The factors that control fat deposition in adipose tissues are poorly understood. It is known that visceral adipose tissues display a range of biochemical properties that distinguish them from adipose tissues of subcutaneous origin. However, we have little information on gene expression either in relation to fat deposition or on inter-species variation in fat deposition. The first step in this study was to identify genes expressed in fat depots of cattle. We used a differential display method to compare PCR-amplified gene transcripts in subcutaneous and visceral adipose tissue of cattle. Twenty-nine transcripts were identified that showed apparent differential expression; these transcripts were classified into eight functional groups. Among the transcripts identified as having differential expression in the two adipose tissues were cell division cycle 42 homolog (CDC42), prefoldin 5, decorin, phosphate carrier, 12S ribosomal RNA gene, and kelch repeat and BTB domain containing 2 (Kbtbd2). In subsequent experiments, we determined the expression levels of these latter genes in the pig and in mice fed either a control or high-fat diet in order to compare the regulation of fat accumulation in other animal species. The levels of CDC42 and decorin mRNA were found to be higher in visceral adipose tissue than in subcutaneous adipose tissue in cattle, pig and mice. However, the other genes studied did not show consistent expression patterns between the two tissues in cattle, pigs and mice. Interestingly, all genes were upregulated in subcutaneous and/or visceral adipose tissues of mice fed the high-fat diet compared to the control diet. The data presented here extend our understanding of gene expression in fat depots and provide further proof that the mechanisms of fat accumulation differ significantly between animal species.