Molecular cloning, characterization of cDNA, and distribution of mRNA encoding the frog prohormone convertase PC1

Abstract

Prohormone convertases (PCs) are calcium-dependent serine endoproteases of the subtilisin/kexin family that play a key role in the posttranslational processing of precursors for biologically active peptides. In this study, we have characterized the cDNA encoding PC1 in the European green frog Rana ridibunda. A frog brain cDNA library was screened by using a heterologous probe at low stringency, and a 2.3-kb cDNA clone encoding PC1 was isolated. This cDNA encodes a 736-residue protein with a 26-amino-acid signal peptide. Comparative structural analysis revealed that frog PC1 exhibits a high degree of amino acid identity with its mammalian counterparts, in particular in the subtilisin-like catalytic domain. Northern blot analysis resolved two major transcripts of 3.0 kb and 5.0 kb that were expressed differentially in the brain and pituitary. In situ hybridization studies showed that, in the frog brain, the highest densities of PC1 mRNA are present in the amygdala, the hypothalamus, and the anterior preoptic area. High concentrations of PC1 mRNA also were found in the pars distalis and pars intermedia of the pituitary, whereas the pars nervosa was devoid of hybridization signal. The wide distribution of PC1 mRNA in the brain and pituitary suggests that, in frog, PC1 is involved in the processing of a number of hormone and neuropeptide precursors.