Population Analysis of Subsaturated 172-12 Nucleosomal Arrays by Atomic Force Microscopy Detects Nonrandom Behavior That Is Favored by Histone Acetylation and Short Repeat Length

Concatameric 5 S rDNA templates reconstituted in vitro into nucleosomal arrays provide very popular chromatin models for many kinds of studies. Here, atomic force microscopy is used to determine the population distributions for one such nucleosomal array, the 172-12, reconstituted to various subsaturated levels with nonacetylated or hyperacetylated HeLa histones. This array is a model for short linker length genomes and transcriptionally active and newly replicated chromatins. The analysis shows that as input histone levels increase, template occupation increases progressively as discrete population distributions. The distributions are random at low (n_av < 4) and high (n_av > 8) loadings but display specific nonrandom features, such as a deficit of molecules with one nucleosome more or less than the peak species in the distribution and enhanced distribution breadths, in the mid-range (n_av = 4-8). Thus, the mid-range of occupation on polynucleosomal arrays may be a special range for chromatin structure and/or assembly. The mid-range nonrandom features are enhanced in distributions from short repeat (172-12) arrays, particularly for unacetylated chromatin, and in distributions from hyperacetylated chromatin, particularly for long repeat (208-12) arrays. Thus, short repeat length and acetylation can affect basic chromatin properties, like population tendencies, in very similar ways and therefore may cause similar changes in chromatin structure. Some possible effects are suggested. The data also indicate that it is thermodynamically more difficult for hyperacetylated nucleosomes to assemble onto the 172-12 templates, a result having implications for in vivo chromatin assembly.