Expression of the endocytic proteins dynamin and amphiphysin in rat gastric enterochromaffin-like cells.

Abstract

Dynamin and amphiphysin play crucial roles in a variety of endocytic processes. Previous investigations of expression and functions of these proteins were performed mostly on neurons. The aim of this study was to investigate the presence and interaction of dyn and amph in gastric enterochromaffin-like cells. These endocrine cells of the gastric mucosa play a pivotal role in the regulation of acid secretion. Exocytosis of histamine-containing secretory vesicles has been described in detail. However, the mechanisms of endocytosis are unknown in this neuroendocrine cell type. Using RT-PCR and western blotting, we detected dynamin-1, -2 and -3 in highly enriched isolated enterochromaffin-like cells. Dynamin-1 and -2 were expressed at similar high levels, whereas dynamin-3 was of low abundance. Immunofluorescence microscopy located dynamin-1 and -2 to the cytoplasm and cell surface, whereas dynamin-3 was distributed differently in the perinuclear area. The presence of amphiphysin-1 and -2 RNAs was revealed by RT-PCR and a new splice variant of amphiphysin-2 was detected. Amphiphysin-1 and -2 were also detected in enterochromaffin-like cells by immunohistochemistry in the same locations as dynamin-1 and -2. Amphiphysin-1 and dynamin-1 co-immunoprecipitated with amphiphysin-2. In addition, dynamin-1 and amphiphysin-2 partially colocalized at the plasma membrane. Our results confirm the interaction of dynamin and amphiphysin and imply a role in endocytosis in enterochromaffin-like cells. To our knowledge, this is the first demonstration of the co-expression of all three dynamin isoforms in a non-tumor cell.