A defining feature of eukaryotic cells is the presence of numerous membrane-bound organelles that subdivide the intracellular space into distinct compartments. Material exchange among most organelles occurs via vesicles that bud off from a source and specifically fuse with a target compartment. How the eukaryotic cell acquired its internal complexity is still poorly understood.
We strive to understand the mechanisms and the evolutionary history of the complex molecular machine that drives this important process. The key factors in vesicle fusion belong to conserved protein families. The core of the machine that drives the fusion of transport vesicles is composed of members of the SNARE protein family. They assemble into tight membrane-bridging complexes to pull two membranes together. Their activity is orchestrated by various other factors including Sec1/Munc18 (SM), Rab, and tethering proteins. Together, they form an ancient molecular machine that diversified during evolution to adapt to the needs of specialized compartments. Rapid exocytosis of neurotransmitters from synaptic vesicles constitutes one of such adaptations, and specific regulatory proteins such as synaptotagmins and complexins evolved in the animal kingdom.
Using biochemical, biophysical, morphological, and bioinformatics approaches we address the following questions:
To better understand the molecular events underlying vesicular fusion we explore the physicochemical properties of the protein-protein interactions involved. We want to identify the domains involved, locate the binding surfaces and study the affinities, stoichiometries and kinetics of the interactions and their interplay with membranes to disentangle the entire network. For our biochemical studies, we almost exclusively use recombinant proteins. Next to standard biochemical techniques, we employ spectroscopic (circular dichroism and fluorescence spectroscopy) and calorimetric methods. Where feasible we use high-resolution structural techniques.
Concomitantly we want to shed light on the evolutionary history of the vesicle fusion machine, which arose from an ancient prototypic mechanism during the rise of the last eukaryotic common ancestor (LECA) from its prokaryotic-like ancestor. We would like to uncover how the mechanism adapted in different eukaryotic lineages and how it was most probably organized in the proto-eukaryotic cell. A particular focus lies on the evolutionary changes of the repertoire of the secretory machine during the rise of animals. Taking advantage of the huge number of available sequence data sets, our group developed a database to store and analyze sequences of the protein families involved in vesicle trafficking. Sequences are analyzed through iterative use of hidden Markov models and tree building.
In conjunction, we plan to scrutinize in vivo, the interaction steps that we identified biochemically in vitro. Ultimately we want to correlate the configuration of the (neuro) secretory machinery in the cell with mutations/diseases-related to transport deficiencies. In addition, we are taking a closer look at the very early stages in the evolution of the secretory apparatus by studying the choanoflagellate Monosiga brevicollis and the placozoan Trichoplax adhaerens. Choanoflagellates are a group of mostly single-celled eukaryotes thought to be the closest known sister group to animals; Trichoplax is an animal positioned near the root of the animal tree. It is a simple, free-living marine animal without a nervous system and that glides using cilia to feed on algae. For this, we use, among others, state-of-the-art light and electron microscopy approaches.
Hidden cell diversity in Placozoa: ultrastructural insights from Hoilungia hongkongensis.
Cell Tissue Res. 2021 Apr 19;10.1007/s00441-021-03459-yDOI / PMID / PDF
A conformational switch driven by phosphorylation regulates the activity of the evolutionarily conserved SNARE Ykt6.
Proc Natl Acad Sci U S A. 2021 Mar 23;118(12):e2016730118DOI / PMID
Choanoflagellates and the ancestry of neurosecretory vesicles.
Philos Trans R Soc Lond B Biol Sci. 2021 Mar 29;376(1821):20190759DOI / PMID
The diversification and lineage-specific expansion of nitric oxide signaling in Placozoa: insights in the evolution of gaseous transmission.
Sci Rep. 2020 Aug 3;10(1):13020DOI / PMID
PI(4,5)P2-dependent regulation of exocytosis by amisyn, the vertebrate-specific competitor of synaptobrevin 2
Proc Natl Acad Sci U S A. 2020 Jun 16;117(24):13468-13479DOI / PMID
Prototypic SNARE Proteins Are Encoded in the Genomes of Heimdallarchaeota, Potentially Bridging the Gap between the Prokaryotes and Eukaryotes.
