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Interaction between Munc13-1 and RIM is Critical for Glucagon-like Peptide-1-mediated Rescue of Exocytotic Defects in Munc13-1-deficient pancreatic ß-cells

Departments of ¹Physiology & Medicine, University of Toronto, Toronto, Ontario, Canada, and
² Department of Clinical and Molecular Pathology, University of Toyama, Toyama, Japan.

Correspondence: herbert.gaisano{at}utoronto.ca

GLP-1 rescues insulin secretory deficiency in type 2 diabetes partly via cAMP actions on Epac2 and PKA-activated Rim2. We had reported haplodeficient Munc13-1^+/– mouse islet ß-cells exhibited reduced insulin secretion causing glucose intolerance. Munc13-1 binds Epac2 and Rim2, but their functional interactions remain unclear.

Aim:We employed Munc13-1^+/– islet ß-cells to examine the functional interactions between Munc13-1 and Epac2 and PKA.

Results:GLP-1 stimulation of Munc13-1^+/– islets normalized the reduced biphasic insulin secretion by its actions on intact islet cAMP production and normal Epac2 and Rim2 levels. To determine which exocytotic steps caused by Munc13-1 deficiency are rescued by Epac2 and PKA, we employed patch-clamp capacitance measurements, showing that: 1) cAMP restored the reduced readily-releasable pool (RRP), and partially restored refilling of a releasable pool of vesicles in Munc13-1^+/– ß-cells. 2) Epac-selective agonist (8-pCPT-2'-O-Me-cAMP) partially restored the reduced RRP and refilling of a releasable pool of vesicles. 3) PKA blockade by H89 (leaving Epac intact) impaired cAMP ability to restore the RRP and refilling of a releasable pool of vesicles. Conversely, PKA-selective agonist (N⁶Bnz-cAMP) completely restored RRP and partially restored refilling of a releasable pool of vesicles. To determine specific contributions within Epac-Rim2-Munc13-1 interaction sites accounting for cAMP rescue of exocytosis caused by Munc13-1 deficiency, we found that blockade of Rim2-Munc13-1 interaction with Rim-Munc13-1-binding domain peptide abolished cAMP rescue, while blockade of Epac-Rim2 interaction with Rim2-PDZ peptide only moderately reduced refilling with little effect on RRP.

Conclusions:cAMP rescue of priming defects caused by Munc13-1 deficiency via Epac and PKA signaling pathways requires downstream Munc13-1-Rim2 interaction.

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