To understand the role of histone H3 sub-domains in chromatin function, 35 histone 113 tandem alanine mutants were generated and tested for their viability and sensitivity to DNA damaging agents. Among 13 non-viable H3 mutants, 6 were mapped around the alpha N helix and preceding tail region. Mutants with individual alanine substitutions in this region were viable but developed multiple sensitivities to DNA damaging agents. The only viable triple Mutant, RE149-50A, in the aN helix region could not grow when combined with histone chaperone mutations, such as asf1 Delta, cac1 Delta, or hir1 Delta, suggesting that this particular region is important when the histone assembly/disassembly pathway is compromised. In addition, further analysis showed that T45, E50, or F54 of the alpha N helix genetically interacted witha histone chaperone(Asf1) and transcription elongation factors (Paf1 and Hpr1). These results Suggest a specific role of the H3 alpha N helix in histone dynamics mediated by histone chaperones, which might be important during transcription elongation. (C) 2008 Elsevier Inc. All rights reserved.