ISWI proteins participate in the genome‐wide nucleosome distribution in Arabidopsis

Chromatin is a highly organized structure with repetitive nucleosome subunits. Nucleosome distribution patterns, which contain information on epigenetic controls, are dynamically affected by ATP‐dependent chromatin remodeling factors (remodelers). However, whether plants have specific nucleosome distribution patterns and how plant remodelers contribute to the pattern formation are not clear. In this study we used the micrococcal nuclease digestion followed by deep sequencing (MNase‐seq) assay to show the genome‐wide nucleosome pattern in Arabidopsis thaliana. We demonstrated that the nucleosome distribution patterns of Arabidopsis are associated with the gene expression level, and have several specific characteristics that are different from those of animals and yeast. In addition, we found that remodelers in the A. thaliana imitation switch (AtISWI) subfamily are important for the formation of the nucleosome distribution pattern. Double mutations in the AtISWI genes, CHROMATIN REMODELING 11 (CHR11) and CHR17, resulted in the loss of the evenly spaced nucleosome pattern in gene bodies, but did not affect nucleosome density, supporting a previous idea that the primary role of ISWI is to slide nucleosomes in gene bodies for pattern formation.     

News & Notes: Stochastic Nucleosome Positioning in a Yeast Chromatin Region Is Not Dependent on Histone H1

To gain a better understanding of the function of the yeast histone H1, its role in nucleosome positioning was studied. With this objective in mind, we analyzed a chromatin region of the yeast Chromosome (Chr) IX, in which there are two closely packed open reading frames (ORFs), POT1 and YIL161w. This locus shows a regular ladder of 13 stochastically positioned nucleosomes, which is unaffected by the absence of the HHO1 gene. This suggests that histone H1 has no effect on nucleosome positioning in yeast. Received: 30 December 1998 / Accepted: 6 May 1999     

Nucleosome Spacing Generated by ISWI and CHD1 Remodelers Is Constant Regardless of Nucleosome Density

Arrays of regularly spaced nucleosomes are a hallmark of chromatin, but it remains unclear how they are generated. Recent genome-wide studies, in vitro and in vivo, showed constant nucleosome spacing even if the histone concentration was experimentally reduced. This counters the long-held assumption that nucleosome density determines spacing and calls for factors keeping spacing constant regardless of nucleosome density. We call this a clamping activity. Here, we show in a purified system that ISWI- and CHD1-type nucleosome remodelers have a clamping activity such that they not only generate regularly spaced nucleosome arrays but also generate constant spacing regardless of nucleosome density. This points to a functionally attractive nucleosome interaction that could be mediated either directly by nucleosome-nucleosome contacts or indirectly through the remodelers. Mutant Drosophila melanogaster ISWI without the HAND-SANT-SLIDE (HSS) domain had no detectable spacing activity even though it is known to remodel and slide nucleosomes. This suggests that the role of ISWI remodelers in generating constant spacing is not just to mediate nucleosome sliding; they actively contribute to the attractive interaction. Additional factors are necessary to set physiological spacing in absolute terms.     

Genome-wide nucleosome specificity and directionality of chromatin remodelers

How chromatin remodelers cooperate to organize nucleosomes around the start and end of genes is not known. We determined the genome-wide binding of remodeler complexes SWI/SNF, RSC, ISW1a, ISW1b, ISW2, and INO80 to individual nucleosomes in Saccharomyces, and determined their functional contributions to nucleosome positioning through deletion analysis. We applied ultra-high-resolution ChIP-exo mapping to Isw2 to determine its subnucleosomal orientation and organization on a genomic scale. Remodelers interacted with selected nucleosome positions relative to the start and end of genes and produced net directionality in moving nucleosomes either away or toward nucleosome-free regions at the 5' and 3' ends of genes. Isw2 possessed a subnucleosomal organization in accord with biochemical and crystallographic-based models that place its linker binding region within promoters and abutted against Reb1-bound locations. Together, these findings reveal a coordinated position-specific approach taken by remodelers to organize genic nucleosomes into arrays.