Curr Biol. 2020 Jul 6;30(13):2468-2480.e5DOI / PMID
Glycine as a signaling molecule and chemoattractant in Trichoplax (Placozoa): insights into the early evolution of neurotransmitters.
Neuroreport. 2020 Apr 8;31(6):490-497DOI / PMID
High Cell Diversity and Complex Peptidergic Signaling Underlie Placozoan Behavior.
Curr Biol. 2018 Nov 5;28(21):3495-3501.e2DOI / PMID
Evidence for a conserved inhibitory binding mode between the membrane fusion assembly factors Munc18 and syntaxin in animals.
J Biol Chem. 292(50):20449-60DOI / PMID
Functional assays for the assessment of the pathogenicity of variants of GOSR2, an ER-to-Golgi SNARE involved in progressive myoclonus epilepsies.
Dis Model Mech. 10(12):1391-1398DOI / PMID
Getting Nervous: An Evolutionary Overhaul for Communication.
Annu Rev Genet. 51:455-476DOI / PMID / Serval
Shedding light on the expansion and diversification of the Cdc48 protein family during the rise of the eukaryotic cell.
BMC Evol Biol. Oct 18;16(1):215DOI / PMID / AAA Database
The SM protein Sly1 accelerates assembly of the ER–Golgi SNARE complex.
PNAS 111:13828-33DOI / PMID
Novel cell types, neurosecretory cells, and body plan of the early-diverging metazoan Trichoplax adhaerens.
Curr Biol. 24:1565-72. (* co-last authors)DOI / PMID / UNIL News / Comment in DOI / PMID
Syntaxin1a variants lacking an N-peptide or bearing the LE mutation bind to Munc18a in a closed conformation.
PNAS. 110:12637-42DOI / PMID
Phosphatidylinositol 4,5-bisphosphate clusters act as molecular beacons for vesicle recruitment.
Nat Struct Mol Biol. 20:679-86DOI / PMID
Untangling the evolution of Rab G proteins: implications of a comprehensive genomic analysis.
BMC Biol. 10:71DOI / PMID / Rab Database
Munc18-1 mutations that strongly impair SNARE-complex binding support normal synaptic transmission.
EMBO J. 31(9):2156-68 (1: equally contributing authors; 2: corresponding authors)DOI / PMID
Primordial neurosecretory apparatus identified in the choanoflagellate Monosiga brevicollis.
PNAS. 108(37):15264-9DOI / PMID / New Scientist
A coiled-coil trigger site is essential for rapid binding of synaptobrevin to the SNARE acceptor complex.
J Biol Chem. 285:21549-59DOI / PMID
Synaptobrevin N-terminally bound to syntaxin-SNAP-25 defines the primed vesicle state in regulated exocytosis.
Cell Biol. 188(3):401-13DOI / PMID
A conserved membrane attachment site in α-SNAP facilitates NSF-driven SNARE complex disassembly.
J Biol Chem. 284(46):31817-26DOI / PMID
Single vesicle millisecond fusion kinetics reveals number of SNARE complexes optimal for fast SNARE-mediated membrane fusion.
J Biol Chem. 284(46):32158-66DOI / PMID
The Ca2+ affinity of synapto-tagmin 1 is markedly increased by a specific interaction of its C2B domain with phosphatidylinositol 4,5-bisphosphate.
J Biol Chem. 284:25749-60DOI / PMID
Is assembly of the SNARE complex enough to fuel membrane fusion?
J Biol Chem. 284:13143-52DOI / PMID / Comment in DOI / PMID
Phylogeny of the SNARE vesicle fusion machinery yields insights into the conservation of the secretory pathway in fungi.
BMC Evolutionary Biology 9:19DOI / PMID / SNARE Database
Imaging the assembly and disassembly kinetics of cis-SNARE complexes on native plasma membranes.
FEBS Letters, 582:3563-8DOI / PMID
SNAREing the basis of multicellularity: Consequences of protein family expansion during evolution.
Mol.Biol.Evol. 25:2055-68DOI / PMID / SNARE Database
Munc18a controls SNARE assembly through its interaction with the syntaxin N-peptide.
EMBO J. 27: 923-33DOI / PMID
Binding of α-SNAP to syntaxin 1 blocks SNARE-dependent exocytosis.
