Chromatinized Templates Reveal the Requirement for the LEDGF/p75 PWWP Domain During HIV-1 Integration in vitro

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Chromatinized Templates Reveal the Requirement for the LEDGF/p75 PWWP Domain During HIV-1 Integration in vitro

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Title: Chromatinized Templates Reveal the Requirement for the LEDGF/p75 PWWP Domain During HIV-1 Integration in vitro
Author: Botbol, Yaïr; Raghavendra, Nidhanapati K.; Lavigne, Marc; Rahman, Shaila; Engelman, Alan N.

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Citation: Botbol, Yaïr, Nidhanapati K. Raghavendra, Shaila Rahman, Alan Engelman, and Marc Lavigne. 2008. Chromatinized templates reveal the requirement for the LEDGF/p75 PWWP domain during HIV-1 integration in vitro. Nucleic Acids Research 36(4): 1237-1246.
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Abstract: Integration is an essential step in the retroviral lifecycle, and the lentiviral integrase binding protein lens epithelium-derived growth factor (LEDGF)/p75 plays a crucial role during human immunodeficiency virus type 1 (HIV-1) cDNA integration. In vitro, LEDGF/p75 stimulates HIV-1 integrase activity into naked target DNAs. Here, we demonstrate that this chromatin-associated protein also stimulates HIV-1 integration into reconstituted polynucleosome templates. Activation of integration depended on the LEDGF/p75-integrase interaction with either type of template. A differential requirement for the dominant DNA and chromatin-binding elements of LEDGF/p75 was however observed when using naked DNA versus polynucleosomes. With naked DNA, the complete removal of these N-terminal elements was required to abate cofactor function. With polynucleosomes, activation mainly depended on the PWWP domain, and to a lesser extent on nearby AT-hook DNA-binding motifs. GST pull-down assays furthermore revealed a role for the PWWP domain in binding to nucleosomes. These results are completely consistent with recent ex vivo studies that characterized the PWWP and integrase-binding domains of LEDGF/p75 as crucial for restoring HIV-1 infection to LEDGF-depleted cells. Our studies therefore establish novel in vitro conditions, highlighting chromatinized DNA as target acceptor templates, for physiologically relevant studies of LEDGF/p75 in lentiviral cDNA integration.
Published Version: doi:10.1093/nar/gkm1127
Other Sources: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2275106/pdf/
Terms of Use: This article is made available under the terms and conditions applicable to Other Posted Material, as set forth at http://nrs.harvard.edu/urn-3:HUL.InstRepos:dash.current.terms-of-use#LAA
Citable link to this page: http://nrs.harvard.edu/urn-3:HUL.InstRepos:4743732

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