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1.
Incorporation of [125I]IgG into spleen cells was studied in vivo and in vitro. In vivo, the antigen after uptake into the cytoplasm migrated into cell nuclei, where it was bound to chromatin up to the saturation level. One day after immunization the constant level of [125I]IgG was 1.3 X 10(12) molecules per spleen (10(8) cells). The same number of [125I]IgG molecules were bound to chromatin in cell cultures. The uptake of [125I]IgG was competitively inhibited by non-labelled IgG. Binding of [125I]IgG molecules reextracted from cytoplasm and chromatin with specific anti-human IgG serum argues against the uptake of degraded [125I]IgG molecules. [125I]IgG was tightly bound to DNA. Approximately 50 per cent of [125I]IgG was present in the residual chromatin fraction (after removal of 0.35 M and 2 M NaCl-soluble fractions) and 40 per cent was complexed with DNA (after removal of histones and non-histones AP1, AP2, AP3 and AP4). Binding of [125I]IgG by isolated chromatin was inhibited by the cytoplasmic fraction but not by BSA. Binding of [125I]IgG by fractionated chromatin, (chromatins remaining after removal of 0.35M, and 2M NaCl-soluble fractions or histones + non-histones AP1 + AP2 + AP3 + AP4) occurred at a level similar to that observed with native chromatin. The results suggest that interaction of antigen with immunocompetent cells is not restricted to the cell surface but that antigen seems to be taken up into cytoplasm, migrates to the nuclei and is bound to chromatin, probably directly to DNA. The results are discussed in relation to the induction of the immune reaction. 相似文献
2.
Noll M 《Nucleic acids research》1974,1(11):1573-1578
The digestion of chromatin in situ with DNase I reveals, after denaturation, a regular series of single stranded DNA fragments the lengths of which represent multiples of 10 bases. These experiments are compatible with the DNA being on the outside of the chromatin subunit and suggest that the subunit structure itself contains repetitive structural elements. Possible models are discussed. 相似文献
3.
Detlef Doenecke 《Cell》1976,8(1):59-64
Cesium chloride equilibrium density centrifugation shows that treatment of rat liver nuclei with low concetrations of micrococcal nuclease for extremely short periods of time results in the appearance of chromatin fractions of low protein/DNA ratio and even free DNA. The DNA of these chromatin fractions is shorter than the DNA moiety of one chromatin subunit. The amount of high buoyant density material is decreased with increasing digestion time. We conclude that this material belongs to the minor chromatin fraction which is not organized according to the subunit model. 相似文献
4.
DNA organization in nucleosomes 总被引:3,自引:0,他引:3
5.
Ermekova Vanadiya M. Melkonyan Ovsep S. Nanazashvili Michael G. Umansky Samuil R. 《Molecular and cellular biochemistry》1984,62(2):133-139
Summary Rabbit antibodies against rat thymus and liver chromatin are obtained. Antithymus IgG are found to interact only with homologous chromatin, while antiliver IgG interact with both liver and thymus chromatin. After preincubation of antiliver IgG with thymus chromatin the antibodies interact with homologous chromatin only. Thus chromatin of both organs contains tissue-specific proteins. Antiliver and antithymus IgG are used to investigate a distribution of tissue-specific and tissue-non-specific immunogenic proteins in thymocyte nuclei. It is shown that these proteins are practically lacking in nuclear membranes, matrix, nuclear sap, nucleous chromatin and do not participate in attaching DNA to the nuclear matrix.Abbreviations NHP
non-histone chromatin proteins
- IgG
immunoglobulins G
- PMSF
PhMeSO3 相似文献
6.
Differences and similarities in chromatin structure of Neurospora crassa and higher eucaryotes. 总被引:47,自引:0,他引:47
The subunit structure of Neurospora chromatin which contains a full histone complement (Goff, 1976) exhibits both differences and similarities to chromatin of higher eucaryotes. The size of the DNA per subunit is only 170 +/- 5 base pairs, as compared to 200 base pairs in higher eucaryotes. However, the internal structures of the subunits are closely related. They contain 140 base pairs of DNA that are more tightly associated with the histone core and similarly arranged on the outside of the subunit. Hence the difference in structure resides in a shorter linker region of adjacent subunits in Neurospora chromatin. This is supported by a reduced primary cutting site and a lower content of lysines in histone H1. The role of H1 and its relation to the linker region are discussed. 相似文献
7.
