Selective Recapture of Secretory Granule Components After Full Collapse Exocytosis in Neuroendocrine Chromaffin Cells

Abstract

In secretory cells, calcium-regulated exocytosis is rapidly followed by compensatory endocytosis. Neuroendocrine cells secrete hormones and neuropeptides through various modes of exo-endocytosis, including kiss-andrun, cavicapture and full-collapse fusion. During kissand-run and cavicapture modes, the granule membrane is maintained in an omega shape, whereas it completely merges with the plasma membrane during full-collapse mode. As the composition of the granule membrane is very different from that of the plasma membrane, a precise sorting process of granular proteins must occur. However, the fate of secretory granule membrane after full fusion exocytosis remains uncertain.Here, we investigated the mechanisms governing endocytosis of collapsed granule membranes by following internalization of antibodies labeling the granule membrane protein, dopamine-β-hydroxylase (DBH) in cultured chromaffin cells. Using immunofluorescence and electron microscopy, we observed that after full collapse, DBH remains clustered on the plasma membrane with other specific granule markers and is subsequently internalized through vesicular structures composed mainly of granule components. Moreover, the incorporation of this recaptured granule membrane into an early endosomal compartment is dependent on clathrin and actin. Altogether, these results suggest that after full collapse exocytosis, a selective sorting of granule membrane components is facilitated by the physical preservation of the granule membrane entity on the plasma membrane.

Dynamin 2 regulates biphasic insulin secretion and plasma glucose homeostasis

“…After insulin granule exocytosis and collapse onto the PM, the granule membrane components may be recaptured selectively as a whole unit or internalized by CME after being broken into smaller pieces. Both scenarios of LDCV membrane retrieval have been proposed in chromaffin cells (77,78). Abundant accumulation of CCPs indicates the significant role of CME among other forms of endocytosis in β cells.…”

A Nibbling Mechanism for Clathrin-mediated Retrieval of Secretory Granule Membrane after Exocytosis

“…A recent publication came to a similar conclusion as the present study that chromaffin granule membrane proteins remain associated and punctate after fusion and then are recaptured (42). The nibbling mechanism was not investigated, and the time resolution of the experiments was not sufficient to detect the critical processes that occur within seconds of fusion.…”

Oligophrenin-1: the link between calcium-regulated exocytosis and compensatory endocytosis in neuroendocrine cells

“…Using an assay developed in our laboratory to specifically follow the granule membrane recapture after exocytosis (for more details see [1] ), we observed a severe decrease (around 70%) of the compensatory endocytosis in chromaffin cells cultured from Ophn1 KO mice. This reveals a major role of OPHN1 in the pathway mediating compensatory endocytosis of the secretory granule membrane.…”

“…While the mechanisms underlying exocytosis per se have been extensively characterized in neuroendocrine cells, how the composition, integrity and functionality of the plasma membrane are maintained after exocytosis is obscure. However, in neuroendocrine chromaffin cells from the adrenal gland, we have proposed that secretory granule proteins cluster together at the plasma membrane after full fusion exocytosis before their recapture by compensatory endocytosis [1] . One of our current research goals is to determine the molecular machinery that enables this sorting, segregation and recapture of secretory granule membrane components.…”