Biology subjects

Fasshauer, D.

Publications and source records attributed to Fasshauer, D..

15 recordsLinked to original sources

bioRxiv2026

Heimdall SNAREs reveal an ancestral architecture of eukaryotic SNARE complexes

SNARE proteins are central to eukaryotic cell biology, where they drive membrane fusion by assembling into four-helix bundles that bring opposing membranes into close proximity. Their ubiquitous role in intracellular membrane trafficking raises fundamental questions about their evolutionary origin and the emergence of eukaryotic cellular complexity. Here, we characterize SNARE-like proteins from Heimdallarchaeota, a lineage within the Asgard archaea that is closely related to eukaryotes. We show that a Heimdall SNARE forms a stable homotetrameric four-helix bundle in solution and determine its structure by X-ray crystallography. Strikingly, the Heimdall complex closely resembles eukaryotic SNARE complexes in its overall architecture and core organization, despite being composed of four identical subunits. Molecular dynamics simulations further reveal conserved physical features of the bundle alongside distinctive properties of its central 0-layer. These findings demonstrate that the characteristic four-helix architecture of SNAREs predates the emergence of eukaryotic cells and provide structural insight into the evolution of the membrane fusion machinery.

biochemistry↗

bioRxiv2026

TRACEY: an updated resource for SNARE protein domain annotation with improved HMMs and expanded sequence coverage

MotivationSNARE proteins catalyse membrane fusion across the eukaryotic endomembrane system, from synaptic vesicle exocytosis to intracellular trafficking, endosomal and vacuolar transport, and autophagy, and their accurate domain annotation depends on the quality of profile models and the sequence diversity behind them. The original SNARE domain classification predates the recent expansion of eukaryotic sequence data, leaving its HMM profiles and subgroup coverage unable to resolve divergent and lineage-specific paralogs. ResultsWe present an updated release of TRACEY built on a resynchronized, non-redundant collection of 18,915 curated SNARE proteins spanning 1,188 species, together with a consolidated set of 83 HMM profiles, including 43 models for newly defined subgroups, reconstructed through an iterative, mixture-model-driven procedure. In direct comparison with the legacy models, at least [~]75% of sequences in every overlapping group scored better with the new HMMs, indicating systematic gains in domain detection. A redesigned web interface adds multiparameter querying, FASTA download, and direct scanning of user-submitted sequences against the curated profiles. Availability and implementationTRACEY is freely available at https://tracey.unil.ch. Contactdirk.fasshauer@unil.ch Supplementary informationSupplementary data are available at Bioinformatics online.

bioinformatics↗

bioRxiv2026

Transmembrane domain composition reflects subcellular localization of SNARE proteins

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=128 SRC="FIGDIR/small/713902v1_ufig1.gif" ALT="Figure 1"> View larger version (55K): org.highwire.dtl.DTLVardef@1800a38org.highwire.dtl.DTLVardef@11f19aborg.highwire.dtl.DTLVardef@ce3b90org.highwire.dtl.DTLVardef@1a8935b_HPS_FORMAT_FIGEXP M_FIG C_FIG Transmembrane domains (TMDs) anchor proteins within membranes of distinct composition and biophysical properties, necessitating adaptation to each membranes unique environment. SNARE proteins, the core mediators of eukaryotic membrane fusion, provide an ideal system to study such adaptations: they are ubiquitous across Eukaryota, with paralogs localized to different compartments that have evolved independently to match their membrane context. Here, we combine statistical analyses of approximately 14,000 SNARE TMDs with MD simulations of 28 human SNARE TMDs to uncover how TMDs have adapted to distinct membrane environments. We identify a clear compositional dichotomy that separates SNAREs of the early secretory pathway (ER and Golgi apparatus) from those of the late secretory and endosomal pathways: early TMDs are enriched in bulky Phe residues, whereas late TMDs are dominated by smaller Ile residues. The larger Phe side chains likely enhance hydrophobic interactions in loosely packed ER and Golgi membranes, while Ile likely prevents disruption of the tight lipid-lipid packing in the plasma and endosomal membranes, thereby contributing to protein localization. TMD lengths derived from MD simulations differ from those inferred by sequence-based approaches and show limited variations across compartments. Together, these results reveal how variations in membrane composition shaped SNARE TMDs across eukaryotic compartments.

