Proinsulin processing by the subtilisin-related proprotein convertases furin, PC2, and PC3.

Abstract

Experiments using recombinant vaccinia viruses expressing rat proinsulin I coinfected into COS-7 cells with recombinant vaccinia virus expressing human furin, human PC2, mouse PC3 (subtilisin-related proprotein convertases 1-3, respectively), or yeast Kex2 indicate that in this system both Kex2 and furin produce mature insulin, whereas PC2 selectively cleaves proinsulin at the C-peptide-A-chain junction. This is a property consistent with its probable identity with the rat insulinoma granule type II proinsulin processing activity as described by . PC3 generates mature insulin but cleaves preferentially at the proinsulin B-chain-C-peptide junction. This pattern of cleavage by PC3 is similar, but not identical, to that of the highly B-chain-C-peptide junction-selective type I activity as described by Davidson et aL, perhaps due to the presence of a P4 arginine residue near the C-peptide-A-chain junction unique to the rat proinsulins. These results along with data presented on the expression ofboth PC2 and PC3 in islet 13 cells strongly support the conclusion that these proteases are involved in the conversion of proinsulin to insulin in vivo.

Sorting and processing of secretory proteins

“…PCI and PC2 were localized exclusively to neuronal cells, but PC2 was more widely expressed than PCI. Western blot analysis suggests that PC2 is more strongly expressed than PCI in human insulinoma, while both are strongly expressed in mouse and rat islets [249,250]. A more detailed analysis of expression in rat islet cell subtypes reveals that there is relatively more PCI in insulin-producing B-cells than in non-B-cells, while the opposite is true for PC2 [251].…”

“…PCI mRNA is strongly expressed in the anterior lobe, while only weak expression of PC2 mRNA can be observed. [249]. Indeed, it is not yet clear whether PC2 is expressed at very low levels in these cells, or not at all [249,255].…”

“…[249]. Indeed, it is not yet clear whether PC2 is expressed at very low levels in these cells, or not at all [249,255]. A more precise co-localization study shows that, in the melanotrophs of the intermediate lobe, PC2 and POMC mRNAs are co-localized.…”

“…Finally, it has been shown that in some patients with familial hyperproinsulinaemia in which one of the basic amino acids at one of the cleavage sites has been altered, a partially cleaved proinsulin conversion intermediate is found in the circulation [297]. The cleavage specificity of proinsulin by PCI and PC2 has been analysed in more detail by co-transfection of rat insulin I into COS cells with human PCl and mouse PC2 [249]. PCI generates mature insulin but cleaves preferentially at the Bchain/C-peptide junction, while PC2 selectively cleaves at the Cpeptide/A-chain junction.…”

Hepatic Insulin Gene Therapy in Insulin-Dependent Diabetes Mellitus

“…This conversion can be accomplished in non-b-cells by introduction of minor changes in the amino acid sequence at the cleavage sites (56), such that the modified proinsulin can be cleaved by ubiquitously expressed endoproteases, such as furin (57,58).…”