The structure of chromatin containing DNA complementary to 19 S and 26 S ribosomal RNA in active and inactive stages of Physarum polycephalum

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Abstract

The subunit composition of Physarum chromatin containing DNA complementary to 19 S and 26 S ribosomal RNA was studied by hybridization of 32P-labeled RNA to the DNA of chromatin subunits generated by staphyloeoccal nuclease digestion and separated on sucrose gradients. DNA sequences coding for ribosomal RNAs in growing microplasmodia appear to be more rapidly degraded by micrococcal nuclease than is bulk DNA. The percentage of total nuclear DNA hybridizing to 19 S + 26 S RNA declines progressively with increasing digestion time from 0.18% in undigested nuclei to 0.03% in nuclei treated for 10 min with low levels of nuclease. Similarly, the content of 19 S and 26 S rDNA sequences in nucleosomes released from nuclei declines from 0.07% after 2 min digestion to 0.01% after 10 min digestion. The 19 S and 26 S rDNA sequences are recovered primarily in chromatin subunits containing 140 to 160 nucleotide-pairs of DNA. The distribution of 19 S and 26 S rDNA sequences in nuclease-shielded particles differs from that of the bulk of chromosomal DNA. At all digestion times, the highest content of 19 S and 26 S rDNA sequences is seen in a class of slowly sedimenting particles (peak A) most of which contain a monomer length of DNA but differ from the usual monomer subunit with regard to sedimentation properties and content of nuclear proteins. Electron microscopic observations reveal that isolated peak A particles are in an extended strand configuration as opposed to the compact beaded conformation of isolated monomers. The rapid digestion of 19 S and 26 S rDNA sequences is not due to preferential digestion of newly replicated rDNA. The absence of peak A and slower kinetics of digestion of 19 S and 26 S rDNA sequences in nuclei from microsclerotia, a transcriptionally inactive stage of Physarum, suggest a greater resistance to staphyloeoccal nuclease action of nucleosomes containing inactive rDNA sequences.

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