molecular biology↗

bioRxiv2025

Rapid and Interpretable Protein Contact Map Prediction Using a Pattern-Matching Strategy

Protein sequence determines the structure, function, and dynamics of a protein. In recent years, enormous progress has been made in translating sequence information into structural information using machine learning approaches. However, because of the underlying methodology, it is an immense computational challenge to extract this information from the ever-increasing number of sequences. In the present study, we show that it is possible to create two-dimensional contact maps from sequences, for which only a few exemplary structures are available on a laptop without the need for GPUs or high-performance computing clusters. This is achieved by using a pattern matching approach. The resulting contact maps largely reflect the interactions in the three-dimensional structures. The validity of our method was tested on the 25 protein domains, with abundant structural data, achieving correlations of 0.73-0.94 between predicted and experimental contact maps. To demonstrate broader applicability, we further validated our approach on 7,599 poorly annotated sequences using homologous structural templates, achieving a mean F1-score of 0.609 {+/-} 0.095 and mean accuracy of 0.954 {+/-} 0.036 when compared against high-confidence AlphaFold structures. These results demonstrate that our pattern matching approach maintains robust performance even when relying on a small number of structural templates.

bioinformatics↗

bioRxiv2024

Unveiling Conserved Allosteric Hot Spots in Protein Domains from Sequences

The amino acid sequence determines the structure, function, and dynamics of a protein. In recent years, enormous progress has been made in translating sequence information into 3D structural information using artificial intelligence. However, because of the underlying methodology, it is an immense computational challenge to extract this information from the ever-increasing number of sequences. In the present study, we show that it is possible to create 2D contact maps from sequences, for which only a few exemplary structures are available on a laptop without the need for GPUs. This is achieved by using a pattern-matching approach. The resulting contact maps largely reflect the interactions in the 3D structures and contain information about its function and dynamics. This approach was used to explore the evolutionarily conserved allosteric mechanisms and identify the source- sink (driver-driven) relationships by using an established method that combines Schreibers concept of entropy transfer with a simple Gaussian network model. The validity of our method was tested on the DHFR, PDZ, SH3, and S100 domains, with our predictions consistently aligning with the experimental findings.

bioinformatics↗

bioRxiv2024

A look beyond the QR code of SNARE proteins

Soluble N-ethylmaleimide-sensitive factor Attachment protein REceptor (SNARE) proteins catalyze the fusion process of vesicles with target membranes in eukaryotic cells. To do this, they assemble in a zipper-like fashion into stable complexes between the membranes. Structural studies have shown that the complexes consist of four different helices, which we subdivide into Qa-, Qb-, Qc-, and R-helix on the basis of their sequence signatures. Using a combination of biochemistry, modeling and molecular dynamics, we investigated how the four different types are arranged in a complex. We found that there is a matching pattern in the core of the complex that dictates the position of the four fundamental SNARE types in the bundle, resulting in a QabcR complex. In the cell, several different cognate QabcR-SNARE complexes catalyze the different transport steps between the compartments of the endomembrane system. Each of these cognate QabcR complexes is compiled from a repertoire of about 20 SNARE subtypes. Our studies show that exchange within the four types is largely tolerated structurally, although some non-cognate exchanges lead to structural imbalances. This suggests that SNARE complexes have evolved for a catalytic mechanism, a mechanism that leaves little scope for selectivity beyond the QabcR rule.

biochemistry↗

bioRxiv2023

Arachidonic acid does not grease the exocytotic SNARE machinery

It has been proposed that arachidonic acid stimulates neurite outgrowth by specifically activating syntaxin 3, a SNARE proteins that mediates the fusion of transport vesicles with the plasma membrane and is thus instrumental for the insertion of new membrane needed for cell growth and proliferation. Considering the important role of arachidonic acid and other polyunsaturated fatty acids for human health, these findings would have wide implications for everyday life. Here, I have report that the effect of arachidonic acid is caused by a nonspecific, detergent-like action on SNARE proteins in vitro. The action requires unphysiologically high concentrations, is mimicked by several detergents or detergent-like substances forming micelles regardless of whether arachidonic acid is present or not, and is thus highly unlikely to be of any physiological significance for SNARE function.

