RESUMEN
The Cytomatrix Assembled at the active Zone (CAZ) of a presynaptic terminal displays electron-dense appearance and defines the center of the synaptic vesicle release. The protein constituents of CAZ are multiple-domain scaffolds that interact extensively with each other and also with an ensemble of synaptic vesicle proteins to ensure docking, fusion, and recycling. Reflecting the central roles of the active zone in synaptic transmission, CAZ proteins are highly conserved throughout evolution. As the nervous system increases complexity and diversity in types of neurons and synapses, CAZ proteins expand in the number of gene and protein isoforms and interacting partners. This chapter summarizes the discovery of the core CAZ proteins and current knowledge of their functions.
Asunto(s)
Sinapsis , Vesículas Sinápticas , Humanos , Neuronas , Transmisión SinápticaRESUMEN
Brain's functions, such as memory and learning, rely on synapses that are highly specialized cellular junctions connecting neurons. Functional synapses orchestrate the assembly of ion channels, receptors, enzymes, and scaffold proteins in both pre- and post-synapse. Liprin-α proteins are master scaffolds in synapses and coordinate various synaptic proteins to assemble large protein complexes. The functions of liprin-αs in synapse formation have been largely uncovered by genetic studies in diverse model systems. Recently, emerging structural and biochemical studies on liprin-α proteins and their binding partners begin to unveil the molecular basis of the synaptic assembly. This review summarizes the recent structural findings on liprin-αs, proposes the assembly mechanism of liprin-α-mediated complexes, and discusses the liprin-α-organized assemblies in the regulation of synapse formation and function.
RESUMEN
Synaptic scaffold proteins (e.g., liprin-α, ELKS, RIM, and RIM-BP) orchestrate ion channels, receptors, and enzymes at presynaptic terminals to form active zones for neurotransmitter release. The underlying mechanism of the active zone assembly remains elusive. Here, we report that liprin-α proteins have the potential to oligomerize through the N-terminal coiled-coil region. Our structural and biochemical characterizations reveal that a gain-of-function mutation promotes the self-assembly of the coiled coils in liprin-α2 by disrupting intramolecular interactions and promoting intermolecular interactions. By enabling multivalent interactions with ELKS proteins, the oligomerized coiled-coil region of liprin-α2 enhances the phase separation of the ELKS N-terminal segment. We further show that liprin-α2, by regulating the interplay between two phase separations of ELKS and RIM/RIM-BP, controls the protein distributions. These results imply that the complicated protein-protein interactions allow liprin-α to function with the active zone scaffolds and compartmentalize protein assemblies to achieve comprehensive functions in the active zone.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/química , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas de la Membrana/química , Proteínas de la Membrana/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Multimerización de Proteína , Sinapsis/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas de Unión al GTP/metabolismo , Células HEK293 , Humanos , Proteínas de la Membrana/genética , Mutación/genética , Estructura Secundaria de ProteínaRESUMEN
In neurons, a single motor (dynein) transports large organelles as well as synaptic and dense core vesicles toward microtubule minus ends; however, it is unclear why dynein appears more active on organelles, which are generally excluded from mature axons, than on synaptic and dense core vesicles, which are maintained at high levels. Recent studies in Zebrafish and Caenorhabditis elegans have shown that JIP3 promotes dynein-mediated retrograde transport to clear some organelles (lysosomes, early endosomes, and Golgi) from axons and prevent their potentially harmful accumulation in presynaptic regions. A JIP3 mutant suppressor screen in C. elegans revealed that JIP3 promotes the clearance of organelles from axons by blocking the action of the CSS system (Cdk5, SAD Kinase, SYD-2/Liprin). A synthesis of results in vertebrates with the new findings suggests that JIP3 blocks the CSS system from disrupting the connection between dynein and organelles. Most components of the CSS system are enriched at presynaptic active zones where they normally contribute to maintaining optimal levels of captured synaptic and dense core vesicles, in part by inhibiting dynein transport. The JIP3-CSS system model explains how neurons selectively regulate a single minus-end motor to exclude specific classes of organelles from axons, while at the same time ensuring optimal levels of synaptic and dense core vesicles.
