Functional Analyses of an Evolutionarily Conserved Acidic Patch on the Nucleosome

Abstract

The eukaryotic canonical nucleosome has an acidic patch on each H2A/H2B dimer. This acidic patch is also detected in histone variants, such as the H2A.Z (yeast Htz1)/H2B dimer. Here, we screened a comprehensive histone point mutant library and identified 11 histone residues located in four distinct nucleosome domains (Homologous Recombination (HR) Domain I–IV (HRD-I–IV)) with a potential role in HR. H2A-L66, -E93, and -L94 residues in HRD-I are located in the acidic patch region. Equivalent residues (H2A-L66 and Htz1-L73) partly compensate the function of each dimer. A common residue H2B-L109, which is located underneath of the acidic patch in both dimers, also partly compensates the function of each dimer. Upon exposure to DNA double-strand break (DSB)-inducing agents, the fragmented chromosomes of H2A-L66A mutant cells exhibited slow and limited recovery into intact chromosomes, suggesting that the H2A-L66A mutant is partly deficient in DSB repair. Furthermore, strand invasion, one of critical steps of HR, could be less efficient in H2A-L66A cells. All 11 HRD residues, including H2A-L66, are highly conserved in extant eukaryotic cells; therefore, our screening reported in this study will provide a foundation for future studies about the mechanisms underlying eukaryotic HR based on chromatin.