Synaptotagmin-7 Endows a Subpopulation of Chromaffin Granules with Distinct Calcium Sensing and Fusion Properties

Abstract

Syt-7 is one of two major Ca2+-sensors for regulated release from adrenal chromaffin cells. Its unique biochemistry provides cells with a highly Ca2+ sensitive pool of dense core granules for exocytosis. Syt-7 also participates in limiting the rate at which fusion pores expand via mechanisms that are not fully clear. All previous imaging studies of Syt-7's actions during exocytosis in chromaffin cells have relied on overexpression of WT or mutant Syt protein. Here, we imaged exocytosis in mouse chromaffin cells without Syt-7 for the first time, so that properties which depend on the protein could be directly determined. Our data show that cells lacking Syt-7 exhibit at least a five-fold difference in fusion efficacy compared to WT cells in response to elevated K+ depolarization. Lumenal granule cargos are also released at faster rates from Syt-7 knockout (KO) cells, consistent with data from overexpression studies. Next, we measured the responses of WT and KO cells to endogenous secretogoguges of the sympatho-adrenal synapse, acetylcholine (ACh) and pituitary adenylate cyclase-activating peptide (PACAP). Compared to WT cells, ACh-evoked release in KO cells is significantly impaired with respect to granule fusion kinetics and fusion efficacy. PACAP-evoked release, on the hand, is only mildly suppressed. Our data provide evidence for Syt-7's role in two critical aspects of exocytosis – fusion and fusion pore expansion. Moreover, the separable responses of WT and KO cells to endogenous secretogogues suggest that Syt proteins occupy distinct roles in the physiology of the chromaffin cell system.