Minor groove geometry determines DNA exclusion from nucleosomes (929.1)

Abstract

Like all eukaryotes, the meters of DNA in each of our cells must be compacted many thousands of times to fit inside the nucleus. The first level of compaction, spooling of DNA onto a nucleosome, involves the tight wrapping of DNA around eight histone proteins. Sharply bending DNA to wrap it into nucleosomes comes at an energetic cost related to the stiffness of the DNA. Nucleosomes prefer to form on DNA sequences that have a 10 bp periodicity between AA/AT/TA steps, with CC/CG/GC steps exactly out of phase. The sites of the AA/AT/TA and CC/CG/GC steps exactly correspond to locations of the highest DNA kinking, suggesting that these sequences represent anisotropically bendable sequences. Yet, inclusion of multiple AA steps, in the form of a poly(dA:dT) elements, results in exclusion of the DNA from nucleosomes in vivo. We seek to understand what unique chemical features of these poly‐dA elements are necessary for their exclusion from nucleosomes. Poly‐dA elements are known to adopt a unique structure; characterized crystallographically as a narrowed minor groove and by CD as a double peaked spectrum. Using CD spectroscopy and non‐natural nucleotides, we show that correct minor groove geometry at the base pair level is necessary for formation of the unique poly‐dA structure. Using in vitro competitive nucleosome reconstitution assays, we have further shown that non‐natural DNA sequences that adopt the unique poly‐dA structure are excluded from nucleosomes. Since poly‐dA like sequences are excluded from nucleosomes, this suggests that the unique structure formed by these sequences is energetically stiffer than generic sequence DNA.