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Published October 11, 2013 | Supplemental Material + Published
Journal Article Open

Poly(dA:dT)-Rich DNAs Are Highly Flexible in the Context of DNA Looping

Abstract

Large-scale DNA deformation is ubiquitous in transcriptional regulation in prokaryotes and eukaryotes alike. Though much is known about how transcription factors and constellations of binding sites dictate where and how gene regulation will occur, less is known about the role played by the intervening DNA. In this work we explore the effect of sequence flexibility on transcription factor-mediated DNA looping, by drawing on sequences identified in nucleosome formation and ligase-mediated cyclization assays as being especially favorable for or resistant to large deformations. We examine a poly(dA:dT)-rich, nucleosome-repelling sequence that is often thought to belong to a class of highly inflexible DNAs; two strong nucleosome positioning sequences that share a set of particular sequence features common to nucleosome-preferring DNAs; and a CG-rich sequence representative of high G+C-content genomic regions that correlate with high nucleosome occupancy in vivo. To measure the flexibility of these sequences in the context of DNA looping, we combine the in vitro single-molecule tethered particle motion assay, a canonical looping protein, and a statistical mechanical model that allows us to quantitatively relate the looping probability to the looping free energy. We show that, in contrast to the case of nucleosome occupancy, G+C content does not positively correlate with looping probability, and that despite sharing sequence features that are thought to determine nucleosome affinity, the two strong nucleosome positioning sequences behave markedly dissimilarly in the context of looping. Most surprisingly, the poly(dA:dT)-rich DNA that is often characterized as highly inflexible in fact exhibits one of the highest propensities for looping that we have measured. These results argue for a need to revisit our understanding of the mechanical properties of DNA in a way that will provide a basis for understanding DNA deformation over the entire range of biologically relevant scenarios that are impacted by DNA deformability.

Additional Information

© 2013 Johnson et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Received July 3, 2013; Accepted August 19, 2013; Published October 11, 2013. Editor: James P. Brody. The authors gratefully acknowledge the support of the National Institutes of Health (www.nih.gov, grant numbers DP1 OD000217A [Director's Pioneer Award], R01 GM085286, R01 GM085286-01S1, and 1 U54 CA143869 [Northwestern PSOC center]). SJ gratefully acknowledges the support of a graduate fellowship from the National Science Foundation (www.nsf.gov), and RP the support of the Fondation Pierre Gilles de Gennes (http://www.fondation-pgg.org/). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. We are indebted to the late Jon Widom for the inspiration of this project and for his guidance, mentorship and friendship over many years. We thank Chao Liu, David Wu, David Van Valen, Hernan Garcia, Martin Linde´n, Mattias Rydenfelt, Yun Mou, Tsui-Fen Chou, Eugene Lee, Matthew Raab, Daniel Grilley, Niv Antonovsky, Lior Zelcbuch, Matthew Moore, Ron Milo, Eran Segal, and the Phillips, Mayo, Pierce and Elowitz labs for insightful discussions, equipment and technical help; and Winston Warman at Transgenomic, Inc. (Omaha, NE, USA) and Jin Li at Laragen, Inc (Culver City, CA, USA) for special help with sequencing the poly(dA:dT)-rich DNAs. Author Contributions: Conceived and designed the experiments: SJ YJC RP. Performed the experiments: SJ YJC. Analyzed the data: SJ YJC. Contributed reagents/ materials/analysis tools: SJ YJC. Wrote the paper: SJ YJC RP. Competing Interests: The authors have declared that no competing interests exist. These authors contributed equally to this work.

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Published - journal.pone.0075799.pdf

Supplemental Material - journal.pone.0075799.s001.pdf

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Additional details

Created:
August 19, 2023
Modified:
October 25, 2023