The eukaryotic genome is packaged by wrapping ~147 bp units of DNA around histone octamers to form chains of nucleosomes. The packaging of the DNA within a nucleosome reduces access of the DNA to most transcription factors and polymerases. In between nucleosomes, there are regions of more accessible DNA, called linker regions that vary from a few base pairs to several hundred base pairs.1
Thus within any given cell type, the precise partitioning of the genome into nucleosome-bound and nucleosome-free DNA regions can have large consequences on gene regulation and help define a particular cellular state.2
Recent studies suggest that the genome plays a large role in encoding its own packaging through differences in affinity of the underlying sequence for the histone octamer 3
(). Genome wide nucleosome mapping studies have found that ~80% of all yeast nucleosomes adopt specific positions. 1, 4, 5
The nucleosome positions are often predicted from the primary DNA sequence based on the ability of the DNA sequence to bend in a manner required for nucleosome formation.6, 7
Other work has mapped local changes in nucleosome positions in promoter regions across the yeast genome upon environmental stress such as heat shock.2
These changes in nucleosome positions alter the accessibility of promoter regions to transcription factors. Since such changes in nucleosome positions are generally thought to accompany cellular differentiation and adaptation, it is of interest to understand to what extent and how DNA sequence influences transitions between different chromatin states.
Model: Chromatin Remodeling Enzymes Help Nucleosomes Equilibrate on the Genomic Thermodynamic Landscape
While nucleosomes can assemble on any sequence, under physiological conditions, most nucleosomes do not move on their own,8
but remain kinetically trapped in their locations. In vivo
, transitions between chromatin states are catalyzed by ATP-dependent chromatin remodeling machines that rapidly reorganize histone-DNA contacts.9–12
Thus, understanding whether DNA sequence affects the activity of remodeling enzymes is a key aspect of understanding how DNA sequence might influence transitions between different chromatin states.
One way in which DNA sequence can regulate remodeling activity is by affecting the stability of a nucleosome. This model draws on previous observations that nucleosomes assembled on strong positioning sequences are thermally repositioned more slowly than nucleosomes on weak positioning sequences.8, 13, 14
If the rate-limiting transition state for remodeling primarily involves loosened histone-DNA contacts, then remodeling enzymes, like heat, will also move unstable nucleosomes more readily than stable nucleosomes. In such a mechanism, the main function of a remodeling enzyme would be to catalyze rapid equilibration between different nucleosome positions (). This model predicts that remodeling enzymes will move nucleosomes occupying high affinity DNA sequences more slowly than nucleosomes occupying lower affinity DNA sequences. Another recently proposed possibility is that certain DNA sequences adopt topological properties within nucleosomes that inhibit binding by chromatin remodeling complexes and thereby help stabilize nucleosomes in specific locations.15
While some biochemical studies have shown that ATP-dependent remodeling enzymes can move nucleosomes away from strong nucleosome positioning sequences16–19
other studies suggest that nucleosome positioning properties of the DNA do play an important role in determining the locations of remodeled nucleosomes.14, 20
These observations have suggested a third model in which DNA sequence does not directly affect the activity of remodeling enzymes but instead more locally regulates the outcome of remodeling by influencing the collapse of remodeling intermediates into a particular set of stable nucleosomal positions.14
To distinguish amongst the three models, we separately measured the effects of nucleosome stability and DNA structure on nucleosome remodeling rates. We tested the activities of two major classes of ATP-dependent remodeling complexes, the ISWI class and the SWI/SNF class on nucleosomes with different stabilities. These two classes of complexes are highly abundant, act broadly at a number of different loci, and generate substantially different products. We therefore reasoned that results with these two classes would lead to generalizable principles of how DNA sequence influences ATP-dependent nucleosome reorganization. We chose yeast RSC as a representative member of the SWI/SNF class and human ACF as a representative member of the ISWI class. Surprisingly, we find that the rates of nucleosome movement by these complexes are insensitive to 1000-fold changes in histone-DNA affinity. These and other results presented here support the model that DNA sequence regulates ATP-dependent remodeling by locally influencing the final nucleosome positions adopted by remodeling intermediates.