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The Liprin homology domain is essential for the homomeric interaction of SYD-2/Liprin-α protein in presynaptic assembly.

ABSTRACT: Synapses are asymmetric structures that are specialized for neuronal signal transduction. A unique set of proteins is present at the presynaptic active zone, which is a core structure essential for neurotransmitter release. In Caenorhabditis elegans HSN neurons, SYD-2, a Liprin-α family protein, acts together with a GAP protein SYD-1 to promote presynaptic assembly. Previous studies have shown that elevating the activity of syd-2 can bypass the requirement of syd-1. Liprin-α proteins are composed of coiled-coil-rich regions in the N-terminal half, which mediate interactions with adapter proteins at the presynaptic active zone, and three SAM domains in the C terminus, which bind proteins such as LAR receptor tyrosine phosphatase. To address the molecular mechanism by which SYD-2 activity is regulated, we performed structure-function studies. By monitoring the ability of SYD-2 transgenes to rescue syd-2(lf) and to suppress syd-1(lf) phenotypes in HSN neuron synapses, we identified the N-terminal half of SYD-2 as minimally required for rescuing syd-2(lf) phenotypes. A highly conserved short coiled-coil segment named Liprin Homology 1 (LH1) domain is both necessary and sufficient to suppress syd-1(lf) defects. We show that the LH1 domain forms a dimer and promotes further oligomerization and/or complex formation of SYD-2/Liprin-α proteins. The role of the LH1 domain in presynaptic assembly can be partially complemented by artificial dimerization. These findings suggest a model by which the self-assembly of SYD-2/Liprin-α proteins mediated by the coiled-coil LH1 domain is one of the key steps to the accumulation of presynaptic components at nascent synaptic junctions.

SUBMITTER: Taru H

PROVIDER: S-EPMC3500560 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Presynaptic active zone proteins couple calcium influx with synaptic vesicle exocytosis. However, the control of presynaptic calcium channel localization by active zone proteins is not completely understood. In a Caenorhabditis elegans (C. elegans) forward genetic screen, we find that UNC-10/RIM (Rab3-interacting molecule) and SYD-2/Liprin-α regulate presynaptic localization of UNC-2, the CaV2 channel ortholog. We further quantitatively analyzed live animals using endogenously GFP-tagged UNC-2 and active zone components. Consistent with the interaction between RIM and CaV2 in mammals, the intensity and number of UNC-2 channel puncta at presynaptic terminals were greatly reduced in unc-10 mutant animals. To understand how SYD-2 regulates presynaptic UNC-2 channel localization, we analyzed presynaptic localization of endogenous SYD-2, UNC-10, RIMB-1/RIM-BP (RIM binding protein), and ELKS-1. Our analysis revealed that although SYD-2 is the most critical for active zone assembly, loss of SYD-2 function does not completely abolish presynaptic localization of UNC-10, RIMB-1, and ELKS-1, suggesting an existence of SYD-2-independent active zone assembly. UNC-2 localization analysis in double and triple mutants of active zone components show that SYD-2 promotes UNC-2 localization by partially controlling UNC-10 localization, and ELKS-1 and RIMB-1 also contribute to UNC-2 channel localization. In addition, we find that core active zone proteins are unequal in their abundance. Although the abundance of UNC-10 at the active zone is comparable to UNC-2, SYD-2 and ELKS-1 are twice more and RIMB-1 four times more abundant than UNC-2. Together our data show that UNC-10, SYD-2, RIMB-1, and ELKS-1 control presynaptic UNC-2 channel localization in redundant yet distinct manners.SIGNIFICANCE STATEMENT Precise control of neurotransmission is dependent on the tight coupling of the calcium influx through voltage-gated calcium channels (VGCCs) to the exocytosis machinery at the presynaptic active zones. However, how these VGCCs are tethered to the active zone is incompletely understood. To understand the mechanism of presynaptic VGCC localization, we performed a C. elegans forward genetic screen and quantitatively analyzed endogenous active zones and presynaptic VGCCs. In addition to RIM, our study finds that SYD-2/Liprin-α is critical for presynaptic localization of VGCCs. Yet, the loss of SYD-2, a core active zone scaffolding protein, does not completely abolish the presynaptic localization of the VGCC, showing that the active zone is a resilient structure assembled by redundant mechanisms.

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The Liprin homology domain is essential for the homomeric interaction of SYD-2/Liprin-α protein in presynaptic assembly.

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