Synapse Formation: A Summary of Evolution and Development PDF

SYNAPSE Department of Molecular Life Sciences University of Zurich FORMATION [email protected] HOW DID EVOLUTION ‘BUILD’ A SYNAPSE? 50 nm Synapse Formation PRESYNAPTIC During Evolution HOMEOSTASIS AT THE DROSOPHI...

SYNAPSE Department of Molecular Life Sciences University of Zurich FORMATION [email protected] HOW DID EVOLUTION ‘BUILD’ A SYNAPSE? 50 nm Synapse Formation PRESYNAPTIC During Evolution HOMEOSTASIS AT THE DROSOPHILA NMJ https://www.evogeneao.com/learn/tree-of-life Synapse Evolution? PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Synaptic genes but no synapses? Synapses and neural systems may have evolved more than once! Moroz et al., 2014 nature Synapse Evolution PRESYNAPTIC – Placozoa AT THE DROSOPHILA HOMEOSTASIS NMJ Exocytosis and paracrine signaling of peptidergic cells in Placozoa Trichoplax adhaerens (No evidence for neurons or synapses) Placozoa: Synaptic genes, exocytosis and glutamate receptors, but no evidence for neurons! Evolution of theHOMEOSTASIS PRESYNAPTIC neuronal geneATexpression program THE DROSOPHILA NMJ 2 3 1 1 2 3 Najle, Grau-Bové et al., 2023 Cell Evolution of theHOMEOSTASIS PRESYNAPTIC neuronal geneATexpression program THE DROSOPHILA NMJ 2 3 1 Stepwise 1 evolution of the2 neuronal gene expression program 3 Najle, Grau-Bové et al., 2023 Cell ONTOGENETIC SYNAPSE FORMATION 50 nm OUTLINE How to build a synapse? Presynaptic Postsynaptic assembly assembly Synaptic Transmission PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Ca2+ Pre Cav +++ --- --- +++ Post The Synapse Speed (1 AP) < 1ms Sabatini & Regehr, 1996 Rate (many APs) > 1kHz Ritzau-Jost et al., 2014 Dynamics ~ 106-fold Lou et al., 2006 (Neurotransmitter release rate, 1 AP) 50 nm “Synaptic Modules” PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Pre Post 1. Vesicle fusion Presynaptic 2. Active zone 3. Postsynaptic density (PSD)  Pre Post 1. Vesicle fusion 2. Active zone 3. Postsynaptic density (PSD) Vesicle Fusion: HOMEOSTASIS PRESYNAPTIC SNAREs AT THE DROSOPHILA NMJ (v-SNARE) (t-SNARE) SNARE Soluble NSF Attachment Protein (SNAP) Receptor FRET PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Förster/Fluorescence Resonance Energy Transfer FRET PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Förster/Fluorescence Förster/Fluorescence Resonance Resonance Energy Energy Transfer (FRET) Transfer can be employed to probe intermolecular distance Vesicle Fusion HOMEOSTASIS AT THE DROSOPHILA PRESYNAPTIC NMJ Liposome Synaptic vesicle Syntaxin Synaptobrevin (t-SNARE) (v-SNARE) SNAP-25 FRET (Increase in donor fluorescence) van den Bogaar t et al., 2010 nature struct molec biol Vesicle Fusion HOMEOSTASIS AT THE DROSOPHILA PRESYNAPTIC NMJ FRET (Dequenching due to fusion) Donor Vesicle fusion can be reconstituted in vitro, in the absence of active zone components or Ca2+ Holt et al., 2008 Curr Biol  Pre Post 1. Vesicle fusion 2. Active zone 3. Postsynaptic density (PSD) Active Zone Assembly PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Active zone (AZ): Platform for rapid fusion of synaptic vesicles after Ca2+ influx. AZ-membrane is decorated by a proteinacious cytomatrix (a set of specialized proteins) Pre Post Assembly: Contact with postsynaptic partner required? Delivery as preassembled units or single components? Active Zone Assembly PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Human Embyonic Kidney (HEK) cells Neuron 1 µm Synapsin HEK cell (Pre) Presynaptic differentiation can be induced by expression of a single Neuroligin postsynaptic cell adhesion molecule (Neuroligin) on non-neuronal cells! Cerebellar granule cell culture Scheiffele et al., 2000 Cell Active Zone Assembly PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Injected polylysine-coated beads 100 µm 0.5 µm 1 µm Active-zone formation after contact with polylysine-coated beads! Rat cerebellar cortex Burry, 1983 Brain Res Active Zone Assembly PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Synaptic Dendrite vesicles AZ protein (post) v-SNARE Dendrite (post) AZ protein Synaptic vesicles Active-zone assembly does not require a postsynaptic par tner! Hippocampal cell culture (mouse) Krueger et al., 2003 Neuron Active Zone Assembly PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Pre Active Zone Assembly PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Pre Are “orphan” active zones functional? Probing Exocytosis PRESYNAPTIC using FM Destaining HOMEOSTASIS AT THE DROSOPHILA NMJ fluorophore lipophilic + charge (prevents flipping) FM (Fei Mao)-43 (styryl dye) Polar solvent (H2O) => little fluorescence Fluorescence signal if dye molecule in membrane! Gaffield & Betz et al., 2007 Nat Prot Probing Exocytosis PRESYNAPTIC using FM Destaining HOMEOSTASIS AT THE DROSOPHILA NMJ Bath apply Load (stimulation) FM Wash Unload (stimulation) Gaffield & Betz et al., 2007 Nat Prot Active Zone Assembly PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Krueger et al., 2003 Neuron Active Zone Assembly PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ “Orphan synapses” can undergo exocytosis! Krueger et al., 2003 Neuron Active Zone Assembly PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Stationary vesicle cluster Mobile vesicle cluster Krueger et al., 2003 Neuron Active Zone Assembly PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Stationary vesicle cluster Mobile vesicle cluster Vesicle clusters are mobile and can undergo fusion. Krueger et al., 2003 Neuron Active Zone Assembly PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Self-Assembly of active zones: Functional synapses can form without postsynaptic par tner! Pre Assembly: Contact with postsynaptic partner required? Delivery as preassembled units or single components? Active Zone Assembly PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Pre Assembly: Contact with postsynaptic partner required? Delivery as preassembled units or single components? Active Zone Assembly PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Pre Assembly: Contact with postsynaptic partner required? Delivery as preassembled units or single components? Delivery of Active PRESYNAPTIC Zone Components HOMEOSTASIS AT THE DROSOPHILA NMJ (AZ) (SV) Active zone proteins (Bassoon & Piccolo) and synaptic vesicle proteins (VAMP, SV2) in a preassembled multi-vesicle transpor t vesicle Tao-Cheng, 2007 Neurosci Delivery of Active PRESYNAPTIC Zone Components HOMEOSTASIS AT THE DROSOPHILA NMJ 2 µm/2s Brp Bruchpilot (AZ protein) Syt-1 Synaptotagmin-1 (SV protein) Spinster Lysosome marker Par tial axonal co-transpor t of active zone proteins and synaptic vesicle proteins in presynaptic lysosome-related vesicles (PLVs) Motor neuron axon (Drosophila melanogaster) Vukoja et al., 2018 Neuron Delivery of Active PRESYNAPTIC Zone Components HOMEOSTASIS AT THE DROSOPHILA NMJ Lamp1 lysosome marker Arl8 lysosomal kinesin adaptor Vukoja et al., 2018 Neuron Delivery of Active PRESYNAPTIC Zone Components HOMEOSTASIS AT THE DROSOPHILA NMJ Par tial axonal co-transpor t of active zone proteins and synaptic vesicle proteins in presynaptic lysosome-related vesicles (PLVs) requires Arl8 Arl8 lysosomal kinesin adaptor Vukoja et al., 2018 Neuron Delivery of Active PRESYNAPTIC Zone Components HOMEOSTASIS AT THE DROSOPHILA NMJ Rivero-Ríos & Weisman, 2023 Arl8 lysosomal kinesin adaptor KIF1A kinesin family member 1A PIKFYVE 1-phosphatidylinositol 3-phosphate 5-kinase lipid kinase) Human iPSC-derived neurons Rizalar et al., 2023 Delivery of Active PRESYNAPTIC Zone Components HOMEOSTASIS AT THE DROSOPHILA NMJ Rivero-Ríos & Weisman, 2023 Arl8 lysosomal kinesin adaptor KIF1A Axonal kinesin co-transpor family membert 1A of active zone and synaptic vesicle proteins in PIKFYVEprecursor 1-phosphatidylinositol 3-phosphate vesicles requires Arl8,5-kinase KIF1A,lipid andkinase) PI(3,5)P in human neurons. 