ABSTRACT
Histone acetylation affects many nuclear processes including transcription, chromatin assembly, and DNA damage repair. Acetylation of histone H3 lysine 56 (H3 K56ac) in budding yeast occurs during mitotic S phase and persists during DNA damage repair. Here, we show that H3 K56ac is also present during premeiotic S phase and is conserved in fission yeast. Furthermore, the H3 K56ac modification is not observed in the absence of the histone chaperone Asf1. asf1delta and H3 K56R mutants exhibit similar sensitivity to DNA damaging agents. Mutational analysis of Asf1 demonstrates that DNA damage sensitivity correlates with (i) decreased levels of H3 K56ac and (ii) a region implicated in histone binding. In contrast, multiple asf1 mutants that are resistant to DNA damage display WT levels of K56ac. These data suggest that maintenance of H3 K56 acetylation is a primary contribution of Asf1 to genome stability in yeast.
Subject(s)
Cell Cycle Proteins/metabolism , Histones/metabolism , Lysine/metabolism , Meiosis/physiology , Molecular Chaperones/metabolism , S Phase/physiology , Saccharomyces cerevisiae Proteins/metabolism , Acetylation , Cell Cycle Proteins/chemistry , Cell Cycle Proteins/genetics , DNA Damage , Models, Molecular , Molecular Chaperones/genetics , Phenotype , Protein Conformation , Saccharomyces cerevisiae/cytology , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/metabolism , Saccharomyces cerevisiae Proteins/chemistry , Saccharomyces cerevisiae Proteins/genetics , Spores, Fungal/metabolismABSTRACT
This work describes the nature and sequence information content of the electron capture dissociation mass spectra for the intact Tetrahymena histone H2B. Two major variants of this protein were present bearing nominal modifications of both +42 and +84 Da. This work describes identification of the nature of these two modifications. For example, using gas-phase selection and isolation of the +42-Da modified species, from a background of two H2B variants each present in six or more posttranslationally modified isoforms, we were able to determine that this +42-Da modification isoform bears trimethylation rather than acetylation. LC-CIDMS analysis was also employed on digested preparations to obtain complementary detail of the nature of site-specific posttranslational modifications. This study establishes that integration of the information from these two datasets provides a comprehensive map of posttranslational occupancy for each particular covalent assemblage selected for structural investigation.
