Possible temperature‐dependent blockage of synaptic vesicle recycling induced by a single gene mutation in drosophila

Abstract

The temperature-sensitive Drosophila mutant, shibirets1 (shi), has been shown to exhibit a reversible block in synaptic transmission at 30 degrees C. Various synaptic terminals (neuromuscular, sensory, central) of this mutant were observed by electron microscopy. At 19 degrees C, all terminals of shi showed essentially the same structural features as those of wild-type (Oregon-R) flies, while at 30 degrees C (5 or 10 min of exposure) shi terminals exhibited various structural changes not seen in the wild type. The major structural change observed in all of the various types of terminals was the accumulation of many pitlike structures on the plasma membrane near presynaptic sites. These structures consisted of a spherical head portion, about 50-100 nm in diameter, and a cylindrical neck portion, about 20 nm long and 20-25 nm in diameter. The neck portion was surrounded by a kind of cytoplasmic dense material, about 10 nm thick, reminiscent of a "collar." Thus, these pits are referred to as "collared pits." Similar kinds of pits were observed, although very rarely, in wild-type flies at 19 and 30 degrees C and in shi flies at 19 degrees C. In addition, various degrees of vesicle depletion, and an increase in membranous structures (infoldings and cisternalike or tubulelike structures) often accompanied pit formation. The possibility that these pits are the result of a blocked step in the endocytotic process, which in turn causes vesicle depletion as exocytosis proceeds, is discussed.

A Model for Dynamin Self-assembly Based on Binding Between Three Different Protein Domains

“…Purified dynamin also assembles into spirals or rings in the absence of membrane (5). These structures are strikingly similar to electron-dense collars that were observed many years earlier in neurons of Drosophila shibire mutants (6). Recent experiments demonstrated the ability of brain cytosol and purified dynamin to form lipid tubules (7,8) Purified dynamin can sever the lipid tubules by GTP-dependent constriction (8).…”

Dynamin Assembles into Spirals under Physiological Salt Conditions upon the Addition of GDP and γ-Phosphate Analogues

“…This work revealed that the basic assembly unit of dynamin, a tetramer, can polymerize into rings or spirals that resemble the collar structures seen at the necks of coated pits in the shibire mutant nerve termini (4,25). Dynamin spirals also resemble the striation pattern seen on long tubules, which originate from the plasma membrane and end in a coated pit.…”

A role of amphiphysin in synaptic vesicle endocytosis suggested by its binding to dynamin in nerve terminals.

“…Temperature-sensitive mutations of the dynamin gene (shibire) in Drosophila cause a selective arrest of the synaptic vesicle cycle at the stage of invaginated plasmalemmal pits (3)(4)(5)(6), and transfection of dominant negative dynamin mutants in fibroblastic cells blocks clathrin-mediated endocytosis (7,8). Recent studies have shown that dynamin forms rings at the neck of invaginated clathrin-coated vesicles and suggested that a conformational change of the rings which correlates with GTP hydrolysis leads to vesicle fission (9,10).…”