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Doc2 supports spontaneous synaptic transmission by a Ca(2+)-independent mechanism.

ABSTRACT: Two families of Ca(2+)-binding proteins have been proposed as Ca(2+) sensors for spontaneous release: synaptotagmins and Doc2s, with the intriguing possibility that Doc2s may represent high-affinity Ca(2+) sensors that are activated by deletion of synaptotagmins, thereby accounting for the increased spontaneous release in synaptotagmin-deficient synapses. Here, we use an shRNA-dependent quadruple knockdown of all four Ca(2+)-binding proteins of the Doc2 family to confirm that Doc2-deficient synapses exhibit a marked decrease in the frequency of spontaneous release events. Knockdown of Doc2s in synaptotagmin-1-deficient synapses, however, failed to reduce either the increased spontaneous release or the decreased evoked release of these synapses, suggesting that Doc2s do not constitute Ca(2+) sensors for asynchronous release. Moreover, rescue experiments revealed that the decrease in spontaneous release induced by the Doc2 knockdown in wild-type synapses is fully reversed by mutant Doc2B lacking Ca(2+)-binding sites. Thus, our data suggest that Doc2s are modulators of spontaneous synaptic transmission that act by a Ca(2+)-independent mechanism.

SUBMITTER: Pang ZP

PROVIDER: S-EPMC3102832 | biostudies-literature | 2011 Apr

REPOSITORIES: biostudies-literature

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Doc2 supports spontaneous synaptic transmission by a Ca(2+)-independent mechanism.

Pang Zhiping P ZP Bacaj Taulant T Yang Xiaofei X Zhou Peng P Xu Wei W Südhof Thomas C TC

Neuron 20110401 2

Two families of Ca(2+)-binding proteins have been proposed as Ca(2+) sensors for spontaneous release: synaptotagmins and Doc2s, with the intriguing possibility that Doc2s may represent high-affinity Ca(2+) sensors that are activated by deletion of synaptotagmins, thereby accounting for the increased spontaneous release in synaptotagmin-deficient synapses. Here, we use an shRNA-dependent quadruple knockdown of all four Ca(2+)-binding proteins of the Doc2 family to confirm that Doc2-deficient syna ...[more]

PMID: 21521611

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Project description:Using postsynaptically tethered calcium sensor GCaMP, we investigated spontaneous synaptic transmission at individual active zones (AZs) at the Drosophila (both sexes) neuromuscular junction. Optical monitoring of GCaMP events coupled with focal electrical recordings of synaptic currents revealed "hot spots" of spontaneous transmission, which corresponded to transient states of elevated activity at selected AZs. The elevated spontaneous activity had two temporal components, one at a timescale of minutes and the other at a subsecond timescale. We developed a three-state model of AZ preparedness for spontaneous transmission and performed Monte Carlo simulations of the release process, which produced an accurate quantitative description of the variability and time course of spontaneous transmission at individual AZs. To investigate the mechanisms of elevated activity, we first focused on the protein complexin, which binds the SNARE protein complex and serves to clamp spontaneous fusion. Overexpression of Drosophila complexin largely abolished the high-activity states of AZs, while complexin deletion drastically promoted it. A mutation in the SNARE protein Syntaxin-1A had an effect similar to complexin deficiency, promoting the high-activity state. We next tested how presynaptic Ca2+ transients affect the states of elevated activity at individual AZs. We either blocked or promoted Ca2+ influx pharmacologically, and also promoted Ca2+ release from internal stores. These experiments coupled with computations revealed that Ca2+ transients can trigger bursts of spontaneous events from individual AZs or AZ clusters at a subsecond timescale. Together, our results demonstrated that spontaneous transmission is highly heterogeneous, with transient hot spots being regulated by the SNARE machinery and Ca2+ SIGNIFICANCE STATEMENT Spontaneous synaptic transmission is a vital component of neuronal communication, since it regulates the neuronal development and plasticity. Our study demonstrated that spontaneous transmission is highly heterogeneous and that nerve terminals create transient "hot spots" of spontaneous release of neuronal transmitters. We show that these hot spots are regulated by the protein machinery mediating the release process and by calcium ions. These results contribute to our understanding of spontaneous synaptic transmission as a dynamic, plastic, and tightly regulated signaling mechanism and unravel fundamental biophysical properties of neuronal communication.

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Doc2 supports spontaneous synaptic transmission by a Ca(2+)-independent mechanism.

Publications

Doc2 supports spontaneous synaptic transmission by a Ca(2+)-independent mechanism.

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