Ando K Hagiwara T Beppu M Kikugawa K 《Biochemical and biophysical research communications》2000,275(2):412-417
Human T-lymphoid cell line Jurkat was treated with actinomycin D (ActD) and cycloheximide (CHX). The induction of apoptosis was confirmed by the chromatin condensation and DNA ladder fragmentation. Anti-band 3 IgG, purified from normal human plasma, bound to the ActD- or CHX-treated cells, and the binding was correlated to the degree of apoptosis. Antioxidants, N-acetylcysteine, pilloridine dithiocarbamate, and trolox, inhibited neither induction of DNA fragmentation of ActD-treated cells nor anti-band 3 IgG binding to ActD-treated cells, indicating that formation of the anti-band 3 IgG binding sites on the apoptotic cell surface is caused by nonoxidative mechanism. When Jurkat cells were treated with endo-beta-galactosidase to cleave sialylated poly-N-acetyllactosaminyl saccharide chains from the cell surface before induction of apoptosis, the binding of anti-band 3 IgG was abolished. The results indicate that sialylated poly-N-acetyllactosaminyl saccharide chains on the cell surface are requisite for the binding of anti-band 3 IgG to apoptotic cells. 相似文献
8.
《Cell cycle (Georgetown, Tex.)》2013,12(23):3953-3959
BRG1 is a catalytic subunit of the human SWI/SNF-like BAF chromatin remodeling complexes. Recent findings have shown that inactivation of BRG1 sensitizes mammalian cells to various DNA damaging agents, including ultraviolet (UV) and ionizing radiation. However, it is unclear whether BRG1 facilitates nucleotide excision repair (NER). Here we show that re-expression of BRG1 in cells lacking endogenous BRG1 expression stimulates nucleotide excision repair of UV induced DNA damage. Using a micropore UV radiation technique, we demonstrate that recruitment of the DNA damage recognition protein XPC to sites of UV lesions is significantly disrupted when BRG1 is depleted. Chromatin immunoprecipitation of the endogenous DDB2 protein, which is involved in recruiting XPC to UV-induced CPDs (cyclobutane pyrimidine dimers), shows that elevated levels of BRG1 are associated with DDB2 in chromatin in response to UV radiation. Additionally, we detected slow BRG1 accumulation at sites of UV lesions. Our findings suggest that the chromatin remodeling factor BRG1 is recruited to sites of UV lesions to facilitate NER in human chromatin. 相似文献
9.
《Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms》2020,1863(8):194566
The ATP-dependent BAF chromatin remodeling complex plays a critical role in gene regulation by modulating chromatin architecture, and is frequently mutated in cancer. Indeed, subunits of the BAF complex are found to be mutated in >20% of human tumors. The mechanism by which BAF properly navigates chromatin is not fully understood, but is thought to involve a multivalent network of histone and DNA contacts. We previously identified a composite domain in the BRG1 ATPase subunit that is capable of associating with both histones and DNA in a multivalent manner. Mapping the DNA binding pocket revealed that it contains several cancer mutations. Here, we utilize SELEX-seq to investigate the DNA specificity of this composite domain and NMR spectroscopy and molecular modelling to determine the structural basis of DNA binding. Finally, we demonstrate that cancer mutations in this domain alter the mode of DNA association. 相似文献
10.
Kubota T Hiraga S Yamada K Lamond AI Donaldson AD 《Molecular & cellular proteomics : MCP》2011,10(7):M110.005561
Yeast cells lacking Ctf18, the major subunit of an alternative Replication Factor C complex, have multiple problems with genome stability. To understand the in vivo function of the Ctf18 complex, we analyzed chromatin composition in a ctf18Δ mutant using the quantitative proteomic technique of stable isotope labeling by amino acids in cell culture. Three hundred and seven of the 491 reported chromosomal proteins were quantitated. The most marked abnormalities occurred when cells were challenged with the replication inhibitor hydroxyurea. Compared with wild type, hydroxyurea-treated ctf18Δ cells exhibited increased chromatin association of replisome progression complex components including Cdc45, Ctf4, and GINS complex subunits, the polymerase processivity clamp PCNA and the single-stranded DNA-binding complex RPA. Chromatin composition abnormalities observed in ctf18Δ cells were very similar to those of an mrc1Δ mutant, which is defective in the activating the Rad53 checkpoint kinase in response to DNA replication stress. We found that ctf18Δ cells are also defective in Rad53 activation, revealing that the Ctf18 complex is required for engagement of the DNA replication checkpoint. Inappropriate initiation of replication at late origins, because of loss of the checkpoint, probably causes the elevated level of chromatin-bound replisome proteins in the ctf18Δ mutant. The role of Ctf18 in checkpoint activation is not shared by all Replication Factor C-like complexes, because proteomic analysis revealed that cells lacking Elg1 (the major subunit of a different Replication Factor C-like complex) display a different spectrum of chromatin abnormalities. Identification of Ctf18 as a checkpoint protein highlights the usefulness of chromatin proteomic analysis for understanding the in vivo function of proteins that mediate chromatin transactions. 相似文献
11.
