We identified a novel human gene, NOC4 (Neighbor Of COX4), located 5' to COX4, the gene for cytochrome c oxidase subunit IV, on Chr 16q32-ter. Transcripts from this gene were identified among human expressed sequence tags. A full-length, 1.06-kb human retinal NOC4 cDNA encoded a 24-kDa, 210-amino acid hypothetical protein of unknown function. Northern hybridization analysis of human RNAs from various tissues detected NOC4 transcripts of 2.2 and 1.4 kb in all tissues examined, suggesting that NOC4 expression is ubiquitous. Transcription of both the COX4 and NOC4 genes initiates within a 250-bp intergenic promoter and occurs in opposite directions. The bidirectional promoter is G + C-rich, lacks TATA and CCAAT elements, and contains multiple potential binding sites for Sp1 and NRF-2/GABP. Two of the NRF-2/GABP sites are located within 14-bp direct repeats, a conserved feature of mammalian COX4 promoters. The NOC4 and COX4 genes are also linked in the rat, mouse, and bovine genomes. A NOC4-GFP fusion protein is located in both the nucleus and the cytoplasm, including the mitochondria.