In yeast cells, H2A.Z regulates transcription and is globally associated within a few nucleosomes of the initiator regions of numerous promoters. H2A.Z is deposited at these loci by an ATP-dependent complex, Swr1.com. Here we show that H2A.Z suppresses the p53 → p21 transcription and senescence responses. Upon DNA damage, H2A.Z is first evicted from the p21 promoter, followed by the recruitment of the Tip60 histone acetyltransferase to activate p21 transcription. p400, a human Swr1 homolog, is required for the localization of H2A.Z, and largely colocalizes with H2A.Z at multiple promoters investigated. Notably, the presence of sequence-specific transcription factors, such as p53 and Myc, provides positioning cues that direct the location of H2A.Z-containing nucleosomes within these promoters. Collectively, this study strongly suggests that certain sequence-specific transcription factors regulate transcription, in part, by preferentially positioning histone variant H2A.Z within chromatin. This H2A.Z-centered process is part of an epigenetic process for modulating gene expression.