Biomedical subjects

Tong-Wey Koh

Publications and source records attributed to Tong-Wey Koh.

5 recordsLinked to original sources

PubMed2006

Activity-independent prespecification of synaptic partners in the visual map of Drosophila.

Specifying synaptic partners and regulating synaptic numbers are at least partly activity-dependent processes during visual map formation in all systems investigated to date . In Drosophila, six photoreceptors that view the same point in visual space have to be sorted into synaptic modules called cartridges in order to form a visuotopically correct map . Synapse numbers per photoreceptor terminal and cartridge are both precisely regulated . However, it is unknown whether an activity-dependent mechanism or a genetically encoded developmental program regulates synapse numbers. We performed a large-scale quantitative ultrastructural analysis of photoreceptor synapses in mutants affecting the generation of electrical potentials (norpA, trp;trpl), neurotransmitter release (hdc, syt), vesicle endocytosis (synj), the trafficking of specific guidance molecules during photoreceptor targeting (sec15), a specific guidance receptor required for visual map formation (Dlar), and 57 other novel synaptic mutants affecting 43 genes. Remarkably, in all these mutants, individual photoreceptors form the correct number of synapses per presynaptic terminal independently of cartridge composition. Hence, our data show that each photoreceptor forms a precise and constant number of afferent synapses independently of neuronal activity and partner accuracy. Our data suggest cell-autonomous control of synapse numbers as part of a developmental program of activity-independent steps that lead to a "hard-wired" visual map in the fly brain.

Animals↗

PubMed2005

Synaptic mitochondria are critical for mobilization of reserve pool vesicles at Drosophila neuromuscular junctions.

In a forward screen for genes affecting neurotransmission in Drosophila, we identified mutations in dynamin-related protein (drp1). DRP1 is required for proper cellular distribution of mitochondria, and in mutant neurons, mitochondria are largely absent from synapses, thus providing a genetic tool to assess the role of mitochondria at synapses. Although resting Ca2+ is elevated at drp1 NMJs, basal synaptic properties are barely affected. However, during intense stimulation, mutants fail to maintain normal neurotransmission. Surprisingly, FM1-43 labeling indicates normal exo- and endocytosis, but a specific inability to mobilize reserve pool vesicles, which is partially rescued by exogenous ATP. Using a variety of drugs, we provide evidence that reserve pool recruitment depends on mitochondrial ATP production downstream of PKA signaling and that mitochondrial ATP limits myosin-propelled mobilization of reserve pool vesicles. Our data suggest a specific role for mitochondria in regulating synaptic strength.

Animals↗

PubMed2004

Dap160/intersectin acts as a stabilizing scaffold required for synaptic development and vesicle endocytosis.

We describe the isolation of mutations in dynamin-associated protein 160 kDa (dap160), the Drosophila homolog of intersectin, a putative adaptor for proteins involved in endocytosis, cytoskeletal regulation, and signaling. We show that partial loss-of-function mutants display temperature-sensitive (ts) paralysis, whereas null mutants show ts defects in endocytosis. Loss-of-function mutants exhibit bouton overgrowth at larval neuromuscular junctions (NMJs), but evoked neurotransmission is normal. Mutant NMJs show a mild endocytic defect at 22 degrees C, which is strongly enhanced at 34 degrees C. The levels of dynamin, synaptojanin and endophilin are severely reduced in dap160 mutant NMJs, suggesting that Dap160 serves to stabilize an endocytic macromolecular complex. Electron microscopy reveals fewer vesicles, aberrant large vesicles, and an accumulation of endocytic intermediates at active and periactive zones in mutant terminals. Our data suggest that Dap160, like dynamin, is involved in synaptic vesicle retrieval at active and periactive zones.

Adaptor Proteins, Vesicular Transport↗

PubMed2003

Synaptojanin is recruited by endophilin to promote synaptic vesicle uncoating.

We describe the isolation and characterization of Drosophila synaptojanin (synj) mutants. synj encodes a phosphatidylinositol phosphatase involved in clathrin-mediated endocytosis. We show that Synj is specifically localized to presynaptic terminals and is associated with synaptic vesicles. The electrophysiological and ultrastructural defects observed in synj mutants are strikingly similar to those found in endophilin mutants, and Synj and Endo colocalize and interact biochemically. Moreover, synj; endo double mutant synaptic terminals exhibit properties that are very similar to terminals of each single mutant, and overexpression of Endophilin can partially rescue the functional defects in partial loss-of-function synj mutants. Interestingly, Synj is mislocalized and destabilized at synapses devoid of Endophilin, suggesting that Endophilin recruits and stabilizes Synj on newly formed vesicles to promote vesicle uncoating. Our data also provide further evidence that kiss-and-run is able to maintain neurotransmitter release when synapses are not extensively challenged.

Adaptor Proteins, Signal Transducing↗

PubMed2003

Mapping Drosophila mutations with molecularly defined P element insertions.

The isolation of chemically induced mutations in forward genetic screens is one of the hallmarks of Drosophila genetics. However, mapping the corresponding loci and identifying the molecular lesions associated with these mutations are often difficult and labor-intensive. Two mapping methods are most often used in flies: meiotic recombination mapping with marked chromosomes and deficiency mapping. The availability of the fly genome sequence allows the establishment and usage of molecular markers. Single-nucleotide polymorphisms have therefore recently been used to map several genes. Here we show that thousands of molecularly mapped P element insertions in fly strains that are publicly available provide a powerful alternative method to single-nucleotide polymorphism mapping. We present a strategy that allows mapping of lethal mutations, as well as viable mutations with visible phenotypes, with minimal resources. The most important unknown in using recombination rates to map at high resolution is how accurately recombination data correlate with molecular maps in small intervals. We therefore surveyed distortions of recombination rates in intervals <500 kb. We document the extent of distortions between the recombination and molecular maps and describe the required steps to map with an accuracy of <50 kb. Finally, we describe a recently developed method to determine molecular lesions in 50-kb intervals by using a heteroduplex DNA mutation detection system. Our data show that this mapping approach is inexpensive, efficient, and precise, and that it significantly broadens the application of P elements in Drosophila.

Animals↗