DNA internal motions within nucleosomes during the cell cycle and as a function of ionic strength

We have used 31P NMR spectra to show that DNA internal motions are greatly hindered within oligonucleosomes. The fluctuations seem to be a function of both the cell cycle and the number of nucleosomes interlinked. Namely, the resonance areas, directly related to unbound phosphate, are consistently smaller in M-phase than in S-phase; at the same time, the resonance line width, inversely related to base plane, deoxyribose, and phosphate internal motions, is consistently larger in mononucleosomes than in oligonucleosomes. In all cases, the removal of chromosomal proteins, by a progressive increase of ionic strength up to 2 M NaCl, increases the internal motion, as monitored by a decrease in line width toward that of free DNA. While for both oligo- and mononucleosomes in S-phase the decrease in line width is strictly correlated to a sharp increase in resonance area, in M-phase it is not, with the 31P resonance area rather low even at 2.0 M NaCl extraction. Similarly, while S-phase 31P line widths steadily grow from mono- to oligonucleosomes, in M-phase they do not. Moreover, the increase of the ionic strength to 0.6 M NaCl, as compared to 0.35, 1.2, and 2 M NaCl, displays significant variations on 31P line width and resonance area, independent of the cell cycle phase and the number of nucleosomes interlinked. These observations agree with earlier suggestions on the differential role of the various chromosomal protein subfractions, known to preferentially dissociate at the different ionic strengths in question, in the sealing of mononucleosomes and in the overall stability of polynucleosomes.