The mammalian EGLN family contains three paralagous genes (EGLN1, EGLN2, and EGLN3) encoding prolyl hydroxylase isoforms that mediate the oxygen-dependent targeting of the transcription factor hypoxia inducible factor alpha to the proteosome. The rat orthologue of EGLN3 (SM-20) exhibits tissue-restricted expression, is induced by growth factors in cultured vascular smooth muscle, and is up-regulated during myogenesis. To determine if all three EGLN genes are coordinately regulated, we examined their mRNA expression in murine tissues and in cultured cells. We now report that the three murine EGLN mRNAs have unique but overlapping patterns of tissue expression. The most striking differences were in the heart, where EGLN3 had its highest levels of expression, and the testis, where EGLN2 was the only isoform expressed. In cultured vascular smooth muscle cells, serum treatment led to up-regulation of EGLN1 and EGLN3, but not EGLN2, and only EGLN3 was superinduced by cyclohexamide. In cultured C2C12 myocytes, EGLN3 was up-regulated during differentiation, whereas EGLN1 and EGLN2 were constitutively expressed. The abundance of EGLN3 mRNA in the heart, its induction by growth factors in vascular smooth muscle, and its regulation during C2C12 differentiation suggest a unique role for EGLN3 and might justify the development of isoformspecific inhibitors.