SummaryRegulation of chromatin structure is essential for controlling the access of DNA to factors that require association with specific DNA sequences. The ability to alter chromatin organization in a targeted manner would provide a mechanism for directly manipulating DNA-dependent processes and should provide a means to study direct consequences of chromatin structural changes. Here we describe the development and validation of engineered chromatin remodeling proteins (E-ChRPs) for inducing programmable changes in nucleosome positioning by design. We demonstrate that E-ChRPs function both in vivo and in vitro to specifically reposition target nucleosomes and entire nucleosomal arrays, and possess the ability to evict native DNA-binding proteins through their action. E-ChRPs can be designed with a range of targeting modalities, including the SpyCatcher and dCas9 moieties, resulting in high versatility and enabling diverse future applications. Thus, engineered chromatin remodeling proteins represent a simple and robust means to probe regulation of DNA-dependent processes in different chromatin contexts.
Nucleosome Positioning on Lambda DNA for Single-Molecule Analysis of Chromatin Remodeling - Development of a Versatile Lambda DNA Construct Capable of Reception of any DNA Sequences of Interest