@article{17885, abstract = {The formation of new ribosomes is tightly coordinated with cell growth and proliferation. In eukaryotes, the correct assembly of all ribosomal proteins and RNAs follows an intricate scheme of maturation and rearrangement steps across three cellular compartments: the nucleolus, nucleoplasm, and cytoplasm. We demonstrate that usnic acid, a lichen secondary metabolite, inhibits the maturation of the large ribosomal subunit in yeast. We combine biochemical characterization of pre-ribosomal particles with a quantitative single-particle cryo-EM approach to monitor changes in nucleolar particle populations upon drug treatment. Usnic acid rapidly blocks the transition from nucleolar state B to C of Nsa1-associated pre-ribosomes, depleting key maturation factors such as Dbp10 and hindering pre-rRNA processing. This primary nucleolar block rapidly rebounds on earlier stages of the pathway which highlights the regulatory linkages between different steps. In summary, we provide an in-depth characterization of the effect of usnic acid on ribosome biogenesis, which may have implications for its reported anti-cancer activities.}, author = {Kofler, Lisa and Grundmann, Lorenz and Gerhalter, Magdalena and Prattes, Michael and Merl-Pham, Juliane and Zisser, Gertrude and Grishkovskaya, Irina and Hodirnau, Victor-Valentin and Vareka, Martin and Breinbauer, Rolf and Hauck, Stefanie M. and Haselbach, David and Bergler, Helmut}, issn = {2041-1723}, journal = {Nature Communications}, publisher = {Springer Nature}, title = {{The novel ribosome biogenesis inhibitor usnic acid blocks nucleolar pre-60S maturation}}, doi = {10.1038/s41467-024-51754-3}, volume = {15}, year = {2024}, } @misc{23023, abstract = {In this repository, you will find the R-scripts used to generate the figures, along with the data necessary to make the plots. You will also find the Python-scripts used to compute the pair-wise distance between embryos, to estimate the surface, the volume, the α-parameter, to identify the closest rigid packing and to generate the morphomap. Additionally, you have access to the base model used with Plantseg to predict and label the cells at the 8-cell stage. Note that a more accurate and more specific model was generated for each embryo based on the curation of at least 5 timepoints. Figures The plots in Figure 1-7 and Figure S1-S7 can be generated with the respective files. The code is written in R and requires RStudio to work out-of-the-box. The R scripts are located in figures/. Figures and supplemental figures are generated with the same file (e.g., Figure 1.R generates all the computed panels of Figure 1 and Figure S1). Data required to generate the plots are located in figures/data/. Note that the code to work with the morphomap and the tracking data is located in local packages figures/fabreges.morphomap and figures/fabreges.tracking. An additional package figures/fabreges.generic contains utility functions. Python packages For the needs of this study, we developed a series of package for Python located in python-packages/. The packages can be directly imported within your scripts. Along with the packages, you will find 6 scripts used to compute, extract or estimate the data from the source. The folder is organised as follow: package interfaces is located in python-packages/interfaces/ and can be tested by running the scripts to extract the angles an estimate the surface of contact (compute-angles.py and estimate-contacts.py respectively) in folder python-packages, package morphomap is located in python-packages/morphomap/ and can be tested by running the script to compute the geometrical distances between embryos, project the data in 2D and compute the volume of the cells (compute-distances.py, process-distances.py and compute-volumes.py respectively) in folder python-packages, package splinefit is included in morphomap in python-packages/morphomap/splinefit/, but can function independently, package rbb is located in python-packages/rbb/ and can be tested by running the script to identify the closest rigid packing of an embryo at the 8-cell stage (identify-packing.py) in folder python-packages. We provide example of segmented data in python-packages/samples/. CNN We provide a convolutional neural network (CNN) used in our study, located in cnn. Please note that we optimised the CNN further for each dataset by doing manual curation of at least 5 time points and by retraining the network with those ground truth. The CNN were trained using pytorch-3dunet. The CNN can be used with Plantseg. Currently (plantseg v.1.6.0 and pytorch-3dunet v.1.6.0), the output model from pytorch-3dunet must currently be modified to work with plantseg: # python import torch for model in ('best_checkpoint.pytorch', 'last_checkpoint.pytorch'): state = torch.load(model, map_location='cpu') state = state['model_state_dict'] torch.save(state, model) Requierements Figures The figures were generated with R (v. 4.4.0) and RStudio (v. 2024.04.1+748). The following packages must be installed prior to execution: alphashape3d (v. 1.3.2) devtools (v. 2.4.5) ggplot2 (v. 3.5.1) ggrepel (v. 0.9.5) rstudioapi (v. 0.16.0) To install all these packages, run the following command in the RStudio console: # R install.packages(c("alphashape3d@1.3.2", "devtools@2.4.5", "ggplot2@3.5.1", "ggrepel@0.9.5", "rstudioapi@0.16")) More recent version of R, RStudio and the packages may also work (not tested). Python packages Python 3.12.3 was used to analyse the data. The scripts may work with more recent version of Python and with Python 3.8 or earlier (not tested). To run the code, you must first install the following packages: numpy (v. 1.26.4) imageio (v. 2.34.1) scipy (v. 1.13.1) scikit-image (v. 0.23.2) scikit-learn (v. 1.5.0) Alternatively, you can create a conda environment: # Shell script conda create -y --name fabreges2024 python=3.12.3 conda activate fabreges2024 python -m pip install numpy==1.26.4 imageio==2.34.1 scipy==1.13.1 scikit-image==0.23.2 scikit-learn==1.5.0 More recent version of the dependencies may work (not tested). Usage Figures Open one of the figure script in RStudio and run the entire code. You may have to create a folder output in figures/ if it does not exist. The code has local dependencies that will be installed on your computer when encountering them for the first time. They start with fabreges.* and are necessary to generate the figures. Python packages (example) Compute angles # Shell script # Compute a summary (mean, median, s.d.) of the angles in embryo C1 at time 40 python compute-angles.py -s -r 15 -R 20 samples/C1_t040.tif.gz output/C1_t040_angles.csv # Compute a detailed list of angle values python compute-angles.py -r 15 -R 20 samples/C1_t041.tif.gz output/C1_t041_angles.csv Compute distances # Shell script # Compute the distance between two embryos python compute-distances.py -c 5 -s 5 samples/C1_t040.tif.gz samples/C1_t041.tif.gz output/distances.csv # Append another distance measurement python compute-distances.py -a -c 5 -s 5 samples/C1_t040.tif.gz samples/C1_t042.tif.gz output/distances.csv Compute volumes # Shell script # Compute the volume in isotropic data python compute-volumes.py -r 0.416 0.416 0.416 samples/C1_t040.tif.gz output/volumes.csv Estimate contacts # Shell script # Estimate the contacts and the free surface area python estimate-contacts.py -r 0.416 0.416 0.416 samples/C1_t040.tif.gz output/contacts.csv Identify the closest rigid packing # Shell script # Assuming the file `output/contacts.csv` exists and contains the output # from command `estimate-contacts.py`. # Identify the closest rigid packings python identify-packing.py output/contacts.csv output/backbones.csv Generate the seeds for Surface Evolver # Shell script # Assuming the file `output/contacts.csv` exists and contains the output # from command `estimate-contacts.py`, and the file `output/volumes.csv` # exists and contains the output from command `compute-volumes.py` # Generate the seeds for Surface Evolver using automatic thresholding # Automatic threshold will determine the highest cell-cell contact threshold # that preserves rigidity of the embryo (or 0 if not rigid) python generate-seed.py output/contacts.csv output/volumes.csv output/seed.fe # Generate the seeds for Surface Evolver with custom threshold python generate-seed.py --threshold 0.1 output/contacts.csv output/volumes.csv output/seed.fe Project in 2D and generate a distance matrix # Shell script # Assuming the folder `foo` only contains one or more CSV # from command `compute-distances.py`. # Compute the tSNE 2D projection python process-distances.py -D2 -t output/tsne_2d.csv foo/ # Compute the tSNE 3D projection python process-distances.py -D3 -t output/tsne_2d.csv foo/ # Generate the distance matrix python process-distances.py -m output/dismat.csv foo/}, author = {Corominas-Murtra, Bernat and Moghe, Prachiti and Kickuth, Alison and Ichikawa, Takafumi and Iwatani, Chizuru and Tsukiyama, Tomoyuki and Daniel, Nathalie and Gering, Julie and Stokkermans, Anniek and Wolny, Adrian and Kreshuk, Anna and Duranthon, Véronique and Uhlmann, Virginie and Hannezo, Edouard and Hiiragi, Takashi}, publisher = {Institute of Science and Technology Austria}, title = {{Data and software from: Temporal variability and cell mechanics control robustness in mammalian embryogenesis}}, doi = {10.5281/zenodo.12819780}, year = {2024}, } @inproceedings{18913, abstract = {With the proliferation of blockchain technology in high-value sectors, consensus protocols are becoming critical infrastructures. The rapid innovation cycle in Byzantine fault tolerant (BFT) consensus protocols has culminated in HotStuff, which provides linear message complexity in the partially synchronous setting. To achieve this, HotStuff leverages a leader that collects, aggregates, and broadcasts the messages of other validators. This paper analyzes the security implications of such approaches in practice, from the perspective of liveness and availability. By implementing attacks in a globally-distributed testbed, we show that state-of-the-art leader-based protocols are vulnerable to denial-of-service (DoS) attacks on the leader. Our attacks, demonstrated on committees of up to 64 validators, manage to disrupt liveness within seconds, using only a few tens of Mbps of attack bandwidth per validator. Crucially, the cost and effectiveness of the attacks are independent of the committee size. Based on the outcome of these experiments, we then propose and test effective mitigations. Our findings show that advancements in both protocol design and network-layer defenses can greatly improve the practical resilience of BFT consensus protocols.