Identification of two DNA-binding sites on the globular domain of histone H5.

Goytisolo, Fermín A.; Gerchman, Sue Ellen; Yu, Xinyang; Rees, Christine; Graziano, Vito; Ramakrishnan, V.; Thomas, Jean O.

doi:10.1002/j.1460-2075.1996.tb00708.x

Cited by 143 publications

(151 citation statements)

References 50 publications

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“…Molecules A and B of the dimer differ in the conformation of the winged loop. Molecule A has been typically used for modeling 12,16,18,19). The winged loop of molecule A extends away from the molecule because of hydrophobic intermolecular contacts with the wing of another molecule A in the crystal lattice (12).…”

Section: Resultsmentioning

confidence: 99%

“…DOT more diverse, but all had the DNA backbone clamped between Lys-40 and Lys-97. Usually only two GH5 DNA-binding sites are discussed (12,19,22): a primary site contributed by helix H3 and Lys-85 and a secondary site containing Lys-40, Arg-42, Lys-52, and Arg-94. These analyses were based on molecule A of the GH5 structure in which the perturbed wing conformation causes the Lys-85 side chain to extend toward helix H3.…”

Section: Resultsmentioning

confidence: 99%

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Complex of linker histone H5 with the nucleosome and its implications for chromatin packing

Фан

Roberts

2006

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

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Linker histones are essential for chromatin filament formation, and they play key roles in the regulation of gene expression. Despite the determination of structures of the nucleosome and linker histones, the location of the linker histone on the nucleosome is still a matter of debate. Here we show by computational docking that the globular domain of linker histone variant H5 (GH5) has three distinct DNA-binding sites, through which GH5 contacts the DNA at the nucleosome dyad and the linker DNA strands entering and exiting the nucleosome. Our results explain the extensive mutagenesis and crosslinking data showing that side chains spread throughout the GH5 surface interact with nucleosomal DNA. The nucleosome DNA contacts positively charged side chains that are conserved within the linker histone family, indicating that our model extends to linker histone-nucleosome interactions in general. Furthermore, our model provides a structural mechanism for formation of a dinucleosome complex specific to the linker histone H5, explaining its efficiency in chromatin compaction and transcription regulation. Thus, this work provides a basis for understanding how structural differences within the linker histone family result in functional differences, which in turn are important for gene regulation.computational docking ͉ DNA-protein interactions ͉ DOT ͉ linker DNA ͉ winged-helix protein T he interaction of the linker histone with the nucleosome is an ongoing controversial issue (1-3). Histone H5 has been the focus of linker-histone-related studies in recent years. H5 consists of a central globular domain (GH5) that is essential for nucleosome binding and is flanked by basic N-and C-terminal tails (4). Binding of either H5 or GH5 to nucleosomes protects an additional 20 bp of linker DNA from micrococcal nuclease digestion (5). Early studies based on micrococcal nuclease digestion and DNase I footprinting proposed a symmetrical model in which GH5 contacts the dyad of the nucleosome and both the entering and exiting DNA duplexes (also called DNA arms) (6, 7). A ''bridging'' model was later proposed based on experiments that mapped the binding site of GH5 on mixedsequence chicken chromatosomes by conjugating a crosslinking reagent to specific Ser-to-Cys substitutions (8). In this model, GH5 forms a bridge between one DNA arm and the dyad. A radically different ''off-axis'' model was developed from studies on a DNA fragment containing the Xenopus borealis somatic 5S RNA gene (9). Based on this model, GH5 is positioned Ϸ65 bp away from the dyad and is bound inside the DNA superhelix. The bridging and off-axis models imply there may be two equivalent linker histone-binding sites per nucleosome (1), but a 1:1 ratio of linker histone and nucleosome is observed (10, 11).There have been multiple proposals for defining the DNAbinding sites on the linker histone. GH5 has a winged-helix fold (12), consisting of a three-helix bundle in which helices H2 and H3 are part of a helix-turn-helix motif that is followed by a ␤-hairpin, termed the ...

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Complex of linker histone H5 with the nucleosome and its implications for chromatin packing

Фан

Roberts

2006

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

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“…Previous studies have clearly established that the interaction of H1 with chromatin is dynamic and the overall affinity is governed by a set of interactions mediated by the globular domain (49,50) and the positively charged COOH-terminal tail (51,52). The folded globular domain of H1 binds preferentially to crossovers of duplex strands in superhelical DNA (40), contains two binding sites for DNA (49), and protects an additional 20 bp from micrococcal nuclease digestion (6, 41). The highly basic COOHterminal domain of H1 adopts secondary ␣-helical structure upon binding to DNA (51) and has been identified as the primary determinant of H1 binding to chromatin in vitro (51,53) and in vivo (52).…”

Section: Discussionmentioning

confidence: 99%

Parathymosin Affects the Binding of Linker Histone H1 to Nucleosomes and Remodels Chromatin Structure

