Stable co-occupancy of transcription factors and histones at the HIV-1 enhancer

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Abstract
To investigate mechanisms yielding DNase I‐hypersensitive sites (DHSs) at gene regulatory regions, we have initiated a biochemical analysis of transcription factor binding and nucleosome remodeling with a region of the human immunodeficiency virus 1 (HIV‐1) 5′ long terminal repeat (LTR) that harbors constitutive DHSs in vivo. In vitro reconstitution of an HIV‐1 5′ LTR fragment into nucleosome core particles demonstrates that Sp1, NF‐κB1, LEF‐1, ETS‐1 and USF can gain access to their binding sites in HIV‐1 nucleosomal DNA. The factor‐bound mononucleosomes resist histone displacement from the DNA by the chromatin remodeling activity, SW1–SNF, or the histone chaperone, nucleoplasmin, suggesting that the binding of these factors to nucleosomal HIV‐1 sequences forms a stable complex that includes the underlying histones. However, when the HIV‐1 5′ LTR fragment is incorporated into a nucleosomal array, Sp1 and NF‐κB1 binding produce regions of enhanced DNase I sensitivity specifically at the HIV‐1 nucleosome. These regions resemble the observed in vivo DHSs, yet the HIV‐1 nucleosome remains intact even in the presence of nucleoplasmin. Thus, the constitutive DHSs identified at the HIV‐1 enhancer in native chromatin may reflect the presence of a ternary complex composed of transcriptional activators, histones and DNA.