ELECTRIC STUDIES OF EXOCYTOSIS AND ENDOCYTOSIS

Fusion and fission of membrane-bounded cells and organelles are
fundamental events governing the compartmentalization of
biological space. It is proposed to use video microscopy and an
ultra-sensitive, electrical assay of the cell surface to study, in
living cells, the mechanism of membrane fusion during exocytosis
of single secretory vesicles. In particular we will explore a novel
hypothesis: that the first step in membrane fusion is the
formation of an ion channel (the fusion pore) that connects the
inside of the vesicle to the cell exterior much like a gap junction
would connect two adjacent cells, and that subsequent event in
exocytosis are a consequence of electrolyte fluxes through the
fusion pore. Patch clamp studies of single secretory vesicles will
reveal the type of ion channels present in the vesicle membrane.
We will also explore "fusion pores" possibly formed by virus fusion
proteins stably expressed in a fibroblast-derived cell line, in order
to test whether there are similarities between fusion events
induced by viruses and fusion events during exocytosis. Lastly, we
plan to explore the role of cytosolic (Ca++) and other cytoplasmic
messengers in the control of pinocytosis and membrane turnover.