The p53 protein is a transcription factor that trans-activates various genes in response to DNA-damaging stress. To search for new p53-target genes, we applied a cDNA microarray system using two independent p53-inducible cell lines, followed by in silico analysis to detect p53 response elements. Here, we report on crystallin alpha B gene (CRYAB), which encodes αB-crystallin, and is one of the genes directly trans-activated by p53. We confirmed it is directly transcribed by p53 using promoter analysis, deletion reporter assay, ChIP assay and EMSA. αB-crystallin is also upregulated in a p53-dependent manner and binds to the DNA-binding domain of p53. Overexpression of αB-crystallin increased p53 protein and, in contrast, repression of αB-crystallin decreased p53 protein. Interestingly, both overexpression and repression of αB-crystallin reduced p53-dependent apoptosis. In conclusion, we identified that αB-crystallin was a novel p53-target gene and required for p53-dependent apoptosis using two independent p53-inducible cell lines. This is the first report associating p53 directly with a heat shock protein through trans-activation and physical interaction.