Mitochondrial regulation of insulin production in rat pancreatic islets.
OBJECTIVE
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
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.
Leibowitz
G
G
Endocrinology and Metabolism Service, Department of Medicine, Hadassah-Hebrew University Medical Center, Jerusalem 91120, Israel. gleib@hadassah.org.il
Khaldi
M Z
MZ
Shauer
A
A
Parnes
M
M
Oprescu
A I
AI
Cerasi
E
E
Jonas
J-C
JC
Kaiser
N
N
D000818:Animals; D004305:Dose-Response Relationship, Drug; D005650:Fumarates / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D005973:Glutamine / Q000494:pharmacology; D066298:In Vitro Techniques; D007328:Insulin / Q000096:biosynthesis*; D007515:Islets of Langerhans / Q000378:metabolism*; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects; D008928:Mitochondria / Q000378:metabolism*; D011384:Proinsulin / Q000096:biosynthesis / Q000235:genetics; D011498:Protein Precursors / Q000096:biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013268:Stimulation, Chemical; D013385:Succinate Dehydrogenase / Q000378:metabolism; D019802:Succinic Acid / Q000494:pharmacology
D005650:Fumarates; D007328:Insulin; D011498:Protein Precursors; D012333:RNA, Messenger; D005973:Glutamine; C013737:preproinsulin; C032005:fumaric acid; D011384:Proinsulin; D019802:Succinic Acid; D013385:Succinate Dehydrogenase