Ovine genomic urocortin: cloning, pharmacologic characterization, and distribution of central mRNA

Urocortin (Ucn), the newest member of the corticotropin-releasing factor (C RF) family of peptides, has been demonstrated to have significant physiolog ic and behavioral effects following its peripheral and central administrati on, respectively. In order to assess the differences in Ucn across species, an 18-kb sheep genomic DNA fragment encoding urocortin was isolated by the hybridization screening of a lambda phage Library with a probe generated f rom rat urocortin (rUcn) cDNA. The sheep clone contains a region that is 84 % and 88% homologous to the coding region of rUcn and human Ucn (hUcn), res pectively and encodes an ovine Ucn (oUcn) that is predicted to be identical to the rat peptide. Competitive binding assays demonstrated oUcn to have a high affinity (K-i = 0.1 nM) for the sheep CRF-binding protein (CRF-BP) an d localization studies by in situ hybridization have shown that the distrib ution of oUcn messenger RNA in sheep brain shares with that of rUcn in rat brain a predominant locus of expression in the Edinger-Westphal nucleus of the midbrain, though some secondary sites of expression reported in rat are not conserved. These findings demonstrate that, even across diverse specie s, Ucn is highly conserved with respect to its structure and pharmacology u nlike CRF where significant amino acid substitutions between the rat/human and sheep peptides may underlie differences in neuroendocrine regulation. ( C) 1999 Elsevier Science B.V. All rights reserved.