Abstract
Enhancers and promoters often contain multiple binding sites for the same transcription factor, suggesting that homotypic clustering of binding sites may serve a role in transcription regulation. Here we show that clustering of binding sites for the transcription termination factor TTF-I downstream of the pre-rRNA coding region specifies transcription termination, increases the efficiency of transcription initiation and affects the three-dimensional structure of rRNA genes. On chromatin templates, but not on free rDNA, clustered binding sites promote cooperative binding of TTF-I, loading TTF-I to the downstream terminators before it binds to the rDNA promoter. Interaction of TTF-I with target sites upstream and downstream of the rDNA transcription unit connects these distal DNA elements by forming a chromatin loop between the rDNA promoter and the terminators. The results imply that clustered binding sites increase the binding affinity of transcription factors in chromatin, thus influencing the timing and strength of DNA-dependent processes.
Highlights
An intriguing question for understanding protein-DNA recognition is how low-abundant transcription factors recognize their target sites in genomic DNA [1,2]
We focused on the murine rDNA terminator, which comprises 10 termination sites (T1–T10) spaced by 18–123 bp, preventing the accommodation of nucleosomes in between the TTF-I binding sites
In electrophoretic mobility shift assays (EMSAs) recombinant TTF-I bound with comparable affinity to all terminators assayed
Summary
An intriguing question for understanding protein-DNA recognition is how low-abundant transcription factors recognize their target sites in genomic DNA [1,2]. Empirical studies revealed that regulatory regions, such as enhancers and promoters, comprise modular units of a few hundred base pairs that harbour multiple binding sites for the same transcription factor. Such ‘homotypic clustering sites’ (HTCs) have been identified in 2% of the human genome, being enriched at promoters and enhancers [3]. Studies in mammalian cells have shown that clustering of binding sites facilitate the cooperative binding of nuclear receptors to their target sites in vivo, suggesting that HCTs coordinate the recruitment of transcription initiation factors [8,9,10]. Cooperative binding could arise through indirect effects, e.g. by changing the accessibility of neighbouring binding sites in chromatin [11]
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