}, author = {Giuliari, Giacomo and Sonnino, Alberto and Frei, Marc and Streun, Fabio and Kokoris Kogias, Eleftherios and Perrig, Adrian}, booktitle = {Proceedings of the 19th ACM Asia Conference on Computer and Communications Security}, isbn = {9798400704826}, location = {Singapore, Singapore}, pages = {1345--1360}, publisher = {ACM}, title = {{An empirical study of consensus protocols’ DoS resilience}}, doi = {10.1145/3634737.3656997}, year = {2024}, } @inproceedings{18957, abstract = {Sui Lutris is the first smart-contract platform to sustainably achieve sub-second finality. It achieves this significant decrease by employing consensusless agreement not only for simple payments but for a large variety of transactions. Unlike prior work, Sui Lutris neither compromises expressiveness nor throughput and can run perpetually without restarts. Sui Lutris achieves this by safely integrating consensuless agreement with a high-throughput consensus protocol that is invoked out of the critical finality path but ensures that when a transaction is at risk of inconsistent concurrent accesses, its settlement is delayed until the total ordering is resolved. Building such a hybrid architecture is especially delicate during reconfiguration events, where the system needs to preserve the safety of the consensusless path without compromising the long-term liveness of potentially misconfigured clients. We thus develop a novel reconfiguration protocol, the first to provably show the safe and efficient reconfiguration of a consensusless blockchain. Sui Lutris is currently running in production and underpins the Sui smart-contract platform. Combined with the use of Objects instead of accounts it enables the safe execution of smart contracts that expose objects as a first-class resource. In our experiments Sui Lutris achieves latency lower than 0.5 seconds for throughput up to 5,000 certificates per second (150k ops/s with transaction blocks), compared to the state-of-the-art real-world consensus latencies of 3 seconds. Furthermore, it gracefully handles validators crash-recovery and does not suffer visible performance degradation during reconfiguration.}, author = {Blackshear, Sam and Chursin, Andrey and Danezis, George and Kichidis, Anastasios and Kokoris Kogias, Eleftherios and Li, Xun and Logan, Mark and Menon, Ashok and Nowacki, Todd and Sonnino, Alberto and Williams, Brandon and Zhang, Lu}, booktitle = {Proceedings of the 2024 on ACM SIGSAC Conference on Computer and Communications Security}, isbn = {9798400706363}, location = {Salt Lake City, UT, United States}, pages = {2606--2620}, publisher = {ACM}, title = {{Sui Lutris: A blockchain combining broadcast and consensus}}, doi = {10.1145/3658644.3670286}, year = {2024}, } @article{17269, abstract = {The directed migration of epithelial cell collectives through coordinated movements plays a crucial role in various physiological processes and is increasingly understood at the level of large confluent monolayers. However, numerous processes rely on the migration of small groups of polarized epithelial clusters in complex environments, and their responses to external geometries remain poorly understood. To address this, we cultivate primary epithelial keratocyte tissues on adhesive microstripes to create autonomous epithelial clusters with well-defined geometries. We show that their migration efficiency is strongly influenced by the contact geometry and the orientation of cell–cell contacts with respect to the direction of migration. A combination of velocity and polarity alignment with contact regulation of locomotion in an active matter model captures quantitatively the experimental data. Furthermore, we predict that this combination of rules enables efficient navigation in complex geometries, which we confirm experimentally. Altogether, our findings provide a conceptual framework for extracting the interaction rules of active systems from their interaction with physical boundaries, as well as design principles for collective navigation in complex microenvironments.}, author = {Vercruysse, Eléonore and Brückner, David and Gómez-González, Manuel and Remson, Alexandre and Luciano, Marine and Kalukula, Yohalie and Rossetti, Leone and Trepat, Xavier and Hannezo, Edouard B and Gabriele, Sylvain}, issn = {1745-2481}, journal = {Nature Physics}, pages = {1492--1500}, publisher = {Springer Nature}, title = {{Geometry-driven migration efficiency of autonomous epithelial cell clusters}}, doi = {10.1038/s41567-024-02532-x}, volume = {20}, year = {2024}, } @article{18446, abstract = {How living systems achieve precision in form and function despite their intrinsic stochasticity is a fundamental yet ongoing question in biology. We generated morphomaps of preimplantation embryogenesis in mouse, rabbit, and monkey embryos, and these morphomaps revealed that although blastomere divisions desynchronized passively, 8-cell embryos converged toward robust three-dimensional shapes. Using topological analysis and genetic perturbations, we found that embryos progressively changed their cellular connectivity to a preferred topology, which could be predicted by a physical model in which actomyosin contractility and noise facilitate topological transitions, lowering surface energy. This mechanism favored regular embryo packing and promoted a higher number of inner cells in the 16-cell embryo. Synchronized division reduced embryo packing and generated substantially more misallocated cells and fewer inner-cell–mass cells. These findings suggest that stochasticity in division timing contributes to robust patterning.