Nucleosome positioning : Impact on transcription factors access
Sammanfattning: Nucleosome Positioning: Impact on Transcription Factors Access Qiao Li Laboratory of Molecular Genetics, Department of Cell and Molecular Biology Medical Nobel Institute, Karolinska Institute, S-171 77 Stockholm, SwedenIn the eukaryotic cell nucleus, DNA is organized into nucleosomes. Some of theregulatory regions are integrated into sequence-specifically positionednucleosomes. What impact nucleosome positioning has on the accessibility of theregulatory sequences, and how transcription factors gain access to their bindingsites within nucleosomes, are fundamental to the understanding of the eukaryoticgene expression. These issues were addressed in a purified system by binding twodifferent transcription factors to various nucleosomes displaying their cognatesites in a defined rotational and translational position. Glucocorticoid receptor (GR) is able to recognize a glucocorticoid responseelement (GRE) that is located at the nucleosome dyad and faces away from thehistone octamer. The binding affinity is only 2-fold lower than that of a free GRE.The contacts of GR with this nucleosomal GRE are identical to that with the freeGRE as revealed by dimethylsulfate methylation analysis. Conversely, a GRE at asimilar translational position but facing towards the histone octamer isinaccessible for GR recognition. However, the influence of rotational positioningcan be compromised by translational positioning of the GRE in a nucleosome. GRis capable of binding to an inward-facing GRE near the nucleosome boundaryalbeit with a 6-fold decreased affinity relative to an outward-facing GRE. Theinteraction of the GR homodimer with the GRE is via the two consecutive majorgrooves that face toward the histone octamer, and the formation of the ternarycomplex is accompanied by a local alteration of histone-DNA contacts. The helical path of the DNA on the histone octamer is anisotropic and thisis reflected in the decreased GR binding to an outward-facing GRE located 20-bpfrom the nucleosome dyad. However, the reduced GR affinity can be restored byintroducing a 5-bp A/T segment immediately after the GRE at the major groovecompression. The introduction of an A/T segment at a major groove compressionis detrimental to the nucleosomal formation, which alleviates the constraintsimposed by the histone-DNA interaction at this translational position. The affinity of GR to two GREs organized in one nucleosome depends onthe spatial proximity of the two GREs within the nucleosome rather than thelinear distance along the DNA axis. The facilitated binding of GR is observed forthe two GREs juxtapositioned by one turn of nucleosomal DNA (80-bp). This is insharp contrast to the cooperative GR binding previously reported for free GRE. In contrast to GR, NF1 could not bind to its recognition sequences within anucleosome regardless of the rotational and translational position. Taken together, these data demonstrate that both rotational andtranslational positioning of the regulatory sequences within nucleosomes modulatetranscription factor access. However, the effect of translational positioning can beoverridden by the sequences which flank the regulatory element. Furthermore, theability of a transcription factor to recognize its cognate element in a nucleosome isan inherent property that differs among different transcription factors.Key words: glucocorticoid receptor, transcription factor, gene regulation, nucleosome,nucleosome positioning, DNA-bending sequenceISBN 91-628-1917-8 Stockholm 1996
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