Doc2b serves as a scaffolding platform for concurrent binding of multiple Munc18 isoforms in pancreatic islet β-cells

Latha Ramalingam, Jingping Lu, Andy Hudmon, Debbie C Thurmond

Research output: Contribution to journalArticlepeer-review

14 Scopus citations

Abstract

Biphasic glucose-stimulated insulin secretion (GSIS) from pancreatic β-cells involves soluble N-ethylmaleimide-sensitive fusion protein-attachment protein receptor (SNARE) protein-regulated exocytosis. SNARE complex assembly further requires the regulatory proteins Munc18c, Munc18-1 and Doc2b. Munc18-1 and Munc18c are required for first- and second-phase GSIS respectively. These distinct Munc18-1 and Munc18c roles are related to their transient high-affinity binding with their cognate target (t-)SNAREs, Syntaxin 1A and Syntaxin 4 respectively. Doc2b is essential for both phases of GSIS, yet the molecular basis for this remains unresolved. Because Doc2b binds to Munc18-1 and Munc18c via its distinct C2A and C2B domains respectively, we hypothesized that Doc2b may provide a plasma membrane-localized scaffold/platform for transient docking of these Munc18 isoforms during GSIS. Towards this, macromolecular complexes composed of Munc18c, Doc2b and Munc18-1 were detected in β-cells. In vitro interaction assays indicated that Doc2b is required to bridge the interaction between Munc18c and Munc18-1 in the macromolecular complex; Munc18c and Munc18-1 failed to associate in the absence of Doc2b. Competition-based GST-Doc2b interaction assays revealed that Doc2b could simultaneously bind both Munc18-1 and Munc18c. Hence these data support a working model wherein Doc2b functions as a docking platform/scaffold for transient interactions with the multiple Munc18 isoforms operative in insulin release, promoting SNARE assembly.

Original languageEnglish (US)
Pages (from-to)251-8
Number of pages8
JournalBiochemical Journal
Volume464
Issue number2
DOIs
StatePublished - Dec 1 2014
Externally publishedYes

Keywords

  • Animals
  • Biphasic Insulins/metabolism
  • Calcium-Binding Proteins/chemistry
  • Exocytosis
  • Glucose/chemistry
  • Humans
  • Insulin-Secreting Cells/metabolism
  • Islets of Langerhans/metabolism
  • Mice
  • Mice, Knockout
  • Multiprotein Complexes
  • Munc18 Proteins/chemistry
  • Nerve Tissue Proteins/chemistry
  • Rats
  • SNARE Proteins/metabolism

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