biochemistry↗

bioRxiv2022

Exploring the conformational changes of the Munc18-1/syntaxin 1a complex

Neurotransmitters are released from synaptic vesicles, the membrane of which fuses with the plasma membrane upon calcium influx. This membrane fusion reaction is driven by the formation of a tight complex comprising the plasma membrane SNARE proteins syntaxin-1a and SNAP-25 with the vesicle SNARE protein synaptobrevin. The neuronal protein Munc18-1 forms a stable complex with syntaxin-1a. Biochemically, syntaxin-1a cannot escape the tight grip of Munc18-1, so formation of the SNARE complex is inhibited. However, Munc18-1 is essential for the release of neurotransmitters in vivo. It has therefore been assumed that Munc18-1 makes the bound syntaxin-1a available for SNARE complex formation. Exactly how this occurs is still unclear, but it is assumed that structural rearrangements occur. Here, we used a series of mutations to specifically weaken the complex at different critical positions in order to induce these rearrangements biochemically. Our approach was guided through sequence and structural analysis and supported by molecular dynamics simulations. Subsequently, we created a homology model showing the complex in an altered conformation. This conformation presumably represents a more open arrangement of syntaxin-1a that permits the formation of SNARE complex to be initiated while still bound to Munc18-1. In the future, research should investigate how this central reaction for neuronal communication is controlled by other proteins.

biochemistry↗

bioRxiv2022

Megaviruses contain various genes encoding for eukaryotic vesicle trafficking factors

Many intracellular pathogens, such as bacteria and large viruses, enter eukaryotic cells via phagocytosis, then replicate and proliferate inside the host. To avoid degradation in the phagosomes, they have developed strategies to modify vesicle trafficking. Although several strategies of bacteria have been characterized, it is not clear whether viruses also interfere with the vesicle trafficking of the host. Recently, we came across SNARE proteins encoded in the genomes of several bacteria of the order Legionellales. These pathogenic bacteria may use SNAREs to interfere with vesicle trafficking, since SNARE proteins are the core machinery for vesicle fusion during transport. They assemble into membrane-bridging SNARE complexes that bring membranes together. We now have also discovered SNARE proteins in the genomes of diverse giant viruses. Our biochemical experiments showed that these proteins are able to form SNARE complexes. We also found other key trafficking factors that work together with SNAREs such as NSF, SM, and Rab proteins encoded in the genomes of giant viruses, suggesting that viruses can make use of a large genetic repertoire of trafficking factors. Most giant viruses possess different collections, suggesting that these factors entered the viral genome multiple times. In the future, the molecular role of these factors during viral infection need to be studied. O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY SynopsisGiant viruses enter their eukaryotic host cells by phagocytosis. For reproduction, they hijack the host cells membranes by an unknown mechanism. Here, we found that giant viruses express several core factors of the eukaryotic vesicle fusion machinery, including SNARE, Rab, SM proteins, and the disassembly protein NSF. Very probably, these genes were transferred from different eukaryotic hosts to different viruses. Whether the viruses deploy these factors for interfering with the vesicle trafficking of the host cell needs to be investigated.

genomics↗

bioRxiv2020

Hidden cell diversity in Placozoa: Ultrastructural Insights from Hoilungia hongkongensis

From a morphological point of view, placozoans are among the most simple free-living animals. This enigmatic phylum is critical for our understanding of the evolution of animals and their cell types. Their millimeter-sized, disc-like bodies consist of only three cell layers that are shaped by roughly six major cell types. Placozoans lack muscle cells and neurons but are able to move using their ciliated lower surface and take up food in a highly coordinated manner. Intriguingly, the genome of Trichoplax adhaerens, the founding member of the enigmatic phylum, has disclosed a surprising level of genetic complexity. Moreover, recent molecular and functional investigations have uncovered a much larger, so-far hidden cell-type diversity. Here, we have extended the microanatomical characterization of a recently described placozoan species - Hoilungia hongkongensis. In H. hongkongensis, we recognized the established canonical three-layered placozoan body plan but also came across several morphologically distinct and potentially novel cell types, among them novel gland cells and "shiny spheres"-bearing cells at the upper epithelium. Thus, the diversity of cell types in placozoans is indeed higher than anticipated.

