Assembly and Disassembly of Nucleosome Core Particles Containing Histone Variants by Human Nucleosome Assembly Protein I

Graduate School of Comprehensive Human Sciences and Institute of Basic Medical Sciences, University of Tsukuba, 1-1-1 Tennohdai, Tsukuba 305-8575, Japan.
Molecular and Cellular Biology (Impact Factor: 4.78). 01/2006; 25(23):10639-51. DOI: 10.1128/MCB.25.23.10639-10651.2005
Source: PubMed


Histone variants play important roles in the maintenance and regulation of the chromatin structure. In order to characterize
the biochemical properties of the chromatin structure containing histone variants, we investigated the dynamic status of nucleosome
core particles (NCPs) that were assembled with recombinant histones. We found that in the presence of nucleosome assembly
protein I (NAP-I), a histone chaperone, H2A-Barr body deficient (H2A.Bbd) confers the most flexible nucleosome structure among
the mammalian histone H2A variants known thus far. NAP-I mediated the efficient assembly and disassembly of the H2A.Bbd-H2B
dimers from NCPs. This reaction was accomplished more efficiently when the NCPs contained H3.3, a histone H3 variant known
to be localized in the active chromatin, than when the NCPs contained the canonical H3. These observations indicate that the
histone variants H2A.Bbd and H3.3 are involved in the formation and maintenance of the active chromatin structure. We also
observed that acidic histone binding proteins, TAF-I/SET and B23.1, demonstrated dimer assembly and disassembly activity,
but the efficiency of their activity was considerably lower than that of NAP-I. Thus, both the acidic nature of NAP-I and
its other functional structure(s) may be essential to mediate the assembly and disassembly of the dimers in NCPs.

Download full-text


Available from: Shin-ichi Tate
  • Source
    • "A rabbit polyclonal antibody against MV C protein was generated as follows: Haemmagulutinin (HA)-tagged recombinant C protein between amino acid positions 1e93 was obtained using E. coli expression system [14]. Recombinant C protein was purified using Ni-chelation resins according to the method by the manufacturer (Novagen). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Measles virus (MV) C protein has been known a multifunctional protein involved in anti-IFN response, viral RNA synthesis, and so on. Recent studies have clarified that double-stranded RNA (dsRNA) is derived from viral genomic RNA (vRNA), and the amount of dsRNA was increased in an MV lacking C protein (MV(C-))-infected cells, suggesting that C protein blocks viral RNA synthesis. However, detailed roles of C protein in viral RNA synthesis remain unknown. Here, we have confirmed through time course experiments using Vero/hSLAM cells that as reported previously, the amount of mRNA is increased in MV(C-)-infected cells at 36 hours post infection (hpi). In contrast, we found that the transcription level is lower in MV(C-)-infected cells than wild-type MV-infected cells in early phases of infection. Immunoprecipitation assays were performed to find an interactor(s) of C protein, revealed that C protein interacts with N protein in the absence of vRNA and P protein. RNA immunoprecipitation (RIP) assays showed that the interaction between N protein and vRNA was increased in MV(C-)-infected cells. These results suggest that in early phases of infection C protein facilitates viral transcription to control the formation of nascent nucleocapsid composed of N protein and vRNA. Copyright © 2015. Published by Elsevier Inc.
    Preview · Article · Jun 2015 · Biochemical and Biophysical Research Communications
  • Source
    • "H2A.B does not form a stable histone octamer without DNA28, and nucleosomal H2A.B exchanges rapidly in nuclei25. Consistently, H2A.B confers a more flexible nucleosome structure, as compared to the other mammalian histone H2A variants, H2A.X, H2A.Z, macroH2A, and canonical H2A38. Intriguingly, these characteristics are common to human histone H3T14, which is also highly expressed in testis394041. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Human histone H2A.B (formerly H2A.Bbd), a non-allelic H2A variant, exchanges rapidly as compared to canonical H2A, and preferentially associates with actively transcribed genes. We found that H2A.B transiently accumulated at DNA replication and repair foci in living cells. To explore the biochemical function of H2A.B, we performed nucleosome reconstitution analyses using various lengths of DNA. Two types of H2A.B nucleosomes, octasome and hexasome, were formed with 116, 124, or 130 base pairs (bp) of DNA, and only the octasome was formed with 136 or 146 bp DNA. In contrast, only hexasome formation was observed by canonical H2A with 116 or 124 bp DNA. A small-angle X-ray scattering analysis revealed that the H2A.B octasome is more extended, due to the flexible detachment of the DNA regions at the entry/exit sites from the histone surface. These results suggested that H2A.B rapidly and transiently forms nucleosomes with short DNA segments during chromatin reorganization.
    Full-text · Article · Dec 2013 · Scientific Reports
  • Source
    • "Nucleosome core particle (NCP) assembly assays were performed using 147-bp DNA fragment containing 5S rRNA gene sequence and core histones prepared as described previously (24). For DNA-binding assays shown in Figure 4, 196-bp DNA fragment was mixed with recombinant NPM2 proteins in 20 mM Tris–HCl pH 7.9, 100 mM NaCl, 10% Glycerol, 0.1 mg/ml of BSA. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Sperm chromatin remodeling after oocyte entry is the essential step that initiates embryogenesis. This reaction involves the removal of sperm-specific basic proteins and chromatin assembly with histones. In mammals, three nucleoplasmin/nucleophosmin (NPM) family proteins-NPM1, NPM2 and NPM3-expressed in oocytes are presumed to cooperatively regulate sperm chromatin remodeling. We characterized the sperm chromatin decondensation and nucleosome assembly activities of three human NPM proteins. NPM1 and NPM2 mediated nucleosome assembly independently of other NPM proteins, whereas the function of NPM3 was largely dependent on formation of a complex with NPM1. Maximal sperm chromatin remodeling activity of NPM2 required the inhibition of its non-specific nucleic acid-binding activity by phosphorylation. Furthermore, the oligomer formation with NPM1 elicited NPM3 nucleosome assembly and sperm chromatin decondensation activity. NPM3 also suppressed the RNA-binding activity of NPM1, which enhanced the nucleoplasm-nucleolus shuttling of NPM1 in somatic cell nuclei. Our results proposed a novel mechanism whereby three NPM proteins cooperatively regulate chromatin disassembly and assembly in the early embryo and in somatic cells.
    Full-text · Article · Feb 2012 · Nucleic Acids Research
Show more