}, author = {Fabrèges, Dimitri and Corominas-Murtra, Bernat and Moghe, Prachiti and Kickuth, Alison and Ichikawa, Takafumi and Iwatani, Chizuru and Tsukiyama, Tomoyuki and Daniel, Nathalie and Gering, Julie and Stokkermans, Anniek and Wolny, Adrian and Kreshuk, Anna and Duranthon, Véronique and Uhlman, Virginie and Hannezo, Edouard B and Hiiragi, Takashi}, issn = {1095-9203}, journal = {Science}, number = {6718}, publisher = {AAAS}, title = {{Temporal variability and cell mechanics control robustness in mammalian embryogenesis}}, doi = {10.1126/science.adh1145}, volume = {386}, year = {2024}, } @inproceedings{18071, abstract = {Recent advancements on DAG-based consensus protocols allow for blockchains with improved metrics and properties, such as throughput and censorship-resistance. Variants of the Bullshark [18] consensus protocol are adopted for practical use by the Sui blockchain, for improved latency. However, the protocol is leader-based, and is strongly affected by crashed leaders that can lead to various performance issues, for example, decreased transaction throughput. In this paper, we propose HammerHead, a DAG-based consensus protocol, that is inspired by Carousel [8] and provides Leader-Utilization. Our proposal differs from Carousel, which is built for a chained consensus protocol; in HammerHead chain quality is inherited by the DAG. HammerHead needs to preserve safety and liveness, despite validators committing leader vertices asynchronously. The key idea is to update leader schedules dynamically, based on the validators' scores during the previous schedule. We implement HammerHead and show a minor improvement in performance for cases without faults. The major improvements in comparison to Bullshark appear in faulty settings. Specifically, we show a drastic, 2x-latency improvement and up to 40% increased throughput when crash faults occur (100 validators, 33 faults).}, author = {Tsimos, Giorgos and Kichidis, Anastasios and Sonnino, Alberto and Kokoris Kogias, Eleftherios}, booktitle = {Proceedings - International Conference on Distributed Computing Systems}, isbn = {9798350386059}, issn = {2575-8411}, location = {Jersey City, NJ, United States}, pages = {1377--1387}, publisher = {IEEE}, title = {{HammerHead: Leader reputation for dynamic scheduling}}, doi = {10.1109/ICDCS60910.2024.00129}, year = {2024}, } @article{17284, abstract = {Platelet homeostasis is essential for vascular integrity and immune defence1,2. Although the process of platelet formation by fragmenting megakaryocytes (MKs; thrombopoiesis) has been extensively studied, the cellular and molecular mechanisms required to constantly replenish the pool of MKs by their progenitor cells (megakaryopoiesis) remains unclear3,4. Here we use intravital imaging to track the cellular dynamics of megakaryopoiesis over days. We identify plasmacytoid dendritic cells (pDCs) as homeostatic sensors that monitor the bone marrow for apoptotic MKs and deliver IFNα to the MK niche triggering local on-demand proliferation and maturation of MK progenitors. This pDC-dependent feedback loop is crucial for MK and platelet homeostasis at steady state and under stress. pDCs are best known for their ability to function as vigilant detectors of viral infection5. We show that virus-induced activation of pDCs interferes with their function as homeostatic sensors of megakaryopoiesis. Consequently, activation of pDCs by SARS-CoV-2 leads to excessive megakaryopoiesis. Together, we identify a pDC-dependent homeostatic circuit that involves innate immune sensing and demand-adapted release of inflammatory mediators to maintain homeostasis of the megakaryocytic lineage.}, author = {Gärtner, Florian R and Ishikawa-Ankerhold, Hellen and Stutte, Susanne and Fu, Wenwen and Weitz, Jutta and Dueck, Anne and Nelakuditi, Bhavishya and Fumagalli, Valeria and Van Den Heuvel, Dominic and Belz, Larissa and Sobirova, Gulnoza and Zhang, Zhe and Titova, Anna and Navarro, Alejandro Martinez and Pekayvaz, Kami and Lorenz, Michael and Von Baumgarten, Louisa and Kranich, Jan and Straub, Tobias and Popper, Bastian and Zheden, Vanessa and Kaufmann, Walter and Guo, Chenglong and Piontek, Guido and Von Stillfried, Saskia and Boor, Peter and Colonna, Marco and Clauß, Sebastian and Schulz, Christian and Brocker, Thomas and Walzog, Barbara and Scheiermann, Christoph and Aird, William C. and Nerlov, Claus and Stark, Konstantin and Petzold, Tobias and Engelhardt, Stefan and Sixt, Michael K and Hauschild, Robert and Rudelius, Martina and Oostendorp, Robert A.J. and Iannacone, Matteo and Heinig, Matthias and Massberg, Steffen}, issn = {1476-4687}, journal = {Nature}, pages = {645--653}, publisher = {Springer Nature}, title = {{Plasmacytoid dendritic cells control homeostasis of megakaryopoiesis}}, doi = {10.1038/s41586-024-07671-y}, volume = {631}, year = {2024}, } @article{18920, abstract = {The globally distributed marine alga Emiliania huxleyi has cooling effect on the Earth’s climate. The population density of E. huxleyi is restricted by Nucleocytoviricota viruses, including E. huxleyi virus 201 (EhV-201). Despite the impact of E. huxleyi viruses on the climate, there is limited information about their structure and replication. Here, we show that the dsDNA genome inside the EhV-201 virion is protected by an inner membrane, capsid, and outer membrane. EhV-201 virions infect E. huxleyi by using fivefold vertices to bind to and fuse the virus’ inner membrane with the cell plasma membrane. Progeny virions assemble in the cytoplasm at the surface of endoplasmic reticulum–derived membrane segments. Genome packaging initiates synchronously with the capsid assembly and completes through an aperture in the forming capsid. The genome-filled capsids acquire an outer membrane by budding into intracellular vesicles. EhV-201 infection induces a loss of surface protective layers from E. huxleyi cells, which enables the continuous release of virions by exocytosis.}, author = {Homola, Miroslav and Büttner, Renate Carina and Füzik, Tibor and Křepelka, Pavel and Holbová, Radka and Nováček, Jiří and Chaillet, Marten L. and Žák, Jakub and Grybchuk, Danyil and Förster, Friedrich and Wilson, William H. and Schroeder, Declan C. and Plevka, Pavel}, issn = {2375-2548}, journal = {Science Advances}, number = {15}, publisher = {American Association for the Advancement of Science}, title = {{Structure and replication cycle of a virus infecting climate-modulating alga Emiliania huxleyi}}, doi = {10.1126/sciadv.adk1954}, volume = {10}, year = {2024}, } @inbook{18052, abstract = {Sodium dodecyl sulfate-digested freeze-fracture replica labeling (SDS-FRL) is an electron microscope (EM) sample preparation technique which allows for high-resolution visualization of membrane proteins with high sensitivity. However, image acquisition of specific replica profiles such as synapses in a large field of EM view needs a valid experience and a long time for manual searching. Here, we describe how to utilize deep learning for automatizing image acquisition of specific profiles of interest in replica samples. This protocol facilitates the labor-intensive collection of EM images, in particular for rare profiles. We provide instructions for using SerialEM image acquisition software in conjunction with object detection by our newly developed deep learning software DarEM, to automatically acquire tilt series of all synapses in a selected region. We then show how to perform a mostly automated analysis of gold particle labeling in the acquired images by utilizing Darea software.}, author = {Kleindienst, David and Costanzo, Tommaso and Shigemoto, Ryuichi}, booktitle = {New Aspects in Analyzing the Synaptic Organization of the Brain}, editor = {Lübke, Joachim H.R. and Rollenhagen, Astrid}, isbn = {9781071640180}, issn = {1940-6045}, pages = {123--137}, publisher = {Springer Nature}, title = {{Automated Imaging and Analysis of Synapses in Freeze-Fracture Replica Samples with Deep Learning}}, doi = {10.1007/978-1-0716-4019-7_8}, year = {2024}, } @misc{14842, abstract = {Eva Benkova received a PhD in Biophysics at the Institute of Biophysics of the Czech Academy of Sciences in 1998. After working as a postdoc at the Max Planck Institute in Cologne and the Center for Plant Molecular Biology (ZMBP) in Tübingen, she became a group leader at the Plant Systems Biology Department of the Vlaams Instituut voor Biotechnologie (VIB) in Gent. In 2012, she transitioned to an Assistant Professor position at the Institute of Science and Technology Austria (ISTA) where she was later promoted to Professor. Since 2021, she has served as the Dean of the ISTA Graduate School. As a plant developmental biologist, she focuses on unraveling the molecular mechanisms and principles that underlie hormonal interactions in plants. In her current work, she explores the intricate connections between hormones and regulatory pathways that mediate the perception of environmental stimuli, including abiotic stress and nitrate availability.}, author = {Benková, Eva}, booktitle = {Current Biology}, issn = {1879-0445}, number = {1}, pages = {R3--R5}, publisher = {Elsevier}, title = {{Eva Benkova}}, doi = {10.1016/j.cub.2023.11.039}, volume = {34}, year = {2024}, } @misc{23025, abstract = {The globally distributed marine alga Emiliania huxleyi produces reflective calcite disks (coccoliths) that increase the albedo of ocean water and thus reduce the heat absorption in the ocean, which cools the Earth’s climate. The population density of E. huxleyi is restricted by Nucleocytoviricota viruses, including E. huxleyi virus 201 (EhV-201). Despite the impact of E. huxleyi viruses on the climate, there is limited information about their structure and replication. Here we show that the dsDNA genome inside the EhV-201 virion is protected by an inner membrane, capsid, and outer membrane decorated with numerous transmembrane proteins. The virions are prone to deformation, and parts of their capsids