Mitochondrial regulation of insulin production in rat pancreatic islets

Abstract

Aims/hypothesis: The study was designed to identify the key metabolic signals of glucose-stimulated proinsulin gene transcription and translation, focusing on the mechanism of succinate stimulation of insulin production. Methods: Wistar rat islets were incubated in 3.3 mmol/l glucose with and without esters of different mitochondrial metabolites or with 16.7 mmol/l glucose. Proinsulin biosynthesis was analysed by tritiated leucine incorporation into newly synthesised proinsulin. Preproinsulin gene transcription was evaluated following transduction with adenoviral vectors expressing the luciferase reporter gene under the control of the rat I preproinsulin promoter. Steady-state preproinsulin mRNA was determined using relative quantitative PCR. The mitochondrial membrane potential was measured by microspectrofluorimetry using rhodamine-123. Results: Succinic acid monomethyl ester, but not other mitochondrial metabolites, stimulated preproinsulin gene transcription and translation. Similarly to glucose, succinate increased specific preproinsulin gene transcription and biosynthesis. The inhibitor of succinate dehydrogenase (SDH), 3-nitropropionate, abolished glucose-and succinate-stimulated mitochondrial membrane hyperpolarisation and proinsulin biosynthesis, indicating that stimulation of proinsulin translation depends on SDH activity. Partial inhibition of SDH activity by exposure to fumaric acid monomethyl ester abolished the stimulation of preproinsulin gene transcription, but only partially inhibited the stimulation of proinsulin biosynthesis by glucose and succinate, suggesting that SDH activity is particularly important for the transcriptional response to glucose. Conclusions/interpretation: Succinate is a key metabolic mediator of glucose-stimulated preproinsulin gene transcription and translation. Moreover, succinate stimulation of insulin production depends on its metabolism via SDH. The differential effect of fumarate on preproinsulin gene transcription and translation suggests that these processes have different sensitivities to metabolic signals.

Mitochondrial succinate dehydrogenase is involved in stimulus-secretion coupling and endogenous ROS formation in murine beta cells

“…3a). Compared with the effects observed by Leibowitz et al [16], who tested 3-NPA in rat islets and observed complete inhibition of glucose-induced hyperpolarisation of Δψ m , the effect of 3-NPA was less pronounced in our experiments. This discrepancy might be due to the experimental design: we switched between 15 and 0.5 mmol/l glucose whereas Leibowitz et al [16] compared the effect of 3-NPA on Δψ m at 16.7 and 3 mmol/l glucose (levels at which respiratory chains are already partly activated).…”

“…Leibowitz et al [16] showed that for rat islets, SDH inhibition by 3-nitropropionic acid (3-NPA) or fumaric acid monoethyl ester (MEF) impaired insulin secretion by lowering gene transcription of preproinsulin and synthesis of proinsulin [16]. As a structural analogue of succinate, 3-NPA covalently and irreversibly binds to the active site of the enzyme thereby preventing complex-II-mediated FADH 2 generation [17].…”

The Pancreatic β-Cell: A Bioenergetic Perspective

“…rescence of a cationic probe such as TMRM (173,183), or the more slowly equilibrating rhodamine 123 (R123) (135,286,304). This technique is complicated by the plasma membrane potential, ⌬ p , which controls the gradient of the probe between the incubation medium and the cytosol.…”

“…Bearing in mind that the unstimulated ␤-cell maintains an unusually low ⌬G p , could protein synthesis be directly controlled by the phosphorylation potential? Leibowitz et al (286) observed that the cell-permeant monomethyl succinate produced a hyperpolarization of the in situ rat islet mitochondria, and enhanced insulin secretion, to an extent comparable to that with elevated glucose. [ 3 H]leucine incorporation into proinsulin was enhanced by both high glucose and monomethyl succinate, and the authors proposed a direct bioenergetic role for succinate, with a raised ⌬G p as a proximal signal for proinsulin biosynthesis.…”

Cell-Permeable Pentapeptide V5 Inhibits Apoptosis and Enhances Insulin Secretion, Allowing Experimental Single-Donor Islet Transplantation in Mice

“…The mitochondrial metabolite, succinate, is a key metabolic mediator of glucose-stimulated preproinsulin gene transcription and translation (43). Therefore, we examined mitochondrial function in V5-treated islets and found that mitochondrial function was increased by ϳ42% compared with untreated islets.…”