evolutionary biology↗

bioRxiv2020

Choanoflagellates and the ancestry of neurosecretory vesicles

Neurosecretory vesicles are highly specialized trafficking organelles important for metazoan cell-cell signalling. Despite the high anatomical and functional diversity of neurons in metazoans, the protein composition of neurosecretory vesicles in bilaterians appears to be similar. This similarity points towards a common evolutionary origin. Moreover, many key neurosecretory vesicle proteins predate the origin of the first neurons and some even the origin of the first animals (metazoans). However, little is known about the molecular toolkit of these vesicles in non-bilaterian metazoans and their closest unicellular relatives, making inferences about the evolutionary origin of neurosecretory vesicles extremely difficult. By comparing 28 proteins of the core neurosecretory vesicle proteome in 13 different species, we demonstrate that most of the proteins are already present in unicellular organisms. Surprisingly, we find that the vesicle residing SNARE protein synaptobrevin is localized to the vesicle-rich apical and basal pole in the choanoflagellate Salpingoeca rosetta. Our 3D vesicle reconstructions reveal that the choanoflagellates Salpingoeca rosetta and Monosiga brevicollis exhibit a polarized and diverse vesicular landscape. This study sheds light on the ancestral molecular machinery of neurosecretory vesicles and provides a framework to understand the origin and evolution of secretory cells, synapses, and neurons.

evolutionary biology↗

bioRxiv2020

Nitric oxide (NO) signaling in Trichoplax and related species: Microchemical characterization and the lineage-specific diversification

Nitric oxide (NO) is a free radical gaseous messenger with a broad distribution across the animal kingdom. However, the early evolution of nitric oxide-mediated signaling in animals is unclear due to limited information about prebilaterian metazoans such as placozoans. Here, we analyzed NO synthases (NOS) in four different species of placozoans (haplotypes H1, H2, H4, H13). In contrast to all other invertebrates studied, Hoilungia and Trichoplax have three distinct NOS genes, including PDZ domain-containing NOS. To characterize NOS activity in Trichoplax adhaerens, we used capillary electrophoresis for microchemical assays of NO-related metabolites. Specifically, we quantified nitrites (products of NO oxidation) and L-citrulline (co-product of NO synthesis from L-arginine), which were affected by NOS inhibitors confirming the presence of functional NOS. Next, using fluorescent single-molecule in situ hybridization, we showed that distinct NOSs are expressed in different subpopulations of cells, with a noticeable distribution close to the edge regions of Trichoplax. These data suggest the compartmentalized release of this messenger and a greater diversity of cell types in placozoans than anticipated. We also revealed a dramatic diversification of NO receptor machinery, including identification of both canonical and novel NIT-domain containing soluble guanylate cyclases as putative NO/nitrite/nitrate sensors. Thus, although Trichoplax is considered to be one of the morphologically simplest free-living animals, the complexity of NO-cGMP-mediated signaling is greater to those in vertebrates. This situation illuminates multiple lineage-specific diversifications of NOSs and NO/nitrite/nitrate sensors from the common ancestor of Metazoa. Short AbstractNitric oxide (NO) is a ubiquitous gaseous messenger, but we know little about its early evolution. Here, we analyzed NO synthases (NOS) in four different species of placozoans - one of the early-branching animal lineages. In contrast to other invertebrates studied, Trichoplax and Hoilungia have three distinct NOS genes, including PDZ domain-containing NOS. Using ultra-sensitive capillary electrophoresis assays, we quantified nitrites (products of NO oxidation) and L-citrulline (co-product of NO synthesis from L-arginine), which were affected by NOS inhibitors confirming the presence of functional enzymes in Trichoplax. Using fluorescent single-molecule in situ hybridization, we showed that distinct NOSs are expressed in different subpopulations of cells, with a noticeable distribution close to the edge regions of Trichoplax. These data suggest both the compartmentalized release of NO and a greater diversity of cell types in placozoans than anticipated. NO receptor machinery includes both canonical and novel NIT-domain containing soluble guanylate cyclases as putative NO/nitrite/nitrate sensors. Thus, although Trichoplax and Hoilungia exemplify the morphologically simplest free-living animals, the complexity of NO-cGMP-mediated signaling in Placozoa is greater to those in vertebrates. This situation illuminates multiple lineage-specific diversifications of NOSs and NO/nitrite/nitrate sensors from the common ancestor of Metazoa.

