Saccharomyces cerevisiae encodes a single linker histone, Hho1p, with two globular domains. This raised the possibility that Hho1p could bind to two nucleosome cores simultaneously. To evaluate this idea, we studied the ability of a four-way junction, immobilized on the surface of a magnetic bead, to pull down a radiolabeled four-way junction in the presence of different Hho1 proteins. Four-way junctions are known to bind to H1, presumably due to structure similarities to the DNA at the nucleosomal entry/exit point. We found a significant increase in the ability of full-length Hho1p to pull down radiolabeled four-way junction DNA under ionic conditions where both globular domains could bind. The binding was structure specific, since the use of double-stranded DNA, or a mutant Hho1p in which the second DNA binding site of globular domain 1 was abolished, resulted in a significant decrease in bridged binding. Additionally, bridged binding required a covalent attachment between the two globular domains, since factor Xa protease treatment of the complex formed by a modified Hho1p that contained a factor Xa cleavage site between the two globular domains resulted in a significant release of radiolabeled four-way junction. These findings demonstrated that the two globular domains independently associated with two different four-way junction molecules in a manner that required amino acid residues implicated in structure-specific binding in the nucleosome. We discuss the implication of these findings on the chromatin structure of yeast and propose a model where a single Hho1 protein binds to two serially adjacent nucleosomes.
|Evidence ID||Analyze ID||Interactor||Interactor Systematic Name||Interactor||Interactor Systematic Name||Type||Assay||Annotation||Action||Modification||Phenotype||Source||Reference||Note|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Gene Ontology Term||Gene Ontology Term ID||Qualifier||Aspect||Method||Evidence||Source||Assigned On||Reference||Annotation Extension|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Phenotype||Experiment Type||Experiment Type Category||Mutant Information||Strain Background||Chemical||Details||Reference|
|Evidence ID||Analyze ID||Regulator||Regulator Systematic Name||Target||Target Systematic Name||Experiment||Conditions||Strain||Source||Reference|