Selective displacement of histone H1 from whole HeLa nuclei: effect on chromatin structure in situ as probed by micrococcal nuclease

Gm, Lawson; Rd, Cole

doi:10.1021/bi00578a005

Cited by 51 publications

(32 citation statements)

References 42 publications

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“…Digestion of H1-depleted nuclei with micrococcal nuclease, for instance, shows an increased availability of chromatin to the nucleolytic attack (14,18,22). With fluorescent probes, the role of H1 is not well defined, as to fluorescence response in situ.…”

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confidence: 99%

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Flow cytometric evaluation of DNA stainability with propidium iodide after histone H1 extraction

et al. 1989

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A flow cytometric evaluation of the effect of the histone H1 extraction on DNA stainability with propidium iodide was performed on isolated HeLa nuclei. Selective removal of the lysine-rich protein was attained by using two established techniques involving treatment with 0.7 M NaCl or low pH. DNA stainability was monitored at different dye/DNA-P ratios, varying from low to high saturating concentrations. Depletion of the histone Hlfrom nuclei results in the transition from low to high affinity of a portion of binding sites, as shown by 1) the increase in fluorescence intensity after staining with the dye at low saturating concentrations and 2) the higher value of the fluorescence intensity ratio (F15/F15,J exhibited by H1-depleted nuclei stained with a low (5 pg/ml) vs. a high (50 pg/ml) concentration, as compared with control samples.

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confidence: 99%

“…Extraction was performed with two established techniques (0.7 M NaCl or acid buffer, pH 2.8), that are reported to remove selectively the basic protein, without significant displacement of histones from the nucleosome core (6,18).…”

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Flow cytometric evaluation of DNA stainability with propidium iodide after histone H1 extraction

et al. 1989

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“…The nuclei were depleted of histone Hl by the method in [3]. The nuclei were extracted with a pH 2.2 buffer and selective extraction of histone Hl was thus achieved ( fig.1).…”

Section: Experimental Methods and Resultsmentioning

confidence: 99%

Structural organization of calf thymus chromatin depleted of histone H1 by acidic treatment

Azorı́n

Junca

1981

FEBS Letters

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“…Histones Hi and H5 can be selectively removed from chromatin under a variety of conditions (21,(24)(25)(26)(27)(28)(29), some of which, however, also lead to a redistribution of the histone octamers on the DNA (21,24,30). In the present study, we have developed procedures for the preparation of Hi-and H5-depleted chicken reticulocyte chromatin that do not disturb the characteristic spacing of the residual core protein-DNA complex.…”

Section: Introductionmentioning

confidence: 97%

“…Upon reconstitution of core histones with DNA, nucleosomes are formed (10,(15)(16)(17)(18)(19), but the unique spacing is not regained (19)(20)(21). The resulting irregularity evidently vitiates the reconstitution of the native chromatin structure when the linker histones are introduced (20)(21)(22)(23).Histones Hi and H5 can be selectively removed from chromatin under a variety of conditions (21,(24)(25)(26)(27)(28)(29), some of which, however, also lead to a redistribution of the histone octamers on the DNA (21,24,30). In the present study, we have developed procedures for the preparation of Hi-and H5-depleted chicken reticulocyte chromatin that do not disturb the characteristic spacing of the residual core protein-DNA complex.…”

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Reversible dissociation of linker histone from chromatin with preservation of internucleosomal repeat.

Allan¹,

Staynov²,

Gould³

1980

Proc. Natl. Acad. Sci. U.S.A.

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Procedures are described for the dissociation of histones HI and H5 from chicken reticulocyte chromatin without disruption of the native core histone-DNA complex. The comparative properties of native and depleted chromatin with respect to sedimentation, thermal denaturation, and sensitivity to nuclease digestion have been studied. The changes in these properties resulting from removal of the linker histones are fully reversed when histone H5 is added back to the depleted chromatin.Chromatin contains two functionally distinct groups of histones-namely, the "core" histones (H2A, H2B, H3, and H4) and the linker histones (Hi, largely replaced by H5 in chicken reticulocytes). The core histones are responsible for the primary level of DNA folding, with formation of nucleosomes and a consequent diminution of the contour length by a factor of 7 (1). In the nucleosome, the core histones form an octameric complex around which 145 base pairs of DNA are wound. In native chromatin these nucleosome cores are separated by a uniform length of DNA, characteristic of the tissue, which in chicken reticulocytes is about 70 base pairs (2). Histones Hi and H5 are associated with the linker regions between the nucleosomes (3-6), as well as with the core histone complex (7). Moreover, it is thought that HI and H5 direct the next higher level of DNA folding, apparently helping to bend the chain of nucleosome beads into the form of a solenoid (8) and reducing the contour length by a further factor of 6 (8, 9).The reconstitution of chromatin from separated components provides the most direct approach to the clarification of the structural and functional roles of these constituents in the complex. Thus, in studies of the reconstitution of core histones with DNA, histones H3 and H4 have been identified as the agents of supercoiling of the DNA in the nucleosome (10)(11)(12)(13)(14). Upon reconstitution of core histones with DNA, nucleosomes are formed (10,(15)(16)(17)(18)(19), but the unique spacing is not regained (19)(20)(21). The resulting irregularity evidently vitiates the reconstitution of the native chromatin structure when the linker histones are introduced (20)(21)(22)(23).Histones Hi and H5 can be selectively removed from chromatin under a variety of conditions (21,(24)(25)(26)(27)(28)(29), some of which, however, also lead to a redistribution of the histone octamers on the DNA (21,24,30). In the present study, we have developed procedures for the preparation of Hi-and H5-depleted chicken reticulocyte chromatin that do not disturb the characteristic spacing of the residual core protein-DNA complex. We show that by all the criteria available to us, replacement of histone H5 on the depleted chromatin results in substantial regain of the native chromatin structure. MATERIAL AND METHODSPreparation of "Native" Chromatin. Nuclei were prepared from chicken reticulocytes as described (31). The nuclei were washed twice with 0.2 M sucrose/1 mM CaCl2/5 mM Tris-HCl, pH 7.5/80 mM NaCl/0.2 mM phenylmethylsulfonyl fluoride (PMSF), and res...

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Selective displacement of histone H1 from whole HeLa nuclei: effect on chromatin structure in situ as probed by micrococcal nuclease

Cited by 51 publications

References 42 publications

Flow cytometric evaluation of DNA stainability with propidium iodide after histone H1 extraction

Flow cytometric evaluation of DNA stainability with propidium iodide after histone H1 extraction

Structural organization of calf thymus chromatin depleted of histone H1 by acidic treatment

Reversible dissociation of linker histone from chromatin with preservation of internucleosomal repeat.

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