We have determined histone stoichiometries in nuclei from several sources by a direct chemical method, with the particular aim of quantitating histone H1 and, in chicken erythrocytes, H5, and of distinguishing between one and two molecules per nucleosome. The four histones H3, H4, H2A and H2B are found in equimolar amounts, as expected for the core histone octamer. The molar ratio of H1 in lymphocyte and glial nuclei is 1.0 per octamer, and in liver nuclei from three species 0.8 per octamer. These results suggest that each nucleosome has one H1 molecule; nucleosomes could acquire two molecules of H1 only at the expense of others containing none. The stoichiometry of H5 in chicken erythrocyte nuclei is similar to that of H1 in other nuclei, being about 0.9 molecules per nucleosome; the H1 also present in these nuclei amounts to 0.4 molecules per nucleosome.

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A detailed hydrodynamic study of chicken erythrocyte chromatin as a function of ionic strength, and comparison with results reported earlier for rat liver chromatin [Butler, P. J. G. and Thomas, J. 0. (1980) J . Mol. Bid. 140,529; Thomas, J. 0. and Butler, P. J. G. (1980) J . Mol. Biol. 144,[89][90][91][92][93] shows similar behaviour in the two cases, but quantitative differences indicate a greater stability of the chicken erythrocyte chromatin. The hydrodynamic method may thus provide a general assay for the stability in solution of the higher-order structure of chromatin -a helical coiling into a solenoid.When sedimentation coefficients of nucleosoine oligomers of various lengths from chicken erythrocytes are measured as a function of ionic strength, two main changes in behaviour are observed. The first is between pentamer and hexamer as in rat liver chromatin, suggesting that the solenoid has 5 -6 nucleosomes per turn. The longer repeat length of chicken erythrocyte chromatin (212 base pairs compared with 200 base pairs for rat liver chromatin) is thus accommodated into the solenoid without obvious change of structure.The second change in behaviour is a jump of about 9 % in the sedimentation coefficient at ionic strength 50 mM for nucleosome oligomers above a critical size, which we attribute to the relative instability of highcrorder structure in long oligomers at lower ionic strength. In chicken erythrocyte chromatin this critical size is about 60 nucleosomes, compared with 50 nucleosomes in rat liver chromatin. The increased axial stability in the solenoid that this indicates possibly derives from the erythrocyte-specific histone H5, and would be expected in this chromatin which is virtually inactive in transcription.During erythropoiesis in nucleated erythrocytes, pronounced structural and functional changes occur in the chromatin. The appearance [I] and dephosphorylation [2,3] of the tissue-specific histone H5 is accompanied by a reduction in nuclear activity, and the nucleosome repeat length increases from about 200 to 212 base pairs [1,4]. In the mature erythrocyte transcription is barely detectable [5], and H I has been replaced by [2,6], or supplemented with [I], substantial amounts of H5, which has a higher arginine content than H I but is sufficiently homologous to be regarded as an extreme H I variant [7,8]. It might be expected that such profound modification would be reflected in the physical properties of the chromatin compared with that from a biologically active nucleus such as rat liver.We have previously examined the higher-order structure of rat liver chromatin using measurement of the sedimentation coefficient of a range of nucleosome oligomers to monitor salt-induced conformational changes, and two salt-dependent structural transitions were observed [9,10]. One, occurring between nucleosome pentamers and hexamers, was attributed to the appearance of the simplest element of higherorder structure, a single stable turn of a helix. The second, evident only in oligomers of more than 50 nuc...