Presynaptic terminals contain several specialized compartments, which have been described by electron microscopy. We show in an identified Drosophila neuromuscular synapse that of these compartments-synaptic vesicle clusters, presynaptic plasma membrane, cytosol, and axonal cytoskeleton-labeled specific reagents may be resolved from one another laser scanning confocal Using a panel compartment-specific markers shibire(ts1) mutants to trap intermediate stage synaptic recycling, we examined...

We have developed a primary cell culture system derived from embryonic and larval stages of Drosophila. This allows for high-resolution imaging genetic analyses endocytic processes. Here, we investigated pathways three types molecules: an endogenous receptor that binds anionic ligands (ALs), glycosylphosphatidylinositol (GPI)-anchored protein (GPI-AP), markers the fluid phase in hemocytes. find AL-binding (ALBR) is internalized into Rab5-positive endosomes, whereas major portion taken up...

Endosomal degradation is severely impaired in primary hemocytes from larvae of eye color mutants Drosophila. Using high resolution imaging and immunofluorescence microscopy these cells, products genes, deep-orange (dor) carnation (car), are localized to large multivesicular Rab7-positive late endosomes containing Golgi-derived enzymes. These structures mature into small sized Dor-negative, Car-positive structures, which subsequently fuse form tubular lysosomes. Defective endosomal mutant...

In an attempt to identify genes that control or encode the targets of general anesthetics, we have chemically mutagenized fruit flies and selected four lines show abnormal response volatile anesthetic halothane. Specifically, about 2-fold higher concentrations halothane are required induce loss motor in mutant flies. Fine mapping two isolates indicates they alter a previously uncharacterized gene Drosophila. absence these mutants display alterations behavior imply changes adult larval...

Abstract Individual contributions made by different calcium release and sequestration mechanisms to various aspects of excitable cell physiology are incompletely understood. SERCA, a sarco-endoplasmic reticulum ATPase, being the main agent for uptake into ER, plays central role in this process. By isolation extensive characterization conditional mutations Drosophila SERCA gene, we describe novel roles key protein neuromuscular enable genetic analysis function. At motor nerve terminals,...

D-serine is an important signalling molecule, which activates N-methyl D-aspartate receptors (NMDARs) in conjunction with its fellow co-agonist, the neurotransmitter glutamate. Despite involvement plasticity and memory related to excitatory synapses, cellular source sink remain a question. We hypothesise that astrocytes, type of glial cell surrounds are likely candidates control extracellular concentration D-Serine by removing it from synaptic space. Using situ patch clamp recordings...

Syndapins belong to the F-BAR domain protein family whose predicted functions in membrane tubulation remain poorly studied vivo. At Drosophila neuromuscular junctions, syndapin is associated predominantly with a tubulolamellar postsynaptic system known as subsynaptic reticulum (SSR). We show that overexpression greatly expands this system. Syndapin can expand SSR absence of dPAK and Dlg, two regulators development. Syndapin's N-terminal domain, required for cultured cells, expansion....

The shibirets2 mutation of Drosophila melanogaster causes a temperature sensitive inhibition endocytosis; this in turn leads to synaptic-vesicle depletion and consequent paralysis. Heat-pulses delivered during development individuals affect the morphology number adult structures. A simple screening protocol has been used isolate several mutations that partially suppress temperature-sensitive paralytic phenotype mutant animals. All these very tightly linked shibire are likely be second site...

T-1-family conotoxins belong to the T-superfamily and are composed of 10−17 amino acids. They share a common cysteine framework disulfide connectivity exhibit unusual posttranslational modifications, such as tryptophan bromination, glutamic acid carboxylation, threonine glycosylation. We have isolated characterized novel peptide, Mo1274, containing 11 acids, that shows same pattern, -CC-CC, linkage those members. The complete sequence, GNWCCSARVCC, in which W denotes bromotryptophan, was...

