We are working to understand the inherent DNA accessibility within nucleosomes and chromatin. Numerous protein complexes must bind DNA binding sites that are located within the nucleosome to regulate transcription. We are investigating the impact of thermally excited nucleosome structural transitions on DNA-protein binding within the nucleosome. We are focusing on DNA binding transcription factors (TFs). However, our results should apply to any DNA binding protein.
We recently made a surprising discovery with single molecule Fluorescence Resonance Energy Transfer (smFRET) experiments that detect individual TF binding and dissociation events at the TF’s target site within a single nucleosome. We find that nucleosomes increase protein dissociation from their DNA target sites by three orders of magnitude! Before this, it was assumed that nucleosomes only controlled DNA-protein binding by regulating the probability that a nucleosome unwrapped. Our results show that nucleosomes regulate TF occupancy by controlling both the binding and dissociation rates. We recently published this observation in Nucleic Acids Research. We are now working to understand how nucleosomes increase TF dissociation in thermal equilibrium without an energy source such as ATP. We are pursuing two hypotheses: (i) nucleosome rewrapping competes with partially bound states of the TF (Fig.1) and (ii) that the nucleosome directly introduces changes that increase TF dissociation.
Fig. 1 Dimeric model of how nucleosomes increase TF dissociation
In addition, we are investigating other properties of the nucleosome including (i) the impact of the TF binding site position within the nucleosomes on TF occupancy, binding rate and dissociation rate, (ii) the DNA accessibility within hexasomes and tetrasomes (partially formed nucleosomes) and (iii) the DNA accessibility at TF binding sites between nucleosomes within chromatin.