Histone acetylation has been shown to be required for the proper regulation of 
many cellular processes including transcription, DNA repair, and chromatin 
assembly. Acetylation of histone H3 on lysine 56 (H3K56) occurs both during the 
premeiotic and mitotic S phase and persists throughout DNA damage repair. To 
learn more about the molecular mechanism of H3K56 acetylation and factors 
required for this process, we surveyed the genome of the yeast Saccharomyces 
cerevisiae to identify genes necessary for this process. A comparative global 
proteomic screen identified several factors required for global H3K56 
acetylation, which included histone chaperone Asf1 and a protein of an unknown 
function Rtt109 but not Spt10. Our results indicate that the loss of Rtt109 
results in the loss of H3K56 acetylation, both on bulk histone and on chromatin, 
similar to that of asf1Delta or the K56Q mutation. RTT109 deletion exhibits 
sensitivity to DNA damaging agents similar to that of asf1Delta and H3K56Q 
mutants. Furthermore, Rtt109 and H3K56 acetylation appear to correlate with 
actively transcribed genes and associate with the elongating form of polymerase 
II in yeast. This histone modification is also associated with some of the 
transcriptionally active puff sites in Drosophila. Our results indicate a new 
role for the Rtt109 protein in the proper regulation of H3K56 acetylation.