NF-IL6 was originally identified as a DNA binding protein regulating interleukin-1 (IL-1)-stimulated IL-6 expression. Direct cloning of NF-IL6 showed its homology with C/EBP, a hepatocyte- and adipocyte-specific transcription factor. This study showed that the expression of NF-IL6 messenger RNA (mRNA) increased markedly during the differentiation to a (mRNA) increased markedly during the differentiation to a macrophage lineage in mouse myeloid leukemia cells M1, human histiocytic leukemia cells U937, promyelocytic leukemia cells HL-60, and human peripheral monocytes. Particularly in HL-60 cells that undergo granulocyte or macrophage differentiation depending on inducers, NF-IL6 mRNA was specifically upregulated during macrophage differentiation but not granulocyte differentiation. It was also shown that the functional NF-IL6 protein increased during the differentiation of U937 cells. Furthermore, recombinant NF-IL6 was found to bind to the regulatory regions of the IL-1, tumor necrosis factor, granulocyte colony-stimulating factor, and lysozyme genes, which are expressed in mature macrophages. These results suggest that NF-IL6 may possibly be involved as an important transcription factor in the process of activation and/or differentiation of macrophages.