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Transporte Axonal/fisiología , Neuronas/metabolismo , Animales , HumanosRESUMEN
Kinesin-3 UNC-104(KIF1A) is the major axonal transporter of synaptic vesicles. Employing yeast two-hybrid and co-immunoprecipitation (Co-IP) assays, we characterized a LIN-2(CASK) binding site overlapping with that of reported UNC-104 activator protein SYD-2(Liprin-α) on the motor's stalk domain. We identified the L27 and GUK domains of LIN-2 to be the most critical interaction domains for UNC-104. Further, we demonstrated that the L27 domain interacts with the sterile alpha motifs (SAM) domains of SYD-2, while the GUK domain is able to interact with both the coiled coils and SAM domains of SYD-2. LIN-2 and SYD-2 colocalize in Caenorhabditis elegans neurons and display interactions in bimolecular fluorescence complementation (BiFC) assays. UNC-104 motor motility and Synaptobrevin-1 (SNB-1) cargo transport are largely diminished in neurons of LIN-2 knockout worms, which cannot be compensated by overexpressing SYD-2. The absence of the motor-activating function of LIN-2 results in increased motor clustering along axons, thus retaining SNB-1 cargo in cell bodies. LIN-2 and SYD-2 both positively affect the velocity of UNC-104, however, only LIN-2 is able to efficiently elevate the motor's run lengths. From our study, we conclude that LIN-2 and SYD-2 act in a functional complex to regulate the motor with LIN-2 being the more prominent activator.
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Transporte Axonal/fisiología , Axones/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Movimiento Celular/fisiología , Proteínas del Helminto/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Vesículas Sinápticas/metabolismo , Animales , Animales Modificados Genéticamente/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas del Tejido Nervioso/genética , Fosfoproteínas/metabolismoRESUMEN
Since Caenorhabditis elegans was chosen as a model organism by Sydney Brenner in 1960's, genetic studies in this organism have been instrumental in discovering the function of genes and in deciphering molecular signaling network. The small size of the organism and the simple nervous system enable the complete reconstruction of the first connectome. The stereotypic developmental program and the anatomical reproducibility of synaptic connections provide a blueprint to dissect the mechanisms underlying synapse formation. Recent technological innovation using laser surgery of single axons and in vivo imaging has also made C. elegans a new model for axon regeneration. Importantly, genes regulating synaptogenesis and axon regeneration are highly conserved in function across animal phyla. This mini-review will summarize the main approaches and the key findings in understanding the mechanisms underlying the development and maintenance of the nervous system. The impact of such findings underscores the awesome power of C. elegans genetics.
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Axones/fisiología , Caenorhabditis elegans/fisiología , Regeneración Nerviosa/fisiología , Sinapsis/fisiología , Animales , Axones/metabolismo , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Modelos Neurológicos , Mutación , Regeneración Nerviosa/genética , Transducción de Señal/genética , Sinapsis/metabolismoRESUMEN
SYD-2/liprin-α is a multi-domain protein that associates with and recruits multiple active zone molecules to form presynaptic specializations. Given SYD-2's critical role in synapse formation, its synaptogenic ability is likely tightly regulated. However, mechanisms that regulate SYD-2 function are poorly understood. In this study, we provide evidence that SYD-2's function may be regulated by interactions between its coiled-coil (CC) domains and sterile α-motif (SAM) domains. We show that the N-terminal CC domains are necessary and sufficient to assemble functional synapses while C-terminal SAM domains are not, suggesting that the CC domains are responsible for the synaptogenic activity of SYD-2. Surprisingly, syd-2 alleles with single amino acid mutations in the SAM domain show strong loss of function phenotypes, suggesting that SAM domains also play an important role in SYD-2's function. A previously characterized syd-2 gain-of-function mutation within the CC domains is epistatic to the loss-of-function mutations in the SAM domain. In addition, yeast two-hybrid analysis showed interactions between the CC and SAM domains. Thus, the data is consistent with a model where the SAM domains regulate the CC domain-dependent synaptogenic activity of SYD-2. Taken together, our study provides new mechanistic insights into how SYD-2's activity may be modulated to regulate synapse formation during development.