Phorbolester-activated Munc13-1 and ubMunc13-2 exert opposing effects on dense-core vesicle secretion

Elife. 2022 Oct 10:11:e79433. doi: 10.7554/eLife.79433.

Abstract

Munc13 proteins are priming factors for SNARE-dependent exocytosis, which are activated by diacylglycerol (DAG)-binding to their C1-domain. Several Munc13 paralogs exist, but their differential roles are not well understood. We studied the interdependence of phorbolesters (DAG mimics) with Munc13-1 and ubMunc13-2 in mouse adrenal chromaffin cells. Although expression of either Munc13-1 or ubMunc13-2 stimulated secretion, phorbolester was only stimulatory for secretion when ubMunc13-2 expression dominated, but inhibitory when Munc13-1 dominated. Accordingly, phorbolester stimulated secretion in wildtype cells, or cells overexpressing ubMunc13-2, but inhibited secretion in Munc13-2/Unc13b knockout (KO) cells or in cells overexpressing Munc13-1. Phorbolester was more stimulatory in the Munc13-1/Unc13a KO than in WT littermates, showing that endogenous Munc13-1 limits the effects of phorbolester. Imaging showed that ubMunc13-2 traffics to the plasma membrane with a time-course matching Ca2+-dependent secretion, and trafficking is independent of Synaptotagmin-7 (Syt7). However, in the absence of Syt7, phorbolester became inhibitory for both Munc13-1 and ubMunc13-2-driven secretion, indicating that stimulatory phorbolester x Munc13-2 interaction depends on functional pairing with Syt7. Overall, DAG/phorbolester, ubMunc13-2 and Syt7 form a stimulatory triad for dense-core vesicle priming.

Keywords: adrenal chromaffin cell; capacitance measurements; exocytosis; mouse; neuroscience; neurotransmitter release; vesicle fusion; vesicle priming.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Dense Core Vesicles
  • Diglycerides*
  • Exocytosis
  • Mice
  • Phorbol Esters*
  • SNARE Proteins / metabolism
  • Synaptotagmins

Substances

  • Diglycerides
  • Phorbol Esters
  • SNARE Proteins
  • Synaptotagmins
  • Unc13a protein, mouse
  • Unc13b protein, mouse

Grants and funding

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.