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Albany 2019: 20th Conversation - Abstracts

category image Albany 2019
Conversation 20
June 11-15 2019
Adenine Press (2019)

Mismatched base pairs locally distort DNA structure, leading to increased DNA-binding by transcription factor proteins

The local structure of genomic DNA can vary drastically from the ideal B-form double helix, and one cause for structural deformations is the pairing of non-complementary bases (i.e. mis-paired bases, or mismatches). DNA mismatches are frequently formed by spontaneous chemical reactions (such as nucleotide deamination), replication errors, and genetic recombination. Mismatches alter the local DNA structure (Figure 1a), which can affect interactions with DNA-binding proteins, including regulatory transcription factors (TFs). Currently, very little is known about the effects of mismatches on TF binding.

We present Saturation Mismatch Binding Assay (SaMBA), the first assay to characterize the effects of mismatches on TF-DNA binding in high throughput. For genomic sequences of interest, SaMBA generates DNA duplexes containing all possible single-base mismatches, and quantitatively assesses the effects of the mismatches on TF-DNA interactions.
We applied SaMBA to measure binding of 21 TFs (covering 14 structural families) to thousands of mismatched sequences, and mapped the impact of mismatches on these TFs. Remarkably, for all 21 TFs examined, introduction of mismatches at certain positions resulted in significantly increased binding, with some mismatches creating high-affinity binding sites in nonspecific DNA and some converting known binding sites into “super-sites” stronger than any canonical Watson-Crick site.
Structural analyses of mismatches that increase TF binding revealed that these mismatches are oftentimes distorting the naked DNA such that its structure becomes similar to that of bound DNA sites (Fig. 1b), thus explaining the increased binding measured in our assay. Our results reveal that the intrinsic energy cost of deforming the DNA structure is an important, widespread layer of control in protein-DNA recognition. Furthermore, since several recent studies have shown that bound TFs at damaged DNA are likely to be a significant cause for mutations (Reijns, et al. 2015), characterizing the binding preferences of TFs to mismatched DNA is an important step toward understanding the mutational landscape in the genome.

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Figure 1. (a) DNA mismatches have non-canonical base-pair geometries and induce large structural. (b) An example of protein-induced DNA structural deformation that could be mimicked by specific mismatches.

References
    Rohs, R. et al. (2010). Origins of specificity in protein-DNA recognition. Annual review of biochemistry 79, 233-269

    Reijns, M. A. et al. (2015). Lagging-strand replication shapes the mutational landscape of the genome. Nature 518, 502-506.

Ariel Afek 1,2
Honglue Shi 3
Atul Rangadurai 4
Hashim M Al-Hashimi 3,4
Raluca M Gordan 1,2

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Ariel Afek completed his PhD in Chemistry at Ben-Gurion University under the supervision of Dr. David Lukatsky, and he is now a postdoctoral fellow in the laboratory of Dr. Raluca Gordan at Duke University’s Center for Genomic and Computational biology. he will deliver a short oral from the platform.

1Center for Genomic and Computational Biology
2Department of Biostatistics and Bioinformatics
3Department of Chemistry
4Department of Biochemistry
Duke University
Durham, NC 27708, USA

Ph: (984) 3770565
Email: ariel.afek@duke.edu