Native nucleosome-positioning elements for the investigation of nucleosome repositioning.
Nucleosome repositioning is essential for establishing nucleosome-depleted regions (NDRs) to initiate transcription. This process has been extensively studied using structural, biochemical, and single-molecule approaches, which require homogenously positioned nucleosomes. This is often achieved using the Widom 601 sequence, a highly efficient nucleosome positioning element (NPE) selected for its unusually strong binding to the H3-H4 histone tetramer. Due to the artificial nature of 601, a library of native NPEs is needed to explore the role of DNA sequence in nucleosome repositioning. Here, we characterize the position distributions and nucleosome formation free energy for a set of yeast native nucleosomes (YNNs) from Saccharomyces cerevisiae. We show these native NPEs can be used in biochemical studies of nucleosome repositioning by transcription factors (TFs) and the chromatin remodeler Chd1. TFs could directly reposition a fraction of nucleosomes containing native NPEs, but not 601-containing nucleosomes. In contrast, partial unwrapping was similar for 601 and native NPE sequences. The rate of ATP-dependent remodeling by Chd1 was within the range of the fast and slow directions of the 601 nucleosomes. This set of native NPEs provides an alternative to the 601 NPE that can be used for probing the repositioning of nucleosomes that contain native DNA sequences. Overall design: MNase-seq to determine nucleosome positioning of YNN candidates or 601 with and without yeast transcription factors.