RIM Promotes Calcium Channel Accumulation at Active Zones of theDrosophilaNeuromuscular Junction

Abstract

Summary Synaptic communication requires the controlled release of synaptic vesicles from presynaptic axon terminals. Release efficacy is regulated by the many proteins that comprise the presynaptic release apparatus, including Ca2+ channels and proteins that influence Ca2+ channel accumulation at release sites. Here we identify Drosophila RIM and demonstrate that it localizes to active zones at the larval neuromuscular junction. In Drosophila RIM mutants, there is a large decrease in evoked synaptic transmission, due to a significant reduction in both the clustering of Ca2+ channels and the size of the readily releasable pool of synaptic vesicles at active zones. Hence, RIM plays an evolutionarily conserved role in regulating synaptic calcium channel localization and readily releasable pool size. Since RIM has traditionally been studied as an effector of Rab3 function, we investigate whether RIM is involved in the newly identified function of Rab3 in the distribution of presynaptic release machinery components across release sites. Bruchpilot (Brp), an essential component of the active zone cytomatrix T bar, is unaffected by RIM disruption, indicating that Brp localization and distribution across active zones does not require wild type RIM. In addition, larvae containing mutations in both RIM and rab3 have reduced Ca2+ channel levels and a Brp distribution that is very similar to that of the rab3 single mutant, indicating that RIM functions to regulate Ca2+ channel accumulation but is not a Rab3 effector for release machinery distribution across release sites.

RIM-binding protein 2 regulates release probability by fine-tuning calcium channel localization at murine hippocampal synapses

“…For both hippocampal and NMJ synapses, however, the RIM/RIM-BP complex is crucial for precise Ca V localization at the AZ and expression of STP and/or long-term plasticity. The comparison of KO phenotypes suggests that in Drosophila, DRBP plays a more pivotal role (6,27), whereas judged from single gene KOs at mammalian synapses, RIM is functionally most important (1,2,8,(28)(29)(30). Still, the importance of RIM-BPs for STP is conserved between flies and mammals (6,8,31), consistent with the highly conserved molecular interactions between RIM-BPs, Ca V s, and RIM (2,4,6).…”

Transmission, Development, and Plasticity of Synapses

“…DiAntonio and colleagues (Graf et al 2012) have characterized several mutations that exhibit this phenotype, highlighting distinct pathways that regulate presynaptic development. A mutation in runningunapposed (rup), which encodes Drosophila rab3, was identified in a screen for AZ mutants.…”

“…These changes appear to be separable; for example, mutants for rim (rab3-interacting molecule) block homeostatic changes in RRP size, but exhibit normal Ca 2+ influx . RIM localizes to AZs and regulates Cac localization (Graf et al 2012), but how RIM contributes to RRP regulation during homeostatic plasticity is unclear. As Rab3-GAP (Rab3 GTPase-activating protein) is also required presynaptically for synaptic homeostasis (Müller et al 2011), there is mounting evidence for a Rab3 signaling module regulating this process.…”

The Role of Calcium Channels in Epilepsy

“…Their localization within the active zones of presynaptic nerve terminals (Westenbroek et al 1995) enables the coupling of calcium influx to vesicular exocytosis through a direct interaction with the SNARE protein complex comprising syntaxin, SNAP-25, and VAMP/synaptobrevin (Catterall 1999;Mochida et al 2003a,b;Weiss and Zamponi 2012). The site of this interaction is the "synprint" (synaptic protein interaction) region located in the large intracellular linker between domains II and III of Ca V (Sheng et al 1994Rettig et al 1996) in the case of Ca V 2.1 and Ca V 2.2 channels; disruption of this motif reduces the efficacy of neurotransmission (Catterall 1999), although emerging evidence suggests that other mechanisms or regulatory proteins, such as CASK, Mint-1, and Rab3-interacting molecules (RIMs), may also facilitate the presynaptic organization of calcium channels (Kaneko et al 2002;Maximov and Bezprozvanny 2002;Kiyonaka et al 2007;Liu et al 2011;Graf et al 2012). In addition to facilitating neurotransmission, the SNARE proteins also exert inhibitory modulation on Ca V 2.1; syntaxin reduces Ca V channel expression and inhibits its activity on heterologous expression (Bezprozvanny et al 1995).…”