Identification and functional analysis of a second RBF-2 binding site within the HIV-1 promoter

Virology. 2011 Sep 15;418(1):57-66. doi: 10.1016/j.virol.2011.07.002. Epub 2011 Aug 2.

Abstract

Transcription from the HIV-1 long terminal repeat (LTR) is mediated by numerous host transcription factors. In this study we characterized an E-box motif (RBE1) within the core promoter that was previously implicated in both transcriptional activation and repression. We show that RBE1 is a binding site for the RBF-2 transcription factor complex (USF1, USF2, and TFII-I), previously shown to bind an upstream viral element, RBE3. The RBE1 and RBE3 elements formed complexes of identical mobility and protein constituents in gel shift assays, both with Jurkat T-cell nuclear extracts and recombinant USF/TFII-I. Furthermore, both elements are regulators of HIV-1 expression; mutations in LTR-luciferase reporters and in HIV-1 molecular clones resulted in decreased transcription, virion production, and proviral expression in infected cells. Collectively, our data indicate that RBE1 is a bona fide RBF-2 binding site and that the RBE1 and RBE3 elements are necessary for mediating proper transcription from the HIV-1 LTR.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding Sites
  • Electrophoretic Mobility Shift Assay
  • Gene Expression
  • Genes, Reporter
  • HIV-1 / genetics*
  • Host-Pathogen Interactions*
  • Humans
  • Jurkat Cells
  • Luciferases / genetics
  • Luciferases / metabolism
  • Models, Biological
  • Promoter Regions, Genetic*
  • Protein Binding
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Transcription Factors, TFII / genetics
  • Transcription Factors, TFII / metabolism*
  • Upstream Stimulatory Factors / genetics
  • Upstream Stimulatory Factors / metabolism*

Substances

  • GTF2I protein, human
  • Recombinant Proteins
  • Transcription Factors, TFII
  • USF1 protein, human
  • USF2 protein, human
  • Upstream Stimulatory Factors
  • Luciferases