The product of the c-myc proto-oncogene, Myc, has been implicated in the transcriptional regulation of several genes, acting either as an activator or repressor of gene expression. To determine whether Myc is involved in the modulation of the expression of the CCAAT/enhancer-binding protein alpha (C/EBP alpha) gene, we used both stable cell lines overexpressing Myc and transient co-transfection assays. We show that the endogenous C/EBP alpha protein level is repressed in stable cell lines overexpressing Myc. We also show that enforced expression of Myc in mouse hibernoma HIB-1B cells dramatically repressed the expression of C/EBP alpha--promoter-reporter fusion genes. This effect of Myc was mediated through the core promoter region. Mutation of the initiator site could not abolish this affect, indicating that Myc may interact with some component(s) of the basal transcription machinery.