We have isolated a novel cDNA clone, named AZ2, from a cDNA library of mRNA prepared from C3H10T1/2 cells that had been transiently exposed to 5-azacytidine, a potent inhibitor of DNA methyltransferase. The elucidated nucleotide sequence revealed that the 5' region of the cDNA was rich in the CpG sequence. The AZ2 cDNA contained a 1215-nucleotide open reading frame, and the expected amino acid sequence had a molecular mass of 46090. The amount of the transcript increased on 5-azacytidine treatment of C3H10T1/2 cells, and the transcript was significantly expressed in mouse testis, brain, lung, kidney, heart and ovary. Specific antibodies raised against a fusion protein including glutathione S-transferase revealed a band of an approximately 48 kDa translation product for testis, brain, lung, and cultured cells that ectopically expressed the AZ2 protein. The AZ2 protein was mainly localized in the cytoplasm. The amino-terminal part of the AZ2 protein was homologous to the previously reported TANK (Cheng and Baltimore, 1996. Genes Dev. 10, 963-973) and I-TRAF (Rothe et al., 1996. Proc. Natl. Acad. Sci. USA 93, 8241-8246), which participate in the signal transduction cascade from the tumor necrosis factor-receptor to the transcription factor, NF kappa B. Overexpression of AZ2 inhibited TNF alpha mediated NF kappa B activation. AZ2 could be a component of a regulator of the NF kappa B activation cascade. (C) 1999 Elsevier Science B.V. All rights reserved.