John H. Frenster
Laboratory of Molecular Biology
The Rockefeller Institute
New York, NY
Much of the DNA of the interphase calf thymus lymphocyte nucleus is visible as large, condensed, Feulgen-positive masses arrayed peripherally within each nucleus in a manner similar to the heterochromatic inactive X chromosome (Barr body).
Such nuclei were isolated, and were incubated with orotic acid-6-C14 to label newly synthesized RNA. The cytoplasmic fragments and nuclear membranes were removed, and the nuclear ribosomes and neutral proteins were extracted at pH 7.1. The extracted nuclei were suspended in cation-free 0.25 M sucrose and sonicated for 5 seconds at 20,000 kc, releasing the condensed masses, extended fibrils, and particles resembling nucleoli. Each of these were isolated by centrifugation.
The isolated heterochromatin masses were composed of a dense reticulum of 40-A fibrils, and contained the bulk of the DNA (72 per cent), little of the total nuclear RNA (17 per cent), and still less of the newly synthesized RNA (14 per cent). The specific activity of the heterochromatic RNA was one-eighth that of nucleolar RNA.
These data support the concept of heterochromatin as condensed interphase chromosome segments which are inactive in RNA synthesis and in genetic expression (Cooper, Ohno et al, Lyons, Russell, German, Hsu, Grumbach et al).
Supported by an award from the NIH (CA-17857).
1. "Ultrastructural Continuity Between Active and Repressed Chromatin".
2. "Nuclear Polyanions as De-repressors of Synthesis of Ribonucleic Acid".
3. "Mechanisms of Repression and De-repression within Interphase Chromatin".