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Component alpha DNA is a homogeneous, highly repetitive fraction that comprises nearly a quarter of the African green monkey (Cercopithecus aethiops) genome. By restriction enzyme analysis, it has a repeat periodicity of 176 +/- 4 nucleotide base pairs, corresponding closely with the length of DNA contained within a nucleosome. The sequence is organized into large blocks of constitutive heterochromatin. A method is described here for the isolation of intact polynucleosomal arrays containing only component alpha sequences. Isolated monkey nuclei are treated with EcoRI, which releases only component alpha nucleosomal arrays; the arrays are then fractionated and purified by sedimentation in sucrose gradients. The method permits a compositional analysis of the proteins associated with a constitutively repressed, heterochromatic sequence. The major differences in the proteins associated with component alpha nucleosomes that distinguish them from the bulk DNA nucleosomes are a decrease in the content of the H1 histones in the component alpha nucleosomes and a concomitant increase in the amount of certain nonhistone proteins. The specific observations are: (i) In the component alpha nucleosomes, 65-70% of the proteins were nonhistone proteins; this contrasts with the value, 40%, for nonhistone proteins associated with nucleosomes containing bulk DNA. (ii) The amount of H1 histone in chromatin containing predominantly bulk DNA was about 13.7%. However, the H1 histone was depleted and possibly absent in component alpha oligonucleosomes. (iii) Coincident with the decrease in the H1 histones and in the same molecular weight range (24,000-43,000), there appeared five minor nonhistone proteins. The minor, low-molecular-weight, nonhistone proteins were not detected in chromatin containing bulk DNA but they represented nearly 12% of the protein in component alpha nucleosomes. The resistance to salt extraction (0.6-2.0 M NaCl) indicates that the low-molecular-weight nonhistone proteins are tenaciously bound to the component alpha nucleosomes. In addition, a class of high-molecular-weight (&gt;100,000) nonhistone proteins was enriched 5- or 6-fold in component alpha oligonucleosomes. The relative amounts of the nucleosome core histones were not changed.

proceedings of the national academy of sciences of the united states of america

Subunit structure of chromatin and the organization of eukaryotic highly repetitive DNA: Nucleosomal proteins associated with a highly repetitive mammalian DNA