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R Grace Zhai

Publications and source records attributed to R Grace Zhai.

10 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↗
PubMed2006

Drosophila NMNAT maintains neural integrity independent of its NAD synthesis activity.

Wallerian degeneration refers to a loss of the distal part of an axon after nerve injury. Wallerian degeneration slow (Wld(s)) mice overexpress a chimeric protein containing the NAD synthase NMNAT (nicotinamide mononucleotide adenylyltransferase 1) and exhibit a delay in axonal degeneration. Currently, conflicting evidence raises questions as to whether NMNAT is the protecting factor and whether its enzymatic activity is required for such a possible function. Importantly, the link between nmnat and axon degeneration is at present solely based on overexpression studies of enzymatically active protein. Here we use the visual system of Drosophila as a model system to address these issues. We have isolated the first nmnat mutations in a multicellular organism in a forward genetic screen for synapse malfunction in Drosophila. Loss of nmnat causes a rapid and severe neurodegeneration that can be attenuated by blocking neuronal activity. Furthermore, in vivo neuronal expression of mutated nmnat shows that enzymatically inactive NMNAT protein retains strong neuroprotective effects and rescues the degeneration phenotype caused by loss of nmnat. Our data indicate an NAD-independent requirement of NMNAT for maintaining neuronal integrity that can be exploited to protect neurons from neuronal activity-induced degeneration by overexpression of the protein.

Animals↗
PubMed2005

The v-ATPase V0 subunit a1 is required for a late step in synaptic vesicle exocytosis in Drosophila.

The V(0) complex forms the proteolipid pore of an ATPase that acidifies vesicles. In addition, an independent function in membrane fusion has been proposed largely based on yeast vacuolar fusion experiments. We have isolated mutations in the largest V(0) component vha100-1 in flies in an unbiased genetic screen for synaptic malfunction. The protein is only required in neurons, colocalizes with markers for synaptic vesicles as well as active zones, and interacts with t-SNAREs. Loss of vha100-1 leads to vesicle accumulation in synaptic terminals, suggesting a deficit in release. The amplitude of spontaneous release events and release with hypertonic stimulation indicate normal levels of neurotransmitter loading, yet mutant embryos display severe defects in evoked synaptic transmission and FM1-43 uptake. Our data suggest that Vha100-1 functions downstream of SNAREs in synaptic vesicle fusion.

Animals↗
PubMed2005

Mutations in Drosophila sec15 reveal a function in neuronal targeting for a subset of exocyst components.

The exocyst is a complex of proteins originally identified in yeast that has been implicated in polarized secretion. Components of the exocyst have been implicated in neurite outgrowth, cell polarity, and cell viability. We have isolated an exocyst component, sec15, in a screen for genes required for synaptic specificity. Loss of sec15 causes a targeting defect of photoreceptors that coincides with mislocalization of specific cell adhesion and signaling molecules. Additionally, sec15 mutant neurons fail to localize other exocyst members like Sec5 and Sec8, but not Sec6, to neuronal terminals. However, loss of sec15 does not cause cell lethality in contrast to loss of sec5 or sec6. Our data suggest a role of Sec15 in an exocyst-like subcomplex for the targeting and subcellular distribution of specific proteins. The data also show that functions of other exocyst components persist in the absence of sec15, suggesting that different exocyst components have separable functions.

Amino Acid Sequence↗
PubMed2004

The architecture of the active zone in the presynaptic nerve terminal.

Active zones are highly specialized sites for release of neurotransmitter from presynaptic nerve terminals. The architecture of the active zone is exquisitely designed to facilitate the regulated tethering, docking, and fusing of the synaptic vesicles with the plasma membrane. Here we present our view of the structural and molecular organization of active zones across species and propose that all active zones are organized according to a common principle in which the structural differences correlate with the kinetics of transmitter release.

Animals↗
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↗
PubMed2003

Unitary assembly of presynaptic active zones from Piccolo-Bassoon transport vesicles.

Recent studies indicate that active zones (AZs)-sites of neurotransmitter release-may be assembled from preassembled AZ precursor vesicles inserted into the presynaptic plasma membrane. Here we report that one putative AZ precursor vesicle of CNS synapses-the Piccolo-Bassoon transport vesicle (PTV)-carries a comprehensive set of AZ proteins genetically and functionally coupled to synaptic vesicle exocytosis. Time-lapse imaging reveals that PTVs are highly mobile, consistent with a role in intracellular transport. Quantitative analysis reveals that the Bassoon, Piccolo, and RIM content of individual PTVs is, on average, half of that of individual presynaptic boutons and shows that the synaptic content of these molecules can be quantitatively accounted for by incorporation of integer numbers (typically two to three) of PTVs into presynaptic membranes. These findings suggest that AZs are assembled from unitary amounts of AZ material carried on PTVs.

Animals↗
PubMed2002

Molecular mechanisms of CNS synaptogenesis.

Synapses of the mammalian CNS are asymmetric sites of cell-cell adhesion between nerve cells. They are designed to mediate the rapid and efficient transmission of signals from the presynaptic bouton of one neuron to the postsynaptic plasma membrane of a second neuron. Significant progress has been made in the characterization of the structural, functional and developmental assembly of CNS synapses. Recent progress has been made in understanding the molecular and cellular mechanisms that underlie synaptogenesis, in particular that of glutamatergic synapses of the CNS.

Animals↗