E M Johnson V C Littau V G Allfrey E M Bradbury H R Matthews 《Nucleic acids research》1976,3(12):3313-3329
Nucleosome DNA repeat lengths in Physarum chromatin, determined by nuclease digestion experiments, are shorter than those observed in most mammalian chromatin and longer than those reported for chromatin of certain other lower eukaryotes. After digestion with staphylococcal nuclease for short periods of time an average repeat length of 190 base pairs is measured. After more extensive digestion an average repeat length of 172 base pairs is measured. Upon prolonged digestion DNA is degraded to an average monomer subunit length of 160 base pairs, with only a small amount of DNA found in lengths of 130 base pairs or smaller. Mathematical analysis of the data suggests that the Physarum nucleosome DNA repeat comprises a protected DNA segment of about 159 base pairs with a nuclease-accessible interconnecting segment which ranges from 13 to 31 base pairs. The spacing data are compatible with measurements from electron micrographs of Physarum chromatin. 相似文献
12.
Properties of chromatin subunits from developing trout testis. 总被引:5,自引:0,他引:5
When a sample of trout testis nuclei is digested with micrococcal nuclease, the DNA is cleaved almost entirely to discrete fragments approximately 200 base pairs long and multiples thereof. The same DNA fragments can be obtained when isolated chromatin, as opposed to intact nuclei, is nuclease digested. These DNA fragments can also be found in discrete chromatin "subunits" isolated from nuclease-digested nuclei. Sedimentation through sucrose gradients or velocity sedimentation in an analytical ultracentrifuge separates these chromatin subunits into 11 S (monomer), 16 S (dimer), and 22 S (trimer) etc. species. Subunits can also be fractionated on a Sepharose 2B column equilibrated and run in low salt. High salt (greater than 40 mM NaCl) or divalent cations (congruent to 5 mM) cause subunit precipitation. Chromatin subunits have a protein to DNA ratio of approximately 1.2 and contain all the histones, including the trout-specific histone T. There are, however, no detectable nonhistone chromosomal proteins. Mg-2+ precipitates of the 11 S chromatin monomers, when pelleted, are thin and clear, while oligomer Mg-2+ pellets are thick and white. This could reflect a more symmetrical or ordered packing of 11 S monomers, which are deficient in histone I. This histone may cross-link the larger oligomers, resulting in a disordered Mg-2+ complex. These results are consistent with the subunit model of chromatin structure, based on 200 base pair long regions of DNA associated with histones. These subunits would be separated by nuclease-sensitive DNA spacer regions and cross-linked by histone I. 相似文献
13.
When chromatin was isolated from cells infected with human cytomegalovirus, the virus DNA remained with the chromatin fraction. If deproteinized virus DNA was added to either isolated nuclei or chromatin, the DNA was lost during the chromatin isolation. When isolated chromatin from cytomegalovirus-infected cells was banded in isopycnic metrizamide gradients, a single peak with a density of 1.18 g/cm3 was present. Analysis of this peak in isopycnic neutral CsCl gradients indicated that it contained both human cytomegalovirus and human embryonic lung cell DNAs. When infected nuclei were treated with micrococcal nuclease, 11S subunit particles which cosedimented with cell nucleosomes and contained virus DNA were isolated. 相似文献
14.
Cleavage of DNA in nuclei and chromatin with staphylococcal nuclease. 总被引:53,自引:0,他引:53
R Axel 《Biochemistry》1975,14(13):2921-2925
Treatment of either rat liver chromatin or intact nuclei with the enzyme staphylococcal nuclease results in the conversion of about half of the DNA to acid-soluble oligonucleotides. As previously described, mild digestion of nuclei results in the liberation of a series of nucleoprotein particles containing DNA fragments which are all integral multiples of a unit length DNA 185 base pairs in length. Analysis of the kinetics of appearance of these fragments suggests that at least 85% of the nuclear DNA is involved in the formation of the repeating subunit profile. More extensive digestion of nuclei however results in the generation of a series of eight unique DNA fragments containing 160 to 50 base pairs. The series of smaller molecular weight DNA is virtually identical with the profile obtained upon limit digestion of isolated chromatin. By velocity centrifugation we have obtained highly purified preparations of the monomeric nucleoprotein particle. Digestion of this monomeric subunit results in the solubilization of 46% of the DNA and analysis of the resistant DNA again reveals the set of eight lower molecular weight fragments. These data suggest that the initial site of nuclease cleavage in chromatin resides within the DNA bridging the repeating monomeric subunits. Further attack results in cleavage at a set of sites within the monomer liberating a pattern of smaller DNA fragments which probably represents the points of intimate contact between the histones and DNA. 相似文献
15.
