Compound Exocytosis and Cumulative Fusion in Eosinophils

Abstract

Focal release of cytotoxic proteins by eosinophils onto the target surface plays an important role in parasite killing. Degranulation was stimulated by intracellular application of calcium and guanosine 5 -3-O-(thio)triphosphate via the recording patch pipette or via streptolysin-O permeabilization. Exocytotic fusion was monitored by capacitance measurements, whereas release of fluorescent weak bases, which accumulate selectively within eosinophil granules, was followed by fluorescence imaging. Several distinct types of granule fusion events were directly observed by simultaneous capacitance and fluorescence measurements. These are fusion of a single granule with the plasma membrane, intracellular granule-granule fusion, fusion of large compounds of pre-fused granules with the plasma membrane (compound exocytosis), and sequential fusion of granules to granules previously fused to the plasma membrane. Extensive granule-granule fusion was also observed by electron microscopy of permeabilized cells. All these fusion mechanisms contribute to focal release. The coexistence of distinct modes of exocytosis suggests that their regulation may modulate effector functions of eosinophils during helminth infection and allergic response.

Glucagon-Like Peptide 1 Regulates Sequential and Compound Exocytosis in Pancreatic Islet β-Cells

“…In rat pituitary lactotrophs, cAMP activation increased secretory granule-secretory granule fusion but not the number of plasma membrane fusion events evoked by KCl stimulation, whereas the latter was selectively enhanced by protein kinase C activation (20). In horse eosinophils, protein kinase C did not exhibit any such effects (28), but instead, GTP␥S, a nonhydrolyzable GTP analog, induced secretory granule-secretory granule fusions and compound exocytic events while reducing the number of exocytic sites (29,30). GTP␥S stimulation was shown to potentiate insulin exocytosis in a cAMP-and Ca 2ϩ -dependent manner (31), but it remains unknown as to which molecular substrate or exocytic event is modulated by GTP␥S.…”

Deploying insulin granule–granule fusion to rescue deficient insulin secretion in diabetes

“…Multivesicular exocytosis was thought to be a less pronounced mode of exocytosis in Fig. 1 Modes of insulin granule exocytosis in the pancreatic islet beta cell beta cells, in contrast to the situation in many secretory cells, such as mast cells [10], eosinophils [11] and neutrophils [12]. In these secretory cells, a very large number of granules undergo rapid homotypic fusion to enable a massive release of granule cargo to effect the desired actions-allergic reactions [10], killing of invading parasites [11] and the acute inflammatory response [12], respectively.…”

“…1 Modes of insulin granule exocytosis in the pancreatic islet beta cell beta cells, in contrast to the situation in many secretory cells, such as mast cells [10], eosinophils [11] and neutrophils [12]. In these secretory cells, a very large number of granules undergo rapid homotypic fusion to enable a massive release of granule cargo to effect the desired actions-allergic reactions [10], killing of invading parasites [11] and the acute inflammatory response [12], respectively. Neuroendocrine cells such as beta cells seem to require less of this mode of compound exocytosis and instead exhibit a slower metered and sustained release, primarily effected by increasing primary exocytosis and, when required, release is further increased by restricted sequential fusion of only a few granules of two or three [6,7].…”

ADP Ribosylation Factor 6 (ARF6) Promotes Acrosomal Exocytosis by Modulating Lipid Turnover and Rab3A Activation

“…Controlled plasma membrane permeabilization has been used to gain access to the membrane fusion machinery required for exocytosis in several secretory cells (29,30). We used a well established streptolysin O (SLO) permeabilization protocol for studying acrosomal granule exocytosis (31)(32)(33)(34) that allows the incorporation of exogenous proteins, lipids, and ions into the cytosol.…”