IMMATURE TRANSFORMED RAT ISLET BETA-CELLS DIFFERENTIALLY EXPRESS C-PEPTIDES DERIVED FROM THE GENES-CODING FOR INSULIN-I AND INSULIN-2 AS WELL AS A TRANSFECTED HUMAN INSULIN GENE

Synthetic peptides representing unique sequences in rat proinsulin C-peptide I and II were used to generated highly specific antisera, which, when applied on sections of normal rat pancreas, confirm a homogeneous coexpression of the two C-peptides in all islet beta-cells. Insulin gene expression is induced in the transformed heterogeneous rat islet cell clone, NHI-6F, by transient in vivo passage. During this process a transfected human insulin gene is coactivated with the endogenous nonallelic rat insulin I and II genes. Newly established cultures from NHI-6F insulinomas having a high frequency of insulin-producing cells showed highly differential expression at the cellular level of the three proinsulin C-peptide immunoreactivities, as follows: C-peptide I > human C-peptide > C-peptide II. The fractions of cells expressing human C-peptide and C-peptide II decreased in time and were absent after more than 50 successive passages, while a C-peptide I-producing population was still present. Double-labeling experiments revealed a heterogeneous distribution of the three different C-peptides. Surprisingly, in the early passages a large fraction of cells would express only a single species of proinsulin-C-peptide immunoreactivity but still at high levels. However, rat C-peptide II and human C-peptide were often colocalized, even in later passages. In situ hybridization studies combined with the immunocytochemical data suggest that the differential expression occurs at the level of transcription. We conclude that NHI-6F cells may represent a level of beta-cell differentiation characterized by a phenotype in which only a single insulin gene is active and that the insulin I gene is preferentially expressed under these conditions. The common behavior, i.e. coexpression and gradual inactivation during successive culture, of a transfected human insulin gene with the homologous rat insulin II gene may suggest the presence of common regulatory characteristics not shared with the retroposed rat insulin I gene.