Differential expression of the two nonallelic proinsulin genes in the developing mouse embryo.

Abstract

In the mouse, insulin is produced from two similar but nonallelic genes that encode proinsulins I and II. We have investigated expression of these two genes during mouse embryonic development, using a PCR to detect the two gene transcripts and immunocytochemistry to visualize the two corresponding proteins. At appearance of the dorsal pancreatic anlage at day 9.5 of gestation, both mRNAs could be detected in the embryos, and both proteins were present together in the same cells of the developing pancreas. At days 9.5 and 10.5, when the ventral anlage appears, there were fewer proinsulin H mRNAs than proinsulin I mRNAs. At day 12.5 this ratio was reversed. Proinsulin II mRNA, but not proinsulin I mRNA, could be detected at day 8.5 in the prepancreatic embryo. Proinsulin II mRNA, but not proinsulin I mRNA, was also found in the heads of embryos at day 9.5 and at all later stages studied. These results indicate that the two proinsulin genes are regulated independently, at least in part. They also suggest that insulin might play a role as a growth factor in the developing mouse brain.Insulin, a key hormone in metabolic homeostasis, is synthesized, stored, and secreted by beta cells of the pancreatic islets. The protein is synthetized in the form of a precursor, preproinsulin, which is highly conserved among animal species. Unlike most mammals, mice and rats express two nonallelic genes that encode proinsulins I and II. In the mouse these two proteins differ by two amino acids in the B chain and three amino acids in the C peptide. Mouse C peptide I also lacks the Gly-Ala residues present in positions 17 and 18 of C peptide 11(1, 2). The corresponding genes in mouse and rat are highly homologous, and their organization is similar, except that the preproinsulin I gene possesses only the first of two introns present in the preproinsulin II gene (1, 3).On previous studies concerning the regulation of these two nonallelic genes, mRNAs were reported to be present in nearly equal quantities in adult pancreas of mouse (1, 4), as well as of rat (5) Here we have investigated regulation of the expression of the two proinsulin genes during mouse development. In the mouse, the pancreas is derived from the duodenum as two evaginations evolving at days 9.5 (dorsal) and 10.5 (ventral) of the gestation. The evaginations coalesce at day 11, and insulin has been first detected in previous studies at day 11.5 (9, 10). We have used a reverse-transcriptase-PCR (RT-PCR) assay, which allows identification and estimation of the relative amounts of mRNA transcribed from each of the two proinsulin genes. This RT-PCR, using a single pair ofprimers and a single probe for the two transcripts, allowed comparison of the relative amounts of proinsulin I and II mRNAs in the samples (11). Immunocytochemistry with antibodies specific for each of the two proinsulins allowed us to distinguish the products of the two genes in embryo sections. We have, thus, been able to detect insulin mRNAs and proteins much earlier, to distinguish the two forms, ...

Mouse MafA, homologue of zebrafish somite Maf 1, contributes to the specific transcriptional activity through the insulin promoter

“…Our results imply that, under conditions of stimulation, the difference between rat insulin promoters I and II depends on Maf regulation. It has been reported that mouse insulin promoters I and II are also differentially regulated [35,36]. Since the RIPE3b elements of both genes are identical, other elements except for RIPE3b may be responsible for the regulational difference between mouse insulin promoters I and II.…”

“…In this study, we indicated that MafA and MafB activate the RIPII and bind to the core site of RIPE3b. Mouse and rat possess two insulin promoters each (insulin promoters I and II), whereas human possesses only one promoter; these insulin promoters have unique transcriptional regulations [33][34][35][36]. Two significant differences appeared when the RIPE3b sequences of these insulin promoters were aligned with the MARE sequence (Fig.…”

“…It is well known that mouse and rat have two insulin genes and that each gene has a specific expression profile [33][34][35][36]. To assess the contribution of large Mafs to the transactivation activity of the insulin gene through the rat insulin promoters I and II (RIPI and II), we first The tree was generated by the UPGMA method using the whole amino acid sequences of the known large Mafs.…”

The MafA transcription factor appears to be responsible for tissue-specific expression of insulin

“…In contrast, other likely MafA-responsive genes, such as pdx-1 (28) and insulin I, were not activated effectively. Differences in the cis-element organization between the rodent insulin genes presumably limits the MafA responsiveness of insulin I because mouse insulin I and II are expressed independently and distinctly during development in the pancreas and thymus (29). In addition, glucagon expression was not influenced by MafA, suggesting that it does not interfere in ␣ cell-enriched MafB-mediated activation (ref.…”

Experimental control of pancreatic development and maintenance

“…Growth and morphogenesis of the bud epithelium leads to a ramification of small ductules containing the precursor cells of the acini, ducts, and islets of Langerhans, the major differentiated tissues of the mature pancreas (7,8). The islets form from dividing cells that escape the ductule epithelium, migrate through the surrounding mesenchyme, coalesce into endocrine cell masses, and differentiate into the four major islet cell types (8,9). The majority of the dividing cells remain within the ductule epithelium and differentiate into either acinar cells, which synthesize digestive hydrolytic enzymes, or mature duct cells, which secrete the fluid that transports the digestive enzymes to the intestine.…”