Anti-silencing function 1 (Asf1) is a highly conserved chaperone of histones 
H3/H4 that assembles or disassembles chromatin during transcription, 
replication, and repair. We have found that budding yeast lacking Asf1 has 
greatly reduced levels of histone H3 acetylated at lysine 9. Lysine 9 is 
acetylated on newly synthesized budding yeast histone H3 prior to its assembly 
onto newly replicated DNA. Accordingly, we found that the vast majority of H3 
Lys-9 acetylation peaked in S-phase, and this S-phase peak of H3 lysine 9 
acetylation was absent in yeast lacking Asf1. By contrast, deletion of ASF1 has 
no effect on the S-phase specific peak of H4 lysine 12 acetylation; another 
modification carried by newly synthesized histones prior to chromatin assembly. 
We show that Gcn5 is the histone acetyltransferase responsible for the 
S-phase-specific peak of H3 lysine 9 acetylation. Strikingly, overexpression of 
Asf1 leads to greatly increased levels of H3 on acetylation on lysine 56 and 
Gcn5-dependent acetylation on lysine 9. Analysis of a panel of Asf1 mutations 
that modulate the ability of Asf1 to bind to histones H3/H4 demonstrates that 
the histone binding activity of Asf1 is required for the acetylation of Lys-9 
and Lys-56 on newly synthesized H3. These results demonstrate that Asf1 does not 
affect the stability of the newly synthesized histones per se, but instead 
histone binding by Asf1 promotes the efficient acetylation of specific residues 
of newly synthesized histone H3.