The C/EBP family of transcription factors regulates viral and cellular CCAAT/enhancer element-mediated transcription. We report the isolation and characterization of genomic and cDNA clones encoding avian CCAAT/enhancer-binding protein-gamma (C/EBP gamma). A partial cDNA clone for a C/EBP-related gene was previously identified by expression library screening for proteins binding the A1 CCAAT/enhancer motif from the avian leukosis virus long terminal repeat [W. Bowers and A. Ruddell (1992) J. Virol. 66, 6578-6586]. Additional cDNA and genomic clones were generated and sequenced to identify the complete protein coding sequence of this gene. Sequence analysis indicates that this gene encodes the avian homolog of C/EBP gamma. As with the murine C/EBP gamma homolog, the avian C/EBP gamma gene is comprised of two exons, with the open reading frame encoded in exon 2. The 150-aa C/EBP gamma protein is highly conserved, as the avian protein shows more than 80% identity with the murine and human homologs. The sequence of the initiation methionine (-3 caaAUGa +4) from the 150-aa open reading frame has a non-optimal Kozak initiation sequence. In vitro transcription and translation assay of this avian cDNA followed by radioimmunoprecipitation assay using a murine C/EBP gamma antiserum indicates that this non-optimal initiation codon is used to express a 22-kDa DNA-binding protein.