DDIT3 is a tightly regulated basic leucine zipper (bZIP) transcription factor and key regulator in cellular stress responses. It is involved in a variety of pathological conditions and may cause cell cycle block and apoptosis. It is also implicated in differentiation of some specialized cell types and as an oncogene in several types of cancer. DDIT3 was originally believed to act as a dominant-negative inhibitor by forming heterodimers with other bZIP transcription factors, preventing their DNA binding and transactivating functions. DDIT3 has, however, been reported to bind DNA and regulate target genes. Here, we employed ChIP sequencing combined with microarray-based expression analysis to identify direct binding motifs and target genes of DDIT3. The results reveal DDIT3 binding to motifs similar to other bZIP transcription factors, known to form heterodimers with DDIT3. Binding to a class III satellite DNA repeat sequence was also detected. DDIT3 acted as a DNA-binding transcription factor and bound mainly to the promotor region of regulated genes. ChIP sequencing analysis of histone H3K27 methylation and acetylation showed a strong overlap between H3K27-acetylated marks and DDIT3 binding. These results support a role for DDIT3 as a transcriptional regulator of H3K27ac-marked genes in transcriptionally active chromatin.