Mutual arrangement of histone H1 molecules in extended chromatin

Abstract

Mutual arrangement of histone H1 molecules in chromatin extended in low salt—EDTA buffer and additionally in the presence of urea was studied by means of reversible cross‐linking combined with chymotryptic digestion. In the chromatins tested, the chymotryptic halves of H1 were cross‐linked in all possible combinations; i.e., C—C, C—N and N—N. The results imply that the mutual arrangement of H1 histones is determined by the structure of extended nucleosomal chain, rather than chromatin superstructure.

The arrangement of H5 molecules in extended and condensed chicken erythrocyte chromatin.

“…Whether this has anything to do with the use of a different cross-linking reagent or a somewhat higher pH (7.9 rather than 7.5), or with the use of chromatin from a different source is not clear; the latter seems unlikely since we obtained the same results for HI in rat liver chromatin (not shown) as described here for H5 in chicken erythrocyte chromatin. The pronounced formation of C-C cross-links in 8 M urea (Nikolaev et al, 1983) is not easily reconciled with our conclusions that C/C cross-linking is promoted by chromatin condensation; however, the relevance of structures in 8 M urea, which denatures the histones but tolerates electrostatic interactions with DNA, is unclear. Ring and Cole (1983) analysed HI-HI cross-links formed in nuclei with two cross-linking reagents of different lengths, and although cross-linking between amino-terminal regions was not detected, strong cross-linking was found between the carboxyterminal regions and between the amino-terminal and carboxyterminal portions of neighbouring HI molecules, in agreement with our results and those of Nikolaev et al (1981).…”

“…Cross-linking was found between all combinations of the N-and C-terminal 'halves' of HI generated by chymotryptic cleavage, both in nuclei and in extended chromatin, and moreover in the presence of 8 M urea; however, short oligonucleosomes (e.g., dinucleosomes) were not examined, nor was the contribution of intramolecular NG/C cross-linking directly determined. Although the relative amounts of C/C and NG/C (or in their terminology N/C) cross-linking is hard to assess from the studies of Nikolaev et al (1981Nikolaev et al ( , 1983 because of poor resolution in the relevant region of the twodimensional gels, there appears to be considerably less NG/C cross-linking, relative to C/C, than we observe, especially in extended chromatin (Figure IA of Nikolaev et al, 1983). Whether this has anything to do with the use of a different cross-linking reagent or a somewhat higher pH (7.9 rather than 7.5), or with the use of chromatin from a different source is not clear; the latter seems unlikely since we obtained the same results for HI in rat liver chromatin (not shown) as described here for H5 in chicken erythrocyte chromatin.…”

“…the HI-helix of Thoma et al, 1979), although such a mechanism is both attractive and plausible. Previous studies of the relative orientation of H] molecules in chromatin While this work was in progress, Nikolaev et al (1981Nikolaev et al ( , 1983 reported studies of the arrangement of H I molecules in calf thymus chromatin using the reagent methyl 4-mercaptobutyrimidate in a two-stage cross-linking procedure. Cross-linking was found between all combinations of the N-and C-terminal 'halves' of HI generated by chymotryptic cleavage, both in nuclei and in extended chromatin, and moreover in the presence of 8 M urea; however, short oligonucleosomes (e.g., dinucleosomes) were not examined, nor was the contribution of intramolecular NG/C cross-linking directly determined.…”

NaCl-Induced Chromatin Condensation. Application of Static Light Scattering at 90° and Stopped Flow Technique

“…Total chromatin was isolated by endonuclease digestion (24). The nuclear pellet was suspended in a 5 mM Na-cacodilate buffer, pH 6.0, containing 0.25 mM sucrose, 4 mM MgC1 2 , 1 mM CaC1 2 , 1 mM PMSF (25) and incubated for 20 min.…”