Abstract: Synaptotagmins are a large family of membrane-trafficking proteins. They are evolutionarily conserved and have 15 members in rodents and humans. Synaptotagmins-1, -2, and -9, are known to have an essential role as calcium sensors for fast synaptic release. Synaptotagmin-7 is a major calcium sensor for the exocytosis of large secretory vesicles in endocrine cells. The functional roles of most synaptotagmin isoforms remain unknown. Here we examined whether synaptotagmins are expressed in the rodent brain in distinct patterns and whether individual neurons and astrocytes coexpress multiple synaptotagmin isoforms. We performed a systematic analysis of expression using radioactive in situ hybridization and quantitative real-time reverse-transcriptase polymerase chain reaction (RT-PCR) as well as multiplex RT-PCR on a single-cell level. Our results demonstrate that most synaptotagmins are expressed in the rodent brain in highly distinctive expression patterns, and that individual neurons express variable subsets of different synaptotagmins. We also show that Syt-11 is the major isoform expressed in astrocytes. This study therefore supports the hypothesis that the functional properties of individual neurons and astrocytes are conferred by the specific subset of synaptotagmins expressed in a cell. J. Comp. Neurol. 512:514-528, 2009. (c) 2008 Wiley-Liss, Inc.
Abstract: Die Weiterleitung eines elektrischen Stimulus von einer Nervenzelle zur nächsten erfolgt an spezialisierten zellulären Schnittstellen, den Synapsen. An chemischen Synapsen erfolgt die Signalübertragung durch die Fusion synaptischer Vesikel mit der präsynaptischen Membran und die Freisetzung darin enthaltener Neurotransmitter in den synaptischen Spalt. Im Gegensatz zu anderen sekretorischen Prozessen erfolgt diese Freisetzung extrem schnell, streng kontrolliert und örtlich begrenzt, gleichzeitig jedoch dynamisch reguliert. Der spezialisierte Bereich der Präsynapse, in dem die Fusion stattfindet, heiÃt Aktive Zone. Ultrastrukturelle Untersuchungen haben gezeigt, dass sie exakt gegenüber dem postsynaptischen Neurotransmitterrezeptionsapparat liegt, und dass an der Plasmamembran auf beiden Seiten des synaptischen Spalts eine elektronendichte Struktur ausgebildet ist. Dieses elektronendichte Material wird an der Präsynapse als Cytomatrix an der Aktiven Zone (CAZ) oder präsynaptisches Netz bezeichnet. Bis heute wurden fünf Proteinfamilien identifiziert, deren Mitglieder spezifisch an der Aktiven Zone angereichert sind: Munc13s, RIMs, ELKS, Bassoon/Piccolo und Liprin-a. In den letzten Jahren haben die Ergebnisse genetischer, biochemischer, struktureller und physiologischer Untersuchungen erste Einblicke in die Funktion dieser Proteine und ihrer Beteiligung an der Regulation der Fusion synaptischer Vesikel, der Vermittlung verschiedener Formen synaptischer Plastizität und der strukturellen Organisation der Aktiven Zone geliefert.
Abstract: RIM-binding proteins (RIM-BPs) were identified as binding partners of the presynaptic active zone proteins RIMs as well as for voltage-gated Ca(2+)-channels. They were suggested to form a functional link between the synaptic-vesicle fusion apparatus and Ca(2+)-channels. Here we show that the RIM-BP gene family diversified in different stages during evolution, but retained their unique domain structure. While invertebrate genomes contain one, and vertebrates include at least two RIM-BPs, we identified an additional gene, RIM-BP3, which is exclusively expressed in mammals. RIM-BP3 is encoded by a single exon of which three copies are present in the human genome. All RIM-BP genes encode proteins with three SH3-domains and two to three fibronectin III repeats. The flanking regions diverge in size and sequence and are alternatively spliced in RIM-BP1 and -2. Quantitative real-time RT-PCR and in situ hybridization analyses revealed overlapping but distinct expression patterns throughout the brain for RIM-BP1 and -2, while RIM-BP3 was detected at high levels outside the nervous system. The modular domain structure of RIM-BPs, their expression pattern and the conservative expansion during evolution shown here support their potential role as important molecular adaptors.
Abstract: Alpha-RIMs (RIM1alpha and RIM2alpha) are multidomain active zone proteins of presynaptic terminals. Alpha-RIMs bind to Rab3 on synaptic vesicles and to Munc13 on the active zone via their N-terminal region, and interact with other synaptic proteins via their central and C-terminal regions. Although RIM1alpha has been well characterized, nothing is known about the function of RIM2alpha. We now show that RIM1alpha and RIM2alpha are expressed in overlapping but distinct patterns throughout the brain. To examine and compare their functions, we generated knockout mice lacking RIM2alpha, and crossed them with previously produced RIM1alpha knockout mice. We found that deletion of either RIM1alpha or RIM2alpha is not lethal, but ablation of both alpha-RIMs causes postnatal death. This lethality is not due to a loss of synapse structure or a developmental change, but to a defect in neurotransmitter release. Synapses without alpha-RIMs still contain active zones and release neurotransmitters, but are unable to mediate normal Ca(2+)-triggered release. Our data thus demonstrate that alpha-RIMs are not essential for synapse formation or synaptic exocytosis, but are required for normal Ca(2+)-triggering of exocytosis.