Mol. Biol. Cell 19:776-84DOI / PMID
Synaptotagmin activates membrane fusion through a Ca2+-dependent trans interaction with phospholipids.
Nat. Struct. Mol. Biol. 14: 904-11DOI / PMID
Budding insights on cell polarity (News & Views).
Nature Structural & Molecular Biology 14: 360-2DOI / PMID
An elaborate classification of SNARE proteins sheds light on the conservation of the eukaryotic endomembrane system.
Mol. Biol. Cell 18: 3463-71DOI / PMID / InCytes / SNARE Database
Determinants of Synaptobrevin regulation in Membranes.
Mol. Biol. Cell 18: 2037-46DOI / PMID / InCytes
Early endosomal SNAREs form a structurally conserved SNARE complex and fuse liposomes with multiple topologies.
EMBO J. 26: 9-18DOI / PMID
N- to C-terminal SNARE complex assembly promotes rapid membrane fusion.
Science 313: 673-6DOI / PMID
Identification of SNAP-47, a novel Qbc-SNARE with ubiquitous expression.
J. Biol. Chem. 281: 17076-83DOI / PMID
Sequential N- to C-terminal ‘zipping-up’ of the SNARE complex drives priming and fusion of secretory vesicles.
EMBO J. 25:955-66DOI / PMID
Alternative Splicing of SNAP-25 Regulates Secretion through Nonconservative Substitutions in the SNARE Domain.
Mol. Biol. Cell 12: 5675-85DOI / PMID
A structural basis for the inhibitory role of tomosyn in exocytosis.
J. Biol. Chem. 279: 47192-200 “JBC Paper of the week”DOI / PMID
A transient N-terminal interaction of SNAP-25 and syntaxin nucleates SNARE assembly.
J. Biol. Chem. 279: 7613-21DOI / PMID
Single-molecule fluorescence resonance energy transfer reveals a dynamic equilibrium between closed and open conformations of syntaxin-1.
PNAS 100: 15516-21DOI / PMID
Structural insights into the SNARE mechanism.
BBA - Molecular Cell Research. Special Issue: Membrane Fusion, 1641: 87-97DOI / PMID
The R-SNARE motif of tomosyn forms SNARE core complexes with syntaxin 1 and SNAP-25 and down-regulates exocytosis.
J. Biol. Chem. 278: 31159-66DOI / PMID
Crystal structure of a complex between human spliceosomal cyclophilin H and a U4/U6 snRNP-60K peptide.
J. Mol. Biol., 331: 45-56DOI / PMID
Habc Domain and SNARE Core Complex Are Connected by a Flexible Linker.
Biochemistry, 42: 4009-14DOI / PMID
SNARE assembly and disassembly exhibit a pronounced hysteresis.
Nat. Struct. Biol. 9: 144-151DOI / PMID / Commented in DOI / PMID
Crystal structure of the endosomal SNARE complex reveals common structural principles of all SNAREs.
Nat. Struct. Biol. 9: 107-111DOI / PMID
Rapid and selective binding to the synaptic SNARE complex suggests a modulatory role of complexins in neuroexocytosis.
J. Biol. Chem. 277: 7838-48DOI / PMID
Homo- and heterooligomeric SNARE complexes studied by site-directed spin labeling.
J. Biol. Chem. 276: 13169-77DOI / PMID
A SNARE complex mediating fusion of late endosomes defines conserved properties of SNARE structure and function.
EMBO J. 19:6453-64DOI / PMID
Selective interaction of complexin with the neuronal SNARE complex: determination of the binding regions.
J. Biol. Chem. 275:19808-18DOI / PMID
Kinetics of Synaptotagmin Respones to Ca2+ and Assembly with the Core SNARE Complex onto Membranes.
Neuron 24: 363-76DOI / PMID
Conserved structural features of the synaptic fusion complex: SNARE proteins reclassified as Q- and R-SNAREs.
Proc. Natl. Acad. Sci. USA 95: 15781-86DOI / PMID
Crystal structure of a SNARE complex involved in synaptic exocytosis at 2.4 Å resolution.
Nature 395: 347-353DOI / PMID / Comment in DOI / PMID
Identification of a minimal core of the synaptic SNARE-complex sufficient for reversible assembly and disassembly.
Biochemistry 37: 10345-55DOI / PMID
Structural changes are associated with SNARE complex formation.