deviate from the icosahedral arrangement. EhV-201 virions infect E. huxleyi by using their fivefold vertex to bind to a host cell and fuse the virus’s inner membrane with the plasma membrane. Whereas the replication of EhV-201 probably occurs in the nucleus, virions assemble in the cytoplasm at the surface of endoplasmic reticulum-derived membrane segments. Genome packaging initiates synchronously with the capsid assembly and completes through an aperture in the forming capsid. Upon the completion of genome packaging, the capsids change conformation, which enables them to acquire an outer membrane by budding into intracellular vesicles. EhV-201 infection induces a loss of surface protective layers from E. huxleyi cells, which allows the continuous release of virions by exocytosis. Our results provide insight into how EhVs bypass the surface protective layers of E. huxleyi and exploit the organelles of an infected cell for progeny assembly.}, author = {Homola, Miroslav and Büttner, Carina and Füzik, Tibor and Krepelka, Pavel and Holbová, Radka and Chaillet, Marten and Nováček, Jiří and Förster, Friedrich and Wilson, William and Schroeder, Declan and Plevka, Pavel}, publisher = {Dryad}, title = {{Data: Structure and replication cycle of a virus infecting climate-modulating alga Emiliania huxleyi}}, doi = {10.5061/dryad.p5hqbzkw0}, year = {2024}, } @article{15146, abstract = {The extracellular matrix (ECM) serves as a scaffold for cells and plays an essential role in regulating numerous cellular processes, including cell migration and proliferation. Due to limitations in specimen preparation for conventional room-temperature electron microscopy, we lack structural knowledge on how ECM components are secreted, remodeled, and interact with surrounding cells. We have developed a 3D-ECM platform compatible with sample thinning by cryo-focused ion beam milling, the lift-out extraction procedure, and cryo-electron tomography. Our workflow implements cell-derived matrices (CDMs) grown on EM grids, resulting in a versatile tool closely mimicking ECM environments. This allows us to visualize ECM for the first time in its hydrated, native context. Our data reveal an intricate network of extracellular fibers, their positioning relative to matrix-secreting cells, and previously unresolved structural entities. Our workflow and results add to the structural atlas of the ECM, providing novel insights into its secretion and assembly.}, author = {Zens, Bettina and Fäßler, Florian and Hansen, Jesse and Hauschild, Robert and Datler, Julia and Hodirnau, Victor-Valentin and Zheden, Vanessa and Alanko, Jonna H and Sixt, Michael K and Schur, Florian KM}, issn = {1540-8140}, journal = {Journal of Cell Biology}, number = {6}, publisher = {Rockefeller University Press}, title = {{Lift-out cryo-FIBSEM and cryo-ET reveal the ultrastructural landscape of extracellular matrix}}, doi = {10.1083/jcb.202309125}, volume = {223}, year = {2024}, } @article{17463, abstract = {Allosteric modulation of protein function, wherein the binding of an effector to a protein triggers conformational changes at distant functional sites, plays a central part in the control of metabolism and cell signalling1,2,3. There has been considerable interest in designing allosteric systems, both to gain insight into the mechanisms underlying such ‘action at a distance’ modulation and to create synthetic proteins whose functions can be regulated by effectors4,5,6,7. However, emulating the subtle conformational changes distributed across many residues, characteristic of natural allosteric proteins, is a significant challenge8,9. Here, inspired by the classic Monod–Wyman–Changeux model of cooperativity10, we investigate the de novo design of allostery through rigid-body coupling of peptide-switchable hinge modules11 to protein interfaces12 that direct the formation of alternative oligomeric states. We find that this approach can be used to generate a wide variety of allosterically switchable systems, including cyclic rings that incorporate or eject subunits in response to peptide binding and dihedral cages that undergo effector-induced disassembly. Size-exclusion chromatography, mass photometry13 and electron microscopy reveal that these designed allosteric protein assemblies closely resemble the design models in both the presence and absence of peptide effectors and can have ligand-binding cooperativity comparable to classic natural systems such as haemoglobin14. Our results indicate that allostery can arise from global coupling of the energetics of protein substructures without optimized side-chain–side-chain allosteric communication pathways and provide a roadmap for generating allosterically triggerable delivery systems, protein nanomachines and cellular feedback control circuitry.