Martic

Karetsou

Kefala

et al. 2005

Journal of Biological Chemistry

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Linker histone H1 is the major factor that stabilizes higher order chromatin structure and modulates the action of chromatin-remodeling enzymes. We have previously shown that parathymosin, an acidic, nuclear protein binds to histone H1 in vitro and in vivo. Confocal laser scanning microscopy reveals a nuclear punctuate staining of the endogenous protein in interphase cells, which is excluded from dense heterochromatic regions. Using an in vitro chromatin reconstitution system under physiological conditions, we show here that parathymosin (ParaT) inhibits the binding of H1 to chromatin in a dose-dependent manner. Consistent with these findings, H1-containing chromatin assembled in the presence of ParaT has reduced nucleosome spacing. These observations suggest that interaction of the two proteins might result in a conformational change of H1. Fluorescence spectroscopy and circular dichroism-based measurements on mixtures of H1 and ParaT confirm this hypothesis. Human sperm nuclei challenged with ParaT become highly decondensed, whereas overexpression of green fluorescent protein-or FLAG-tagged protein in HeLa cells induces global chromatin decondensation and increases the accessibility of chromatin to micrococcal nuclease digestion. Our data suggest a role of parathymosin in the remodeling of higher order chromatin structure through modulation of H1 interaction with nucleosomes and point to its involvement in chromatindependent functions.Eukaryotic DNA is packaged into chromatin (1), a structural framework that plays a key role in the regulation of the transcriptional status of genes and genetic loci (2-4). The basic level of chromatin organization is the nucleosome that consists of DNA wrapped around an octamer of histones (5). Linker histones (e.g. H1 and H5) seal the entry/exit of DNA and stabilize the folding of 10-nm nucleosomal arrays into higher order chromatin structures (6, 7). Binding of histone H1 to nucleosomal arrays affects the spacing of nucleosomes (8), restricts their mobility (9), and reduces the transient exposure of DNA on the nucleosome surface (10). Consistent with its structural role as a major determinant of chromatin folding, histone H1 was found to inhibit the action of enzymes that enhance the accessibility of DNA, such as histone acetyltransferases and ATP-dependent remodeling complexes. For example, changes in linker histone stoichiometry modulate the levels of core histone acetylation (11) and regulate the activity of all classes of ATP-dependent remodeling enzymes (12, 13). Collectively, these findings support the notion that H1 "locks down" chromatin regions and acts as a general inhibitor of transcription (14, 15). However, genetic studies have shown that, rather than being global repressors of transcription, linker histones affect the expression of specific genes (16). Multiple isoforms of linker histone H1 exist in mammalian somatic cells (17) and play differential roles in gene expression (18). Recently, two independent studies highlight the important role of H1 subtypes in dev...

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“…These studies found that HNF3 binding is required for nucleosome positioning over the albumin enhancer (31,32), consistent with the hepatic-specific expression pattern observed for albumin in vivo. HNF3, which structurally resembles linker histone H5 (34), can bind in place of a linker histone molecule to a degenerate HNF3-binding site on the side of a core nucleosome, but lacks the basic amino acids present in linker histones required to mediate compaction of nucleosomal DNA (35). Our in vitro analysis of HNF3 provides the first demonstration that HNF3-directed structural changes functionally mediate derepression of transcription on a chromatin template.…”

Section: Discussionmentioning

confidence: 99%

“…HNF3␣ is of particular interest as it has been shown to position nucleosomes within the enhancer of the liver-specific albumin gene (31,32). A role for HNF3 in organizing chromatin is further supported by the three-dimensional structural similarity of the winged helix conformation to the globular domain of linker histones (33,34); functional conservation of this domain was confirmed by studies demonstrating the nucleosome-binding properties of HNF3 (35). However, the mechanism by which HNF3 transactivates hepatic-specific genes remains unclear.…”

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confidence: 98%

Hepatocyte Nuclear Factor 3 Relieves Chromatin-mediated Repression of the α-Fetoprotein Gene

Crowe¹,

Sang²,

Li³

et al. 1999

Journal of Biological Chemistry

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The ␣-fetoprotein gene (AFP) is tightly regulated at the tissue-specific level, with expression confined to endoderm-derived cells. We have reconstituted AFP transcription on chromatin-assembled DNA templates in vitro. Our studies show that chromatin assembly is essential for hepatic-specific expression of the AFP gene. While nucleosome-free AFP DNA is robustly transcribed in vitro by both cervical (HeLa) and hepatocellular (HepG2) carcinoma extracts, the general transcription factors and transactivators present in HeLa extract cannot relieve chromatin-mediated repression of AFP. In contrast, preincubation with either HepG2 extract or HeLa extract supplemented with recombinant hepatocyte nuclear factor 3 ␣ (HNF3␣), a hepatic-enriched factor expressed very early during liver development, is sufficient to confer transcriptional activation on a chromatin-repressed AFP template. Transient transfection studies illustrate that HNF3␣ can activate AFP expression in a non-liver cellular environment, confirming a pivotal role for HNF3␣ in establishing hepatic-specific gene expression. Restriction enzyme accessibility assays reveal that HNF3␣ promotes the assembly of an open chromatin structure at the AFP promoter. Combined, these functional and structural data suggest that chromatin assembly establishes a barrier to block inappropriate expression of AFP in non-hepatic tissues and that tissue-specific factors, such as HNF3␣, are required to alleviate the chromatin-mediated repression.

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Identification of two DNA-binding sites on the globular domain of histone H5.

Cited by 143 publications

References 50 publications

Complex of linker histone H5 with the nucleosome and its implications for chromatin packing

Complex of linker histone H5 with the nucleosome and its implications for chromatin packing

Parathymosin Affects the Binding of Linker Histone H1 to Nucleosomes and Remodels Chromatin Structure

Hepatocyte Nuclear Factor 3 Relieves Chromatin-mediated Repression of the α-Fetoprotein Gene

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