MOUSE INSULINOMA BETA-TC3 CELLS EXPRESS PRODYNORPHIN MESSENGER-RIBONUCLEIC-ACID AND DERIVED PEPTIDES - A UNIQUE CELLULAR-MODEL FOR THE STUDY OF PRODYNORPHIN BIOSYNTHESIS AND PROCESSING

The tumor cell line beta TC3 has been established from insulinomas der ived from transgenic mice carrying a hybrid insulin promoter-simian vi rus-40 tumor antigen gene. The beta TC3 cells express high steady stat e levels of proinsulin messenger RNA (mRNA). In this same cell line, w e describe in the present study high expression levels of prodynorphin (pro-Dyn) mRNA and its derived peptides. By Northern blot analysis, t he screening of 23 cell lines of endocrine (n = 10) and of nonendocrin e (n = 13) origin revealed the presence of high level of the 2,6-kilob ase pro-Dyn transcript only in beta TC3 cells. The beta TC3 cells expr essed levels of pro-Dyn mRNA comparable to those in rat tissues expres sing pro-Dyn. Chromatographic and radioimmunological studies showed th at pro-Dyn mRNA was translated and fully processed into opioid peptide s with leucine-enkephalin (Leu-Enk)-extended sequences [dynorphin-A-(1 -8), dynorphin-B-(1-13), and alpha-neo-endorphin]. The expression of t he prohormone convertases was also examined in beta TC3 cells by North ern blot analysis. In addition to the ubiquitously expressed furin, be ta TC3 cells have abundant levels of prohormone convertase-1 (PC1) and PC2 mRNAs, but undetectable levels of PACE4 or PC5 mRNAs. Incubation of beta TC3 cells with 8-bromo-cAMP for 24 h stimulated 3-fold both th e pro-Dyn mRNA levels and the secretion of opioid peptides. In contras t to pro-Dyn mRNA, furin, PC1, and PC2 mRNA levels were not affected b y 8-bromo-cAMP. The beta TC3 cells constitute a unique model to elucid ate the biosynthetic pathway of pro-Dyn processing, to identify the pr oteolytic enzymes responsible for the production of pro-Dyn end produc ts, and to assess the potential role of opioid peptides in the regulat ion of pancreatic function.