The proglucagon gene is expressed in the pancreas, intestines, and brain. Tissue-specific post-translational processing of proglucagon results in the liberation of glucagon in the pancreas, and glucagon-like peptide-1 and -2 in the intestines and brain. Proglucagon-derived peptides play essential roles in human physiology. To identify sequences that may have roles in regulation of human proglucagon expression, the strategy of comparative genomics and in vitro transfection were utilized in my research. Human proglucagon genomic sequence was compared with orthologous rodent sequences. An evolutionarily conserved 483 bp region was found in intron I of the proglucagon gene. The transcriptional activity of intron I of the human proglucagon gene and that of the conserved region within intron I were tested in rodent cell lines. I demonstrated that intron I of the human proglucagon gene is an enhancer-like element, and its enhancer activity is confined within the conserved 483 bp segment. Thus, I provided a novel regulatory mechanism underlying the regulation of human proglucagon gene expression.