}, author = {Pillai, Arvind and Idris, Abbas and Philomin, Annika and Weidle, Connor and Skotheim, Rebecca and Leung, Philip J.Y. and Broerman, Adam and Demakis, Cullen and Borst, Andrew J. and Praetorius, Florian M and Baker, David}, issn = {1476-4687}, journal = {Nature}, pages = {911–920 }, publisher = {Springer Nature}, title = {{De novo design of allosterically switchable protein assemblies}}, doi = {10.1038/s41586-024-07813-2}, volume = {632}, year = {2024}, } @article{18934, abstract = {The assembly of biomolecular condensate in eukaryotic cells and the accumulation of amyloid deposits in neurons are processes involving the nucleation and growth (NAG) of new protein phases. To therapeutically target protein phase separation, drug candidates are tested in in vitro assays that monitor the increase in the mass or size of the new phase. Limited mechanistic insight is, however, provided if empirical or untestable kinetic models are fitted to these progress curves. Here we present the web server NAGPKin that quantifies NAG rates using mass-based or size-based progress curves as the input data. A report is generated containing the fitted NAG parameters and elucidating the phase separation mechanisms at play. The NAG parameters can be used to predict particle size distributions of, for example, protein droplets formed by liquid-liquid phase separation (LLPS) or amyloid fibrils formed by protein aggregation. Because minimal intervention is required from the user, NAGPKin is a good platform for standardized reporting of LLPS and protein self-assembly data. NAGPKin is useful for drug discovery as well as for fundamental studies on protein phase separation. NAGPKin is freely available (no login required) at https://nagpkin.i3s.up.pt .}, author = {Sárkány, Zsuzsa and Figueiredo, Francisco and Macedo-Ribeiro, Sandra and Martins, Pedro M.}, issn = {1939-4586}, journal = {Molecular Biology of the Cell}, number = {3}, publisher = {American Society for Cell Biology}, title = {{NAGPKin: Nucleation-and-growth parameters from the kinetics of protein phase separation}}, doi = {10.1091/mbc.e23-07-0289}, volume = {35}, year = {2024}, } @article{18757, abstract = {Segmentation is a critical data processing step in many applications of cryo-electron tomography. Downstream analyses, such as subtomogram averaging, are often based on segmentation results, and are thus critically dependent on the availability of open-source software for accurate as well as high-throughput tomogram segmentation. There is a need for more user-friendly, flexible, and comprehensive segmentation software that offers an insightful overview of all steps involved in preparing automated segmentations. Here, we present Ais: a dedicated tomogram segmentation package that is geared towards both high performance and accessibility, available on GitHub. In this report, we demonstrate two common processing steps that can be greatly accelerated with Ais: particle picking for subtomogram averaging, and generating many-feature segmentations of cellular architecture based on in situ tomography data. Featuring comprehensive annotation, segmentation, and rendering functionality, as well as an open repository for trained models at aiscryoet.org, we hope that Ais will help accelerate research and dissemination of data involving cryoET.}, author = {Last, Mart G.F. and Abendstein, Leoni and Voortman, Lenard M. and Sharp, Thomas H.}, issn = {2050-084X}, journal = {eLife}, publisher = {eLife Sciences Publications}, title = {{Streamlining segmentation of cryo-electron tomography datasets with Ais}}, doi = {10.7554/eLife.98552}, volume = {13}, year = {2024}, } @article{18445, abstract = {Acquisition of specialized cellular features is controlled by the ordered expression of transcription factors (TFs) along differentiation trajectories. Here, we find a member of the Onecut TF family, ONECUT3, expressed in postmitotic neurons that leave their Ascl1+/Onecut1/2+ proliferative domain in the vertebrate hypothalamus to instruct neuronal differentiation. We combined single-cell RNA-seq and gain-of-function experiments for gene network reconstruction to show that ONECUT3 affects the polarization and morphogenesis of both hypothalamic GABA-derived dopamine and thyrotropin-releasing hormone (TRH)+ glutamate neurons through neuron navigator-2 (NAV2). In vivo, siRNA-mediated knockdown of ONECUT3 in neonatal mice reduced NAV2 mRNA, as well as neurite complexity in Onecut3-containing neurons, while genetic deletion of Onecut3/ceh-48 in C. elegans impaired neurocircuit wiring, and sensory discrimination-based behaviors. Thus, ONECUT3, conserved across neuronal subtypes and many species, underpins the polarization and morphological plasticity of phenotypically distinct neurons that descend from a common pool of Ascl1+ progenitors in the hypothalamus.}, author = {Zupančič, Maja and Keimpema, Erik and Tretiakov, Evgenii O. and Eder, Stephanie J. and Lev, Itamar and Englmaier, Lukas and Bhandari, Pradeep and Fietz, Simone A. and Härtig, Wolfgang and Renaux, Estelle and Villunger, Andreas and Hökfelt, Tomas and Zimmer, Manuel and Clotman, Frédéric and Harkany, Tibor}, issn = {2041-1723}, journal = {Nature Communications}, publisher = {Springer Nature}, title = {{Concerted transcriptional regulation of the morphogenesis of hypothalamic neurons by ONECUT3}}, doi = {10.1038/s41467-024-52762-z}, volume = {15}, year = {2024}, } @inproceedings{14213, abstract = {We introduce a method to segment the visual field into independently moving regions, trained with no ground truth or supervision. It consists of an adversarial conditional encoder-decoder architecture based on Slot Attention, modified to use the image as