finally homing of the viral integration machinery onto a suitable region of the host cell chromosomal DNA. LEDGF is a human chromosome-associated protein that has been extensively implicated in integration of HIV and related non-primate viruses. LEDGF binds integrase protein with high affinity and is capable of increasing its activity as much as 150-fold. In cells, LEDGF can protect HIV integrase from degradation and tether the viral protein to chromosomal DNA. All available data strongly indicate that LEDGF is directly involved in targeting HIV integration machinery into actively expressed genes. Recent biochemical and structural studies conducted by the applicant explained some key aspects of the LEDGF-IN recognition. The current research project is aimed to explain mechanism of LEDGF-mediated activation of integration and its selectivity towards a specific subset of retroviruses (lentiviridae). To this end we will try to obtain crystals and determine molecular structures of various integrase-LEDGF complexes. The study will further our understanding of the role that LEDGF plays in retroviral replication. We will also attempt to tailor LEDGF protein to re-target HIV integration machinery to a predefined spot of the human genome, a step towards creating safer gene therapy vectors.