Differential affinity of mammalian histone H1 somatic subtypes for DNA and chromatin

Abstract

BackgroundHistone H1 is involved in the formation and maintenance of chromatin higher order structure. H1 has multiple isoforms; the subtypes differ in timing of expression, extent of phosphorylation and turnover rate. In vertebrates, the amino acid substitution rates differ among subtypes by almost one order of magnitude, suggesting that each subtype might have acquired a unique function. We have devised a competitive assay to estimate the relative binding affinities of histone H1 mammalian somatic subtypes H1a-e and H1° for long chromatin fragments (30–35 nucleosomes) in physiological salt (0.14 M NaCl) at constant stoichiometry.ResultsThe H1 complement of native chromatin was perturbed by adding an additional amount of one of the subtypes. A certain amount of SAR (scaffold-associated region) DNA was present in the mixture to avoid precipitation of chromatin by excess H1. SAR DNA also provided a set of reference relative affinities, which were needed to estimate the relative affinities of the subtypes for chromatin from the distribution of the subtypes between the SAR and the chromatin. The amounts of chromatin, SAR and additional H1 were adjusted so as to keep the stoichiometry of perturbed chromatin similar to that of native chromatin. H1 molecules freely exchanged between the chromatin and SAR binding sites. In conditions of free exchange, H1a was the subtype of lowest affinity, H1b and H1c had intermediate affinities and H1d, H1e and H1° the highest affinities. Subtype affinities for chromatin differed by up to 19-fold. The relative affinities of the subtypes for chromatin were equivalent to those estimated for a SAR DNA fragment and a pUC19 fragment of similar length. Avian H5 had an affinity ~12-fold higher than H1e for both DNA and chromatin.ConclusionH1 subtypes freely exchange in vitro between chromatin binding sites in physiological salt (0.14 M NaCl). The large differences in relative affinity of the H1 subtypes for chromatin suggest that differential affinity could be functionally relevant and thus contribute to the functional differentiation of the subtypes. The conservation of the relative affinities for SAR and non-SAR DNA, in spite of a strong preference for SAR sequences, indicates that differential affinity alone cannot be responsible for the heterogeneous distribution of some subtypes in cell nuclei.

Translocation of histone H1 subtypes between chromatin and cytoplasm during mitosis in normal human fibroblasts

“…Using GFP-labeled proteins, Th'ng et al (12) found histone H1.2 to have lower affinity for chromatin than histone H1.5, whereas histone H1.3 had intermediate binding affinity. In another study, where unlabeled H1 subtypes were used in chromatin competition experiments, H1.2 and H1.5 had lower affinity for chromatin than H1.3 (13). Most likely, differential H1 affinity for chromatin is a result of a combination of phosphorylation and inherent affinity, leading to various distributions of H1 subtypes between the chromosomes and the cytoplasm during the cell cycle.…”

“…Although there seems to be a noteworthy redundancy among the histone H1 subtypes (10), evolutionary data indicate that they posses different functional roles (11). They differ in affinity for chromatin (12,13), ability to affect gene regulation (14), and are found to localize to different parts of the nuclei (12). Thus, GFP-labeled H1.1, H1.2, and H1.3 were mainly found in euchromatin, whereas H1.4 and H1.5 were more common in heterochromatin (12).…”

N- and C-terminal Domains Determine Differential Nucleosomal Binding Geometry and Affinity of Linker Histone Isotypes H10 and H1c

“…H1 isotypes isolated from rat brains were classified as possessing high (H1e, H1 0 , and H1d), intermediate (H1c and H1b), and low binding affinities (H1a) (38). No appreciable differences in the binding affinities between human H1 isotypes H1-0 and H1.2 (the paralog of mouse H1c) were found in in vitro assays (39), although these isotypes were reported to have different binding characteristics by in vivo experiments (40).…”

Caracterización de la estructura secundaria de subtipos de la histona H1 por dicroísmo circular

“…Las histonas H1 difieren en estabilidad evolutiva (10) y en las tasas relativas de síntesis y degradación, en células en división y en células quiescentes, propiedades que permiten cambios en sus proporciones relativas durante el desarrollo y la maduración de los tejidos. Las histonas H1 presentan diferencias de afinidad relativa por el ADN y por la cromatina; la H1a es el subtipo con más baja afinidad, las H1b y H1c tienen afinidades intermedias y las H1d, H1e y H1° tienen las afinidades más altas (11).…”