Chromatin remodeling by ATP-dependent molecular machines 总被引:21,自引:0,他引:21
Lusser A Kadonaga JT 《BioEssays : news and reviews in molecular, cellular and developmental biology》2003,25(12):1192-1200
16.
Jin-Xing Zhou Ping Du Zhang-Wei Liu Chao Feng Xue-Wei Cai Xin-Jian He 《The Plant journal : for cell and molecular biology》2021,107(2):467-479
Association of RNA polymerase V (Pol V) with chromatin is a critical step for RNA- directed DNA methylation (RdDM) in plants. Although the methylated DNA-binding proteins SUVH2 and SUVH9 and the chromatin remodeler-containing complex DRD1-DMS3-RDM1 are known to be required for the association of Pol V with chromatin, the molecular mechanisms underlying the association of Pol V with different chromatin environments remain largely unknown. Here we found that SUVH9 interacts with FVE, a homolog of the mammalian retinoblastoma-associated protein, which has been previously identified as a shared subunit of the histone deacetylase complex and the polycomb-type histone H3K27 trimethyltransferase complex. We demonstrated that FVE facilitates the association of Pol V with chromatin and thus contributes to DNA methylation at a substantial subset of RdDM target loci. Compared with FVE-independent RdDM target loci, FVE-dependent RdDM target loci are more abundant in gene-rich chromosome arms than in pericentromeric heterochromatin regions. This study contributes to our understanding of how the association of Pol V with chromatin is regulated in different chromatin environments. 相似文献
17.
Annunziato AT 《Biochimica et biophysica acta》2012,1819(3-4):196-210
It has been over 35years since the acceptance of the "chromatin subunit" hypothesis, and the recognition that nucleosomes are the fundamental repeating units of chromatin fibers. Major subjects of inquiry in the intervening years have included the steps involved in chromatin assembly, and the chaperones that escort histones to DNA. The following commentary offers an historical perspective on inquiries into the processes by which nucleosomes are assembled on replicating and nonreplicating chromatin. This article is part of a Special Issue entitled: Histone chaperones and Chromatin assembly. 相似文献
18.
A comparison of the processes of chromatin digestion in brain and liver nuclei by Ca, Mg-dependent and staphylococcal endonucleases demonstrates a similarity of the subunit composition of chromatin from both tissues and reveals the same type of linked DNA regions. However, a formation of low molecular weight DNP fragments during hydrolysis and the DNA spectra of soluble and insoluble DNP fragments suggest that brain chromatin contains these fragments alongside with the regions, which are specific for this particular tissue, predominate in it and are resistant to staphylococcal and, particularly, to Ca, Mg-dependent endonucleases. This is paralleled with a non-histone protein enrichment of different brain chromatin fractions and an expansion of the electrophoretic monomer band towards the fragment with a greater molecular weight. It may be assumed that brain nucleosomes are characterized by a higher size heterogeneity of linked DNA, part of which are mostly covered by non-histone proteins, and/or are characterized by a greater set variety. 相似文献
19.
Binding of chromatin-modifying activities to phosphorylated histone H2A at DNA damage sites 总被引:1,自引:0,他引:1
Downs JA Allard S Jobin-Robitaille O Javaheri A Auger A Bouchard N Kron SJ Jackson SP Côté J 《Molecular cell》2004,16(6):979-990
Yeast histone H2A is phosphorylated on Ser129 upon DNA damage, an event required for efficient repair. We show that phosphorylation occurs rapidly over a large region around DNA double-strand breaks (DSBs). Histone H4 acetylation is also important for DSB repair, and we found that the NuA4 HAT complex associates specifically with phospho-H2A peptides. A single NuA4 subunit, Arp4, is responsible for the interaction. The NuA4 complex is recruited to a DSB concomitantly with the appearance of H2A P-Ser129 and Arp4 is important for this binding. Arp4 is also a subunit of the Ino80 and Swr1 chromatin remodeling complexes, which also interact with H2A P-Ser129 and are recruited to DSBs. This association again requires Arp4 but also prior NuA4 recruitment and action. Thus, phosphorylation of H2A at DNA damage sites creates a mark recognized by different chromatin modifiers. This interaction leads to stepwise chromatin reconfiguration, allowing efficient DNA repair. 相似文献
20.