2 Human iPSC-derived neurons Rizalar et al., 2023 Delivery of Active PRESYNAPTIC Zone Components HOMEOSTASIS AT THE DROSOPHILA NMJ Chia et al., 2007 JCB Delivery of Active PRESYNAPTIC Zone Components HOMEOSTASIS AT THE DROSOPHILA NMJ Microtubule (MT) polarity and specific MT-associated proteins target synaptic components to synapses in axons or dendrites. Chia et al., 2007 JCB Active Zone Assembly PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Pre Post Which molecules are involved in AZ assembly? HOW TO BUILD A SYNAPSE? Drosophila PRESYNAPTICmelanogaster HOMEOSTASIS AT THE DROSOPHILA NMJ © Andrew Woolley The Drosophila Neuromuscular Junction (NMJ) The NeuromuscularATJunction DrosophilaHOMEOSTASIS PRESYNAPTIC THE DROSOPHILA 1 µm 5 µm Anti Bruchpilot (Brp, an active zone component) Genes InvolvedHOMEOSTASIS PRESYNAPTIC in Active ZoneATAssembly THE DROSOPHILA NMJ 10 µm 1 µm Drosophila “Synapse defective-1” (dsyd-1) Owald et al., 2010 JBC Drosophila liprin-a (dliprin-a) Genes InvolvedHOMEOSTASIS PRESYNAPTIC in Active ZoneATAssembly THE DROSOPHILA NMJ Adult CNS Drosophila Adult CNS mouse dsyd-1 synaptic vesicles msyd-1A Urwyler et al., 2015 Development Wentzel et al., 2013 Neuron Synapse Defective-1 is required for normal synaptic vesicle targeting in the Drosophila PNS and CNS, as well as in the mouse CNS. Genes InvolvedHOMEOSTASIS PRESYNAPTIC in Active ZoneATAssembly THE DROSOPHILA NMJ rab3- rup 10 µm 1 µm Graf et al., 2011 Neuron Genes InvolvedHOMEOSTASIS PRESYNAPTIC in Active ZoneATAssembly THE DROSOPHILA NMJ Ca2+ channel µm 1 µm The small GTPase Rab3 is required for normal AZ 10 distribution, and pre/post matching Graf et al., 2011 Neuron rab3-rup The small GTPase Rab3 Rab3 Rab3 GTP GDP Rab3-GAP Genes InvolvedHOMEOSTASIS PRESYNAPTIC in Active ZoneATAssembly THE DROSOPHILA NMJ rab3- rup 10 µm 1 µm Graf et al., 2011 Neuron Genes InvolvedHOMEOSTASIS PRESYNAPTIC in Active ZoneATAssembly THE DROSOPHILA NMJ No major defect in synaptic transmission at rab3 mutant synapses Two-Electrode Voltage Clamp Graf et al., 2011 Neuron Drosophila NMJ (10 Hz stim.) Presynaptic PRESYNAPTIC Development of the HOMEOSTASIS Drosophila AT THE NMJ DROSOPHILA NMJ G: Genetic interaction I: Biochem. Interaction rab3 LAR: Leukocyte antigen-related dsyd-1 (LAR)-type receptor RIM cac: Cacophony (Cav2.1), Ca2+ channel Owald & Sigrist et al., 2009 Curr Opin Neurobiol Presynaptic PRESYNAPTIC Development HOMEOSTASIS AT THE DROSOPHILA NMJ rab3 rab3 dsyd-1 RIM Owald & Sigrist et al., 2009 Curr Opin Neurobiol SYD-1 Presynaptic PRESYNAPTIC Development: HOMEOSTASISSearching AT THE for Genes DROSOPHILA NMJ Margeta et al., 2008 Curr Opin Neurobiol Core Active Zone PRESYNAPTIC Proteins AT THE DROSOPHILA HOMEOSTASIS NMJ Rizalaret et al., 2020 Neuron Core Active Zone PRESYNAPTIC Proteins in AT HOMEOSTASIS Placozoa THE DROSOPHILA NMJ AZ proteins in peptidergic cells (no synapses) => paracrine signaling Najle, Grau-Bové et al., 2023 Cell Active Zone Assembly PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Chia et al., 2007 JCB Branch-Specific Axonal Synaptogenesis PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ Prl-1 Phosphatase of regenerating liver Mechanosensory neuron in the adult Drosophila CNS Urwyler et al., 2019 Science Branch-Specific Axonal Synaptogenesis PRESYNAPTIC HOMEOSTASIS AT THE DROSOPHILA NMJ The phosphatase Prl-1 specifies subcellular synapse location and axon Prl-1 Phosphatase branchingof regenerating in an axonliver branch specific fashion in the Drosophila CNS Mechanosensory neuron in the adult Drosophila CNS Urwyler et al., 2019 Science  Pre Post 1. Vesicle fusion 2. Active zone 3. Postsynaptic density (PSD) The Synapse HOMEOSTASIS AT THE DROSOPHILA PRESYNAPTIC EM 50 nm Assembly: Contact with postsynaptic partner required? Delivery as preassembled units or single components? Pre Post 1. Vesicle fusion 2. Active zone 3. Postsynaptic density (PSD) Postsynaptic PRESYNAPTIC Density HOMEOSTASIS AT THE DROSOPHILA NMJ Fibroblast Fibroblast (Pre) (Pre) Postsynaptic Postsynaptic Postsynaptic (inhibitory) (excitatory) (inhibitory) Neurexin expression in non-neuronal cells clusters glutamate- and GABA postsynaptic scaffolding proteins in dendrites Graf et al., 2004 Cell Mouse hippocampal culture and fibroblasts Postsynaptic PRESYNAPTIC Density & AZ Assembly HOMEOSTASIS AT THE DROSOPHILA NMJ vGluT vesicular glutamate transpor ter GAD Glutamate Decarboxylase Neuroligin expression in non-neuronal cells clusters glutamate- and GABA-containing synaptic vesicles. Graf et al., 2004 Cell Postsynaptic PRESYNAPTIC Density & AZ Assembly HOMEOSTASIS AT THE DROSOPHILA NMJ Excitatory synapse Inhibitory synapse Neuroligin-1 and -3 Neuroligin-2 Glutamate receptors GABAA receptors PSD-95 Gephyrin HOW TO BUILD A SYNAPSE? Postsynaptic & transsynaptic molecules Maturation Elimination/competition Plasticity Questions? [email protected]