Biological and Biomedical Science
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Alan Engelman

Division of Pathology
Harvard Medical School

Department of Cancer Immunology and AIDS
Dana Farber Cancer Institute
JFB 815
44 Binney Street
Boston, MA 02115
Tel: (617) 632-4361
Fax: (617) 632-3113
Email: alan_engelman@dfci.harvard.edu
5 postdoctoral fellows, 2 graduate students

Research in the Engelman laboratory focuses on the integration step in the retroviral life cycle. In order to replicate, retroviruses must integrate the cDNA product of reverse transcription into a host cell chromosome. In addition to the viral integrase (IN) protein, host cell proteins play essential roles in integration. IN functions in the context of preintegration complexes (PICs) that contain the cDNA, host cell, and other viral proteins.

PICs isolated from infected cells catalyze cDNA integration in vitro. Although work with purified IN proteins has revealed details of IN structure and function, little is known about the organization and structure of PICs. We previously defined the HIV-1 intasome as the nucleoprotein complex within PICs that mediates integration. Current projects focus on defining the stoichiometries and folding topologies of viral and cell proteins in purified PICs. In vivo roles of host cell factors in HIV-1 integration are evaluated using RNAi and gene knockout strategies.

One cell protein that directly binds to HIV-1 IN, lens epithelium-derived growth factor (LEDGF), greatly stimulates recombinant IN catalytic activity in vitro. LEDGF was knocked out in mice to generate a clean ex vivo model system. HIV-1 titer was reduced approximately 80-fold on LEDGF knockout cells, revealing a crucial role for the host factor in infection. Different retroviruses differentially target genomic features like genes and promoter regions during integration - lentiviruses like HIV-1 preferentially target active genes fairly equally along their lengths. The frequency and distribution of HIV-1 integration were severely disrupted in LEDGF knockout cells: global integration was reduced approximately tenfold, with significant redistributions of residual proviruses to gene-poor regions and transcriptional start sites, mimicking the natural distributions of other retroviruses. The results highlighted a critical role for LEDGF at the genomic targeting step of lentiviral integration.

 

References:

  • Cherepanov P, Sun Z-Y J, Rahman S, Maertens G, Wagner G, Engelman A. Solution structure of the HIV-1 integrase-binding domain in LEDGF/p75. Nat Struct Mol Biol 2005;12:526-32.
  • Cherepanov P, Ambrosio ALB, Rahman S, Ellenberger T, Engelman A. From the Cover: Structural basis for the recognition between HIV-1 integrase and transcriptional coactivator p75. Proc Natl Acad Sci USA 2005;102:17308-13.
  • Daelemans D, Lu R, De Clercq E, Engelman A. Characterization of a replication-competent, integrase defective human immunodeficiency virus (HIV)/simian virus 40 chimera as a powerful tool for the discovery and validation of HIV integrase inhibitors. J Virol 2007;81:4381-5.
  • Shun MC, Raghavendra NK, Vandegraaff N, Daigle JE, Hughes S, Kellam P, Cherepanov P, Engelman A. LEDGF/p75 functions downstream from preintegration complex formation to effect gene-specific HIV-1 integration. Genes Dev 2007;21:1767-78.