evolutionary biology↗

bioRxiv2020

A conformational switch driven by phosphorylation regulates Ykt6 activity in macroautophagy

Membrane fusion, an essential process in all eukaryotes, is driven by SNARE proteins. Ykt6 is an essential SNARE that plays critical roles throughout the secretory, endocytic, and autophagy pathways. Ykt6 activity is thought to be regulated by a conformational change from a closed cytosolic form to an open membrane-bound form, yet the mechanism that regulates this transition is unknown. Through genetic, pharmacologic, and structural modeling approaches in mammalian cells, we found that phosphorylation regulates Ykt6 conversion from a closed to an open state. The phosphorylation site we identified is highly conserved in evolution and is regulated by the Ca2+-dependent phosphatase, calcineurin. We found that phosphorylation is a key determinant for intracellular localization of Ykt6 and its function in macroautophagy. Our studies reveal a novel mechanism by which Ykt6 conformation and activity is regulated by Ca2+ signaling with implications in Parkinsons Disease in which Ykt6 has been shown to play a role.

cell biology↗

bioRxiv2019

Prototypic SNARE proteins are encoded in the genomes of Heimdallarchaeota, potentially bridging the gap between the prokaryotes and eukaryotes

A defining feature of eukaryotic cells is the presence of numerous membrane-bound organelles that subdivide the intracellular space into distinct compartments. How the eukaryotic cell acquired its internal complexity is still poorly understood. Material exchange among most organelles occurs via vesicles that bud off from a source and specifically fuse with a target compartment. Central players in the vesicle fusion process are the Soluble N-ethylmaleimide-sensitive factor Attachment protein REceptor (SNARE) proteins. These small tail-anchored (TA) membrane proteins zipper into elongated four-helix bundles that pull membranes together1-3. SNARE proteins are highly conserved among eukaryotes but are thought to be absent in prokaryotes. Here, we identified SNARE-like factors in the genomes of uncultured organisms of Asgard archaea of the Heimdallarchaeota clade4,5, which are thought to be the closest living relatives of eukaryotes. Biochemical experiments show that the archaeal SNARE-like proteins can interact with eukaryotic SNARE proteins. We did not detect SNAREs in -proteobacteria, the closest relatives of mitochondria, but identified several genes encoding for SNARE proteins in {gamma}-proteobacteria of the order Legionellales, pathogens that live inside eukaryotic cells. Very probably, their SNAREs stem from lateral gene transfer from eukaryotes. Together, this suggests that the diverse set of eukaryotic SNAREs evolved from an archaeal precursor. However, whether Heimdallarchaeota actually have a simplified endomembrane system will only be seen when we succeed studying these organisms under the microscope.

evolutionary biology↗

bioRxiv2018

High cell diversity and complex peptidergic signalling underlie placozoan behaviour

Placozoans, together with sponges, are the only animals devoid of a nervous system and muscles, yet both respond to sensory stimulation in a coordinated manner. How behavioural control in these free-living animals is achieved in the absence of neurons and, more fundamentally, how the first neurons evolved from more primitive communication cells during the rise of animals is not yet understood [1-5]. The placozoan Trichoplax adhaerens is a millimeter-wide, flat, free-living marine animal composed of six morphologically identified cell types distributed across a simple bodyplan [6-9]: a flat upper epithelium and a cylindrical lower epithelium interspersed with a loose layer of fiber cells. Its genome encodes several proneuropeptide genes and genes involved in neurosecretion in animals with a nervous system [10-12]. Here we investigate neuropeptide signalling in Trichoplax adhaerens. We found specific expression of several neuropeptides in non-overlapping cell populations distributed over the three cell layers, revealing an unsuspected cell-type diversity of Trichoplax adhaerens. Using live imaging, we uncovered that treatments with 11 different neuropeptides elicited striking and consistent effects on the animals shape, patterns of movement and velocity that we categorized under three main types: (i) crinkling, (ii) turning, and (iii) flattening and churning. Together, the data demonstrate a crucial role for peptidergic signalling in nerveless placozoans and suggest that peptidergic volume signalling may have predated synaptic signalling in the evolution of nervous systems.

neuroscience↗