Abstract
Gaining the full activity of the insulin receptor (IR) requires the proteolytic cleavage of its proform by intra-Golgi furin-like activity. In mammalian cells, IR is expressed as two isoforms (IRB and IRA) that are responsible for insulin action. However, only IRA transmits the growth-promoting and mitogenic effects of insulin-like growth factor 2. Here we demonstrate that the two IR isoforms are similarly cleaved by furin, but when this furin-dependent maturation is inefficient, IR proforms move to the cell surface where the proprotein convertase PACE4 selectively supports IRB maturation. Therefore, in situations of impaired furin activity, the proteolytic maturation of IRB is greater than that of IRA, and accordingly, the amount of phosphorylated IRB is also greater than that of IRA. We highlight the ability of a particular proprotein convertase inhibitor to effectively reduce the maturation of IRA and its associated mitogenic signaling without altering the signals emanating from IRB. In conclusion, the selective PACE4-dependent maturation of IRB occurs when furin activity is reduced; accordingly, the pharmacological inhibition of furin reduces IRA maturation and its mitogenic potential without altering the insulin effects.
Highlights
The insulin receptor exists as two isoforms: IRA and IRB
We demonstrate that the two IR isoforms are cleaved by furin, but when this furin-dependent maturation is inefficient, IR proforms move to the cell surface where the proprotein convertase PACE4 selectively supports IRB maturation
In the Absence of Furin, IRB Proteolytic Maturation Is Higher than That of IRA—The use of a computational tool for predicting furin cleavage sites (PiTou software) that was developed based on the functional characterization of the 20-residue recognition sequence motif [38] revealed that both proIR isoforms can be cleaved by furin at the unique expected site (Arg735 for IRB and Arg723 for IRA) with similar efficiencies. Consistent with this prediction, comparable amounts of immature and mature IRA and IRB forms were detected in the lysates of HeLa cells that independently overexpressed either of these isoforms (Fig. 1A); the proteolytic maturation of both IR
Summary
The insulin receptor exists as two isoforms: IRA and IRB. Results: IRA and IRB are matured by furin, but when furin activity is reduced, IRB is matured by PACE4. The selective PACE4-dependent maturation of IRB occurs when furin activity is reduced; the pharmacological inhibition of furin reduces IRA maturation and its mitogenic potential without altering the insulin effects. IRA lacks and IRB contains a 12-amino acid segment located in the carboxyl terminus of the insulin-like growth factor; ML, multi-Leu peptide inhibitor (Ac-LLLLRVKRNH2); PEG8-ML, PEGylated multi-Leu peptide inhibitor; PC, proprotein convertase; Phac-RVR-4-Amba, proprotein convertase inhibitor phenylacetylArg-Val-Arg-4-amidinobenzylamide; PKB, protein kinase B; proIR, proform of the insulin receptor. We took advantage of this difference in IRA and IRB maturation to demonstrate that it is possible to reduce the mitogenic signals emanating from the IR by inhibiting IRA maturation
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