context to decode optical flow without attempting to reconstruct the image itself. In the resulting multi-modal representation, one modality (flow) feeds the encoder to produce separate latent codes (slots), whereas the other modality (image) conditions the decoder to generate the first (flow) from the slots. This design frees the representation from having to encode complex nuisance variability in the image due to, for instance, illumination and reflectance properties of the scene. Since customary autoencoding based on minimizing the reconstruction error does not preclude the entire flow from being encoded into a single slot, we modify the loss to an adversarial criterion based on Contextual Information Separation. The resulting min-max optimization fosters the separation of objects and their assignment to different attention slots, leading to Divided Attention, or DivA. DivA outperforms recent unsupervised multi-object motion segmentation methods while tripling run-time speed up to 104FPS and reducing the performance gap from supervised methods to 12% or less. DivA can handle different numbers of objects and different image sizes at training and test time, is invariant to permutation of object labels, and does not require explicit regularization.}, author = {Lao, Dong and Hu, Zhengyang and Locatello, Francesco and Yang, Yanchao and Soatto, Stefano}, booktitle = {1st Conference on Parsimony and Learning}, location = {Hong Kong, China}, title = {{Divided attention: Unsupervised multi-object discovery with contextually separated slots}}, year = {2024}, } @inproceedings{18114, abstract = {This paper presents Mechanistic Neural Networks, a neural network design for machine learning applications in the sciences. It incorporates a new Mechanistic Block in standard architectures to explicitly learn governing differential equations as representations, revealing the underlying dynamics of data and enhancing interpretability and efficiency in data modeling. Central to our approach is a novel Relaxed Linear Programming Solver (NeuRLP) inspired by a technique that reduces solving linear ODEs to solving linear programs. This integrates well with neural networks and surpasses the limitations of traditional ODE solvers enabling scalable GPU parallel processing. Overall, Mechanistic Neural Networks demonstrate their versatility for scientific machine learning applications, adeptly managing tasks from equation discovery to dynamic systems modeling. We prove their comprehensive capabilities in analyzing and interpreting complex scientific data across various applications, showing significant performance against specialized state-of-the-art methods. Source code is available at https://github.com/alpz/mech-nn.}, author = {Pervez, Adeel A and Locatello, Francesco and Gavves, Efstratios}, booktitle = {Proceedings of the 41st International Conference on Machine Learning}, issn = {2640-3498}, location = {Vienna, Austria}, pages = {40484--40501}, publisher = {ML Research Press}, title = {{Mechanistic neural networks for scientific machine learning}}, volume = {235}, year = {2024}, } @article{15257, abstract = {Root gravitropic bending represents a fundamental aspect of terrestrial plant physiology. Gravity is perceived by sedimentation of starch-rich plastids (statoliths) to the bottom of the central root cap cells. Following gravity perception, intercellular auxin transport is redirected downwards leading to an asymmetric auxin accumulation at the lower root side causing inhibition of cell expansion, ultimately resulting in downwards bending. How gravity-induced statoliths repositioning is translated into asymmetric auxin distribution remains unclear despite PIN auxin efflux carriers and the Negative Gravitropic Response of roots (NGR) proteins polarize along statolith sedimentation, thus providing a plausible mechanism for auxin flow redirection. In this study, using a functional NGR1-GFP construct, we visualized the NGR1 localization on the statolith surface and plasma membrane (PM) domains in close proximity to the statoliths, correlating with their movements. We determined that NGR1 binding to these PM domains is indispensable for NGR1 functionality and relies on cysteine acylation and adjacent polybasic regions as well as on lipid and sterol PM composition. Detailed timing of the early events following graviperception suggested that both NGR1 repolarization and initial auxin asymmetry precede the visible PIN3 polarization. This discrepancy motivated us to unveil a rapid, NGR-dependent translocation of PIN-activating AGCVIII kinase D6PK towards lower PMs of gravity-perceiving cells, thus providing an attractive model for rapid redirection of auxin fluxes following gravistimulation.}, author = {Kulich, Ivan and Schmid, Julia and Teplova, Anastasiia and Qi, Linlin and Friml, Jiří}, issn = {2050-084X}, journal = {eLife}, keywords = {General Immunology and Microbiology, General Biochemistry, Genetics and Molecular Biology, General Medicine, General Neuroscience}, publisher = {eLife Sciences Publications}, title = {{Rapid translocation of NGR proteins driving polarization of PIN-activating D6 protein kinase during root gravitropism}}, doi = {10.7554/elife.91523}, volume = {12}, year = {2024}, }