Abstract
Eukaryotic genomes are organized dynamically through the repositioning of nucleosomes. Isw2 is an enzyme that has been previously defined as a genome-wide, nonspecific nucleosome spacing factor. Here, we show that Isw2 instead acts as an obligately targeted nucleosome remodeler in vivo through physical interactions with sequence-specific factors. We demonstrate that Isw2-recruiting factors use small and previously uncharacterized epitopes, which direct Isw2 activity through highly conserved acidic residues in the Isw2 accessory protein Itc1. This interaction orients Isw2 on target nucleosomes, allowing for precise nucleosome positioning at targeted loci. Finally, we show that these critical acidic residues have been lost in the Drosophila lineage, potentially explaining the inconsistently characterized function of Isw2-like proteins. Altogether, these data suggest an 'interacting barrier model,' where Isw2 interacts with a sequence-specific factor to accurately and reproducibly position a single, targeted nucleosome to define the precise border of phased chromatin arrays.
Highlights
Chromatin consists of the nucleic acids and proteins that make up the functional genome of all eukaryotic organisms
We further demonstrate that conserved attributes of this helical domain are observed in the cell cycle regulator Swi[6], which we have identified as a new Isw2- recruitment adapter protein that allows for specific nucleosome positioning at Mbp1/Swi[6] (MBF) and Swi4/Swi[6] (SBF) targets
Close inspection of Isw2-targeted pre-initiation complex sites (PICs) suggests that Isw[2] can only organize a single PIC-proximal nucleosome, while subsequent nucleosomes become more poorly phased as the distance from the initially positioned nucleosome increases (Figure 1A, Figure 1-figure supplement 2)
Summary
Chromatin consists of the nucleic acids and proteins that make up the functional genome of all eukaryotic organisms. A widely accepted model is that ChRPs pack nucleosome arrays against a non-interacting barrier, such as an unrelated DNA-binding protein or another nucleosome[16,20,21] In this way, general regulatory factors (GRFs) could establish chromatin landscapes with differing nucleosome arrays in response to changes in the cellular environment. While this model provides a good explanation for how phased nucleosome arrays can be established throughout the genome by a combination of DNA binding factors and nonspecific chromatin remodeling factors, the fundamental assumptions of the barrier model have not been thoroughly tested It has been shown through genetic and recent biochemical experiments that members of the ISWI family of ChRPs functionally interact with transcription factors in vivo[16,23,24,25,26]. We show that these residues, which are essential for directional, sequence-specific remodeling, were lost in the evolution of the Drosophila lineage, where extensive biochemical, genetic and genomic characterization has been performed on the ITC1 ortholog ACF
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