CIDE-B [cell death-inducing DFF45 (DNA fragmentation factor 45)-like effector B] is a member of the CIDE family of apoptosis-inducing factors. The highly restricted pattern of expression of CIDE - B in the liver and spleen suggests that a mechanism exists for the tissue- and cell-specific regulation of transcription of this gene. We have analysed the promoters of the human CIDE - B gene, particularly the mechanism of cell-specific transcription. Expression of CIDE - B is driven by two promoters which are responsible for the synthesis of two types of transcript, and Sp1 and Sp3 are key regulators of basal transcription from both the upstream and the internal promoter, as indicated by EMSAs (electrophoretic mobility-shift assays) and site-directed mutagenesis. Bisulphite sequencing analysis demonstrated that the upstream promoter was hypermethylated in cells that did not express the long transcript of CIDE - B , but was hypomethylated in cells that expressed this transcript. Furthermore, methylation of this region in vitro reduced the promoter activity to ∼5% of the control. Thus methylation at CpG sites in the upstream promoter region appeared to be important for cell-specific synthesis of the long transcript. By contrast, HNF4α (hepatocyte nuclear factor-4α) bound to the internal promoter and enhanced its activity. Moreover, the short transcript of CIDE - B gene was expressed in cells which do not normally express this transcript upon introduction of exogenous HNF4α, demonstrating the involvement of HNF4α in the cell-specific synthesis of the short transcript. Thus our analysis revealed a novel mechanism for the cell-specific transcription of the human CIDE - B gene, which involves epigenetic and genetic control at separate respective promoters.