NAP1 Modulates Binding of Linker Histone H1 to Chromatin and Induces an Extended Chromatin Fiber Conformation

Abstract

NAP1 (nucleosome assembly protein 1) is a histone chaperone that has been described to bind predominantly to the histone H2A⅐H2B dimer in the cell during shuttling of histones into the nucleus, nucleosome assembly/remodeling, and transcription. Here it was examined how NAP1 interacts with chromatin fibers isolated from HeLa cells. NAP1 induced a reversible change toward an extended fiber conformation as demonstrated by sedimentation velocity ultracentrifugation experiments. This transition was due to the removal of the linker histone H1. The H2A⅐H2B dimer remained stably bound to the native fiber fragments and to fibers devoid of linker histone H1. This was in contrast to mononucleosome substrates, which displayed a NAP1-induced removal of a single H2A⅐H2B dimer from the core particle. The effect of NAP1 on the chromatin fiber structure was examined by scanning/atomic force microscopy. A quantitative image analysis of ϳ36,000 nucleosomes revealed an increase of the average internucleosomal distance from 22.3 ؎ 0.4 to 27.6 ؎ 0.6 nm, whereas the overall fiber structure was preserved. This change reflects the disintegration of the chromatosome due to binding of H1 to NAP1 as chromatin fibers stripped from H1 showed an average nucleosome distance of 27.4 ؎ 0.8 nm. The findings suggest a possible role of NAP1 in chromatin remodeling processes involved in transcription and replication by modulating the local linker histone content.The dynamic organization of chromatin in the eukaryotic nucleus is tightly connected to transcription (1-3). Transcribed chromatin is thought to be in a more open conformation with a higher accessibility to DNase I or micrococcal nuclease (4 -7). On the other hand, highly compacted and dense chromatin regions, referred to as heterochromatin, are often transcriptionally inactive and contain a reduced number of genes (3,7,8). Several factors have been identified that promote a transition between a transcriptionally active and a more dense/inactive chromatin conformation. These include linker histones (9 -11), DNA methylation (2, 12), histone modifications (13-15), and the incorporation of histone variants (16 -19). The dynamic nature of chromatin manifests itself by continuous rearrangements of its three-dimensional structure. These changes involve the activity of histone chaperones that mediate the ordered deposition and the removal and exchange of histones (20 -23). The central carrier of the histone H2A⅐H2B dimer in the cell is NAP1 (nucleosome assembly protein 1) (24). NAP1 is involved in the transport of the histone H2A⅐H2B dimer from the cytoplasm to the nucleus and the deposition of histones onto the DNA as described in several reviews (20 -23). NAP1 and other histone chaperones stimulate the binding of transcription factors to chromatin templates (25, 26). In yeast, loss of NAP1 leads to an altered gene expression of about 10% of the genome (27), and several lines of evidence suggest that NAP1 has activities related to transcription. First, it has been shown that NAP1 is present in com...

Chromatin reprogramming during the somatic-to-reproductive cell fate transition in plants

“…A similar process is likely to be in place in plants, whereas a CDK2 type of activity could possibly contribute to H1 phosphorylation in wheat male meiocytes (Greer et al, 2012). In addition, H1 binding is modulated by the histone chaperone NUCLEOSOME ASSEMBLY PROTEIN 1 (NAP1) (Kepert et al, 2005). Arabidopsis plants lacking NAP1;1-3 activity [triple mutant (Liu et al, 2009)] or the activity of the NAP1-related proteins NRP1 and NRP2 [double mutant (Zhu et al, 2006)] show normal eviction of both H1.1 and H1.2 in the MMC and reincorporation at meiosis (supplementary material Fig.…”

Heparin exerts a dual effect on murine lupus nephritis by enhancing enzymatic chromatin degradation and preventing chromatin binding in glomerular membranes

“…For example, nucleosome assembly protein 1, nucleoplasmin, and Hsp90 induce changes in chromatin conformation that are assumed to increase accessibility for proteinases and nucleases (20)(21)(22)(23)(24)(25). It is not known whether such molecules are tolerated in vivo in doses necessary for a therapeutic effect.…”

Large Multimeric Assemblies of Nucleosome Assembly Protein and Histones Revealed by Small-angle X-ray Scattering and Electron Microscopy

“…The Chz1 chaperone shows very high specificity, binding preferentially to the variant H2A.Z/H2B dimer over the canonical H2A/H2B dimer (10). In contrast, chaperones such as NAP1 can bind all of the core histones as well as the linker histone, although with varying affinities (11)(12)(13). Following the discovery that NAP1 and ACF can facilitate the assembly of chromatin, organizing nucleosomes in regular spaced arrays (14), NAP1 has become a vital constituent for the generation of in vitro chromatin systems (15) in a range of downstream studies (16 -20).…”

“…In more physiological buffers, further oligomerization of yNAP was observed using size exclusion chromatography (SEC) (12) and AUC (25). Several groups reported that the NAP1 dimer can bind an H3/H4 or H2A/H2B dimer or an H1 monomer (11,12,25,26). Alternative stoichiometries have been suggested, including one NAP1 monomer binding one histone dimer (27,28), and therefore, the question of the NAP1-histone stoichiometry remains controversial.…”