In metazoans, lysosomes are characterized by a unique tubular morphology, acidic pH, and specific membrane protein (LAMP) lipid (cholesterol) composition as well soluble (hydrolases) composition. Here we show that perturbation to the eye-color gene, light, results in impaired lysosomal acidification, sterol accumulation, altered endosomal morphology compromised degradation. We find Drosophila homologue of Vps41, Light, regulates fusion subset biosynthetic carriers containing characteristic...

At an initial step during synaptic vesicle recycling, dynamin and adaptor proteins mediate the endocytosis of components from plasma membrane. StonedA stonedB, novel encoded by a single Drosophila gene, have predicted structural similarities to adaptors other implicated in endocytosis. Here, we test possible roles stoned internalization via analyses third instar larval neuromuscular synapses two (stn) mutants, stn(ts) stn(8P1). Both mutations reduce presynaptic levels stonedA although has...

The N-ethylmaleimide-sensitive fusion protein (NSF) is a cytoplasmic implicated in the of intracellular transport vesicles with their target membranes. NSF thought to function essentially all types vesicles, including endoplasmic reticulum, Golgi, and endocytic as well secretory undergoing regulated (for review see Rothman, J. E.(1994) Nature 372, 55-63). However, little [Medline] experimental evidence exists address possibility that organisms might have multiple proteins serving distinct...

Studies in cell-free systems and the lamprey giant synapse have implicated crucial roles for amphiphysin endophilin synaptic transmission. However, null mutants at <i>amphiphysin</i>locus of <i>Drosophila</i> are viable no demonstrable vesicle-recycling defect. This has necessitated a re-examination role Src homology 3 domain-containing proteins vesicle recycling. In this report, we show that endophilin-deficient eye clones an altered electroretinogram. A characteristic defect is its...

Abstract The shibire locus of Drosophila melanogaster encodes dynamin, a GTPase required for the fission endocytic vesicles from plasma membrane. Biochemical studies indicate that mammalian dynamin is part complex containing multiple subunits and other polypeptides. To gain insight into sequences critical its function, we have characterized in detail collection conditional lethal shi alleles. We describe probable null allele show properties are distinct those two classes alleles (termed I...

We have quantitated the response of D. melanogaster to general anesthetics with a device, inebriometer, that assays fly's geotactic and postural behavior. When alleles several loci encode or regulate subunits ion channels were compared control stocks, channel mutants clearly increased anesthetic sensitivity Drosophila. The effects specific in that: a) for alleles, genetic tests indicated anesthesia phenotype was due mutation not extraneous differences between stocks; b) given often affected...

A 26 residue peptide (Am 2766) with the sequence CKQAGESCDIFSQNCCVG‐TCAFICIE‐NH 2 has been isolated and purified from venom of molluscivorous snail, Conus amadis , collected off southeastern coast India. Chemical modification mass spectrometric studies establish that Am 2766 three disulfide bridges. C‐terminal amidation demonstrated by measurements on fragments obtained proteolysis. Sequence alignments belongs to δ‐conotoxin family. inhibits decay sodium current in brain rNav1.2a...

Abstract Rapidly reversible, temperature-sensitive (ts) paralytic mutants of Drosophila have been useful in delineating immediate vivo functions molecules involved synaptic transmission. Here we report isolation and characterization orangi (org), an enhancer shibire (shi), a ts mutant dynamin. org is allele the stress sensitive B (sesB) locus that encodes mitochondrial adenine nucleotide translocase (ANT) results unique behavior accompanied by complete loss transmission visual system....

Mitochondria are the primary source of ATP needed for steps synaptic vesicle cycle. Dynamin‐related protein (DRP) is involved in fission mitochondria and peroxisomes. To assess role function, we characterized a Drosophila DRP mutant combination that shows an acute temperature‐sensitive paralysis. Sequencing reveals single amino acid change guanosine triphosphate hydrolysing domain (GTPase domain) DRP. The these mutants remarkably elongated, suggesting mitochondrial . There loss neuronal...