J. Biol. Chem. 272, 28036-41DOI / PMID
A structural change occurs upon binding of syntaxin to SNAP-25.
J. Biol. Chem. 272: 4582-90DOI / PMID
ARF and VAPP14: two proteins involved in the delivery of heparan sulfate proteoglycan from the trans-Golgi network to the plasma membrane.
Ann N Y Acad Sci. 733:344-56DOI / PMID
Comparative genomics and the nature of placozoan species.
PLoS Biol. 2018 Jul 31;16(7):e2005359DOI / PMID
Loss of Neuroligin3 specifically downregulates retinal GABAAα2 receptors without abolishing direction selectivity.
PLoS One. 2017 Jul 14;12(7):e0181011DOI / PMID
82nd International Titisee Conference held from October 25 to 29, 2000. Mechanisms of membrane fusion.
B.I.F. FUTURA Vol. 16, No. 1, 13-23 (Conference report)Download PDF
Mechanismen intrazellulärer Membranfusion.
Biospektrum No. 1, 2731 (review article; german)Download PDF
Mechanism of SNARE assembly and disassembly.
Landes Bioscience (Book chapter)Download PDF
Dirk Fasshauer studied biology and received his doctorate degree from the University of Göttingen in 1994. He worked as a post-doctoral fellow at Yale University from 1995 to 1997 and then moved to the Max Planck Institute for Biophysical Chemistry in Göttingen and from 2002, headed the “Structural Biochemistry” Research Group in the Department of Neurobiology. In 2009 he joined the DNF as associate professor.
Iman obtained her B.Sc. in Biochemistry in June 2016 at the Mohamed Khidher University in Algeria.Then, she obtained her Master in Molecular biology, Immunology, and Microbiology Specialities at the Eotvos Lorand University, Hungary. She started her PhD thesis in November 2020.
Michela studied Molecular and Medical biotechnology and received her Master's degree in March 2020 at the University of Verona, Italy, with a bioinformatic thesis about RNA sequencing analysis on lymphoma cancer. She started her PhD in August 2020.
Yasmine studied Molecular Biology and received her Master degree from the University of Lausanne in 2020. During her Master project she did an internship for 3 months in the lab. In January 2021 she joined the group as technician.
Ioanna received a BSc in Molecular Biology and Genetics in 2015 from Democritus University of Thrace in Greece and then obtained a Master of Research in Protein Structure and Function from the University of Bath in 2016. In February 2017, she joined the group as a PhD student where she studies the conformational flexibility of the Syntaxin-1a/Munc18-1 complex.
Deepak received his Master's degree in Bioinformatics from IIIT Hyderabad in 2013. He worked as a researcher at TRDDC (TCS Research) Pune from 2013 to 2020, with the research focus on NGS analysis, metagenomics and network biology. He started his PhD thesis in November 2020.
In the SNAREs section of the database you have the possibility to view our classified sequences (SNAREs -> View DB) or to de novo classify protein sequences (SNAREs -> Find Motif). If you choose to view our classified sequences, you find additional information for each of the fields on the ViewDB site by clicking the question mark behind the field. If you choose to de novo classify protein sequences you may paste any protein sequence into the text field of the Find Motif site and hit the Submit-button. The results will be displayed shortly after.Snare DB
In the AAAs section of the database you have the possibility to view our classified sequences (AAAs -> View Database) or to de novo classify protein sequences (AAAs -> Scan Sequence against HMMs). If you choose to view our classified sequences, you find additional information for each of the fields on the ViewDB site by clicking the question mark behind the field. If you choose to de novo classify protein sequences you may paste any protein sequence into the text field of the Find Motif site and hit the Submit-button. The results will be displayed shortly after.AAA DB
In the Rabs section of the database you have the possibility to view our classified sequences (Rabs -> View Database) or to de novo classify protein sequences (Rabs -> Scan Sequence against HMMs). If you choose to view our classified sequences, you find additional information for each of the fields on the ViewDB site by clicking the question mark behind the field. If you choose to de novo classify protein sequences you may paste any protein sequence into the text field of the Find Motif site and hit the Submit-button. The results will be displayed shortly after.RAB DB
Email: dirk.fasshauer [ at ] unil.ch
Phone: +41 21 692 4282