Interaction between Munc13-1 and RIM is critical for glucagon-like peptide-1 mediated rescue of exocytotic defects in Munc13-1 deficient pancreatic beta-cells

Diabetes. 2007 Oct;56(10):2579-88. doi: 10.2337/db06-1207. Epub 2007 Jul 16.

Abstract

Objective: Glucagon-like peptide-1 (GLP-1) rescues insulin secretory deficiency in type 2 diabetes partly via cAMP actions on exchange protein directly activated by cAMP (Epac2) and protein kinase A (PKA)-activated Rab3A-interacting molecule 2 (Rim2). We had reported that haplodeficient Munc13-1(+/-) mouse islet beta-cells exhibited reduced insulin secretion, causing glucose intolerance. Munc13-1 binds Epac2 and Rim2, but their functional interactions remain unclear.

Research design and methods: We used Munc13-1(+/-) islet beta-cells to examine the functional interactions between Munc13-1 and Epac2 and PKA. 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.

Results: To determine which exocytotic steps caused by Munc13-1 deficiency are rescued by Epac2 and PKA, we used 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(+/-) beta-cells, 2) Epac-selective agonist [8-(4-chloro-phenylthio)-2'-O-methyladenosine-3',5'-cyclic monophosphate] partially restored the reduced RRP and refilling of a releasable pool of vesicles, and 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 (N(6)-benzoyladenosine-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, whereas 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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • ATP-Binding Cassette Transporters / physiology*
  • Animals
  • Cyclic AMP / metabolism
  • Exocytosis / drug effects
  • Exocytosis / physiology*
  • Glucagon-Like Peptide 1 / pharmacology*
  • Insulin / deficiency
  • Insulin / metabolism*
  • Insulin Secretion
  • Insulin-Secreting Cells / drug effects
  • Insulin-Secreting Cells / physiology*
  • Mice
  • Mice, Knockout
  • Nerve Tissue Proteins / deficiency
  • Nerve Tissue Proteins / genetics
  • Nerve Tissue Proteins / physiology*
  • Patch-Clamp Techniques

Substances

  • ATP-Binding Cassette Transporters
  • Abca4 protein, mouse
  • Insulin
  • Nerve Tissue Proteins
  • Unc13a protein, mouse
  • Glucagon-Like Peptide 1
  • Cyclic AMP