NuA4 subunit Yng2 function in intra-S-phase DNA damage response.

In eukaryotes, DNA is assembled into nucleosomes, the basic structural unit of chromatin. Factors that target nucleosomes to modulate chromatin structure and provide access to nucleosomal DNA are required to facilitate processes such as transcription, replication, and DNA repair. Covalent modification of core histone N-terminal tails by acetylation of conserved lysine residues is thought to relax chromatin structure and allow access of nonhistone factors to nucleosomal DNA (8, 43). Seminal studies by Smerdon and colleagues showed that histone acetylation might be upregulated to facilitate DNA damage repair in mammalian cells after UV irradiation (40, 41, 46). Recent biochemical results support this model. The mammalian TATA-binding protein-free Taf II complex is recruited along with nucleotide excision repair proteins to UV-damaged DNA (5). Importantly, via its Gcn5 histone acetyltransferase (HAT) subunit, the TATA-binding protein-free Taf II complex preferentially acetylates histone H3 in nucleosomes containing UV-damaged DNA. In turn, acetylation of histone H4 also appears to function in DNA damage repair, as expression of a dominant-negative form of the Tip60 H4 HAT in mammalian cells blocks repair of double strand breaks (23). Studies of histone N-terminal tail acetylation in the budding yeast Saccharomyces cerevisiae also support a role for histone acetylation in DNA repair and genomic integrity. Cells expressing H4 mutants lacking N-terminal acetylation sites activate DNA damage repair signaling and perform a G2/M delay even during unperturbed vegetative growth (33). In turn, conditional mutants in the essential yeast Tip60 homolog Esa1, the catalytic subunit of NuA4, lose nucleosomal histone H4 acetylation and accumulate in G2/M at the nonpermissive temperature. The terminal arrest is partly relieved by abrogating DNA damage checkpoint arrest (13). Analogously, mutants deficient in the Gcn5 or Sas3 histone H3 HAT display a G2/M delay which may also reflect DNA damage checkpoint activation (22, 51). The molecular pathway leading to DNA damage checkpoint activation in histone acetylation mutants remains poorly understood. Perhaps transcription of genes necessary for DNA metabolism is critically impaired, reflecting the known requirement for targeted histone acetylation in transcriptional activation (6, 28). Nonetheless, transcription defects are unlikely to be the only underlying mechanism. Studies of genome-wide expression in yeast histone H4 acetylation mutants revealed surprisingly few significant changes (10, 42). In addition to gene-specific regulation, Esa1 has been implicated in modification of histone H4 across large segments of chromosomes (50). One interpretation is that the DNA-damage-dependent cell cycle arrest with nonacetylatable H4 and esa1 mutants may arise from the effects of decreased genome-wide acetylation per se rather than decreased expression of specific transcripts. Like Esa1, virtually every other NuA4 subunit is essential for cell viability (2, 20). However, mutants lacking the NuA4 subunit Yng2 remain viable and yet are specifically compromised in global nucleosomal histone H4 acetylation (10, 30, 35). Mutants lacking Yng2 display a G2/M delay (10) and are sensitized to methylmethane sulfonate (35). Thus, we have used the yng2 mutant as a tool to study the role of NuA4 activity in DNA damage responses. As for esa1, the yng2 G2/M delay is DNA damage checkpoint dependent. Further, we demonstrate that Yng2 is required specifically to respond to DNA damage during S phase, suggesting a critical role for NuA4 in maintaining genomic integrity during DNA replication.