TY - JOUR AB - Metazoan development relies on the formation and remodeling of cell-cell contacts. Dynamic reorganization of adhesion receptors and the actomyosin cell cortex in space and time plays a central role in cell-cell contact formation and maturation. Nevertheless, how this process is mechanistically achieved when new contacts are formed remains unclear. Here, by building a biomimetic assay composed of progenitor cells adhering to supported lipid bilayers functionalized with E-cadherin ectodomains, we show that cortical F-actin flows, driven by the depletion of myosin-2 at the cell contact center, mediate the dynamic reorganization of adhesion receptors and cell cortex at the contact. E-cadherin-dependent downregulation of the small GTPase RhoA at the forming contact leads to both a depletion of myosin-2 and a decrease of F-actin at the contact center. At the contact rim, in contrast, myosin-2 becomes enriched by the retraction of bleb-like protrusions, resulting in a cortical tension gradient from the contact rim to its center. This tension gradient, in turn, triggers centrifugal F-actin flows, leading to further accumulation of F-actin at the contact rim and the progressive redistribution of E-cadherin from the contact center to the rim. Eventually, this combination of actomyosin downregulation and flows at the contact determines the characteristic molecular organization, with E-cadherin and F-actin accumulating at the contact rim, where they are needed to mechanically link the contractile cortices of the adhering cells. AU - Arslan, Feyza N AU - Hannezo, Edouard