Regulation of insulin exocytosis by Munc13-1.
The slower kinetics of insulin release from pancreatic islet beta cells, as compared with other regulated secretory processes such as chromaffin granule secretion, can in part be explained by the small number of the insulin granules that are docked to the plasma membrane and readily releasable. In type-2 diabetes, the kinetics of insulin secretion become grossly distorted, and, to therapeutically correct this, it is imperative to elucidate the mechanisms that regulate priming and secretion of insulin secretory granules. Munc13-1, a synaptic protein that regulates SNARE complex assembly, is the major protein determining the priming of synaptic vesicles. Here, we demonstrate the presence of Munc13-1 in human, rat, and mouse pancreatic islet beta cells. Expression of Munc13-1, along with its cognate partners, syntaxin 1a and Munc18a, is reduced in the pancreatic islets of type-2 diabetes non-obese Goto-Kakizaki and obese Zucker fa/fa rats. In insulinoma cells, overexpressed Munc13-1-enhanced green fluorescent protein is translocated to the plasma membrane in a temperature-dependent manner. This, in turn, greatly amplifies insulin exocytosis as determined by patch clamp capacitance measurements and radioimmunoassay of the insulin released. The potentiation of exocytosis by Munc13-1 is dependent on endogenously produced diacylglycerol acting on the overexpressed Munc13-1 because it is blocked by a phospholipase C inhibitor (U73122) and abrogated when the diacylglycerol binding-deficient Munc13-1H567K mutant is expressed instead of the wild type protein. Our data demonstrate that Munc13-mediated vesicle priming is not restricted to neurotransmitter release but is also functional in insulin secretion, where it is subject to regulation by the diacylglycerol second messenger pathway. In view of our findings, Munc13-1 is a potential drug target for therapeutic optimization of insulin secretion in diabetes.
Sheu
Laura
L
Department of Medicine, University of Toronto, Toronto, Ontario M5S 1A8, Canada.
Pasyk
Eva A
EA
Ji
Junzhi
J
Huang
Xiaohang
X
Gao
Xiaodong
X
Varoqueaux
Frederique
F
Brose
Nils
N
Gaisano
Herbert Y
HY
D000818:Animals; D000954:Antigens, Surface / Q000378:metabolism; D015153:Blotting, Western; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D003921:Diabetes Mellitus, Experimental; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005089:Exocytosis; D005947:Glucose / Q000378:metabolism; D049452:Green Fluorescent Proteins; D006801:Humans; D015151:Immunoblotting; D007328:Insulin / Q000378:metabolism*; D007340:Insulinoma / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins; D007515:Islets of Langerhans / Q000378:metabolism; D008164:Luminescent Proteins / Q000378:metabolism; D008297:Male; D051379:Mice; D018613:Microscopy, Confocal; D008958:Models, Molecular; D051938:Munc18 Proteins; D009419:Nerve Tissue Proteins / Q000378:metabolism / Q000502:physiology*; D018408:Patch-Clamp Techniques; D011493:Protein Kinase C / Q000378:metabolism; D021381:Protein Transport; D051381:Rats; D011924:Rats, Zucker; D050827:Syntaxin 1; D013696:Temperature; D014162:Transfection; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors; D033921:Vesicular Transport Proteins / Q000378:metabolism
D000954:Antigens, Surface; D004791:Enzyme Inhibitors; D007328:Insulin; D047908:Intracellular Signaling Peptides and Proteins; D008164:Luminescent Proteins; D051938:Munc18 Proteins; D009419:Nerve Tissue Proteins; C494212:STX1A protein, human; C494214:Stx1a protein, mouse; C494213:Stx1a protein, rat; D050827:Syntaxin 1; C485548:UNC13B protein, human; C103912:Unc13a protein, rat; C485549:Unc13b protein, mouse; D033921:Vesicular Transport Proteins; D049452:Green Fluorescent Proteins; D011493:Protein Kinase C; D010738:Type C Phospholipases; D005947:Glucose