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Characterizing interactions of HIV-1 integrase with viral DNA and the cellular cofactor LEDGF

McKee, Christopher J.
http://rave.ohiolink.edu/etdc/view?acc_num=osu1274979614

Abstract Details

2010, Doctor of Philosophy, Ohio State University, Molecular, Cellular, and Developmental Biology.
A hallmark of retroviral replication is the permanent integration of the viral genome into the host cell genome. This integration is mediated by the viral enzyme integrase (IN) which is bound to the ends of the viral DNA in the context of a large nucleoprotein complex known as the pre-integration complex (PIC). Using the two catalytic activities of IN, the viral DNA ends are first processed then used for a strand transfer reaction that simultaneously breaks the host DNA backbone and ligates the viral genome into it. For HIV-1, the site of integration into the host genome is non-random and occurs most often in active transcription units. This bias is likely explained by the recruitment of pre-integration complexes to chromatin via interactions between IN and the cellular protein Lens Epithelium-Derived Growth Factor (LEDGF). Failure of either enzymatic reaction or failure of the PIC to engage chromatin is a replicative dead end for the virus. Integrase has long been considered an attractive therapeutic target due to its essential role in replication and its lack of a cellular counterpart. The first-in-class HIV IN inhibitors block integration by binding specifically to the IN-viral DNA complex, displacing the viral DNA ends from the active site and effectively preventing strand transfer. The specificity of this inhibition led to the rapid emergence of HIV-1 phenotypes resistant to all available strand transfer inhibitors and highlighted the need for new classes of IN inhibitor. The development of these new inhibitors will be driven by an improved understanding of IN structure and of the sequence of pre-integration events. In spite of tremendous efforts, no high-resolution structural data is available for the full-length HIV-1 IN. The following text describes my efforts to characterize three critical molecular interactions that determine the fate of HIV-1 integration: IN with viral DNA, IN with the cellular cofactor LEDGF, and LEDGF with chromatin. Using innovative mass spectrometric footprinting techniques I have mapped and validated biologically essential contacts at IN-viral DNA, IN-IN, and IN-LEDGF interfaces. These experiments provided structural details that revealed previously undescribed changes in integrase conformation and subunit dynamics upon binding viral DNA and LEDGF, respectively. Finally, I have explored epigenetic modifications in chromatin that are recognized by LEDGF. Specific binding of LEDGF to regions featuring these modifications could explain, at least in part, the targeting of active transcription units for HIV-1 integration in infected cells.
Mamuka Kvaratskhelia, PhD (Advisor)
Karin Musier-Forsyth, PhD (Committee Member)
Thomas Schmittgen, PhD (Committee Member)
Mary Jo Burkhard, PhD (Committee Member)
Biochemistry; Cellular Biology; Virology
HIV-1 integrase; LEDGF; mass spectrometric footprinting; histone post-translational modifications

Recommended Citations

Citations

  • McKee, C. J. (2010). Characterizing interactions of HIV-1 integrase with viral DNA and the cellular cofactor LEDGF [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1274979614

    APA Style (7th edition)

  • McKee, Christopher. Characterizing interactions of HIV-1 integrase with viral DNA and the cellular cofactor LEDGF. 2010. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1274979614.

    MLA Style (8th edition)

  • McKee, Christopher. "Characterizing interactions of HIV-1 integrase with viral DNA and the cellular cofactor LEDGF." Doctoral dissertation, Ohio State University, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=osu1274979614

    Chicago Manual of Style (17th edition)

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This open access ETD is published by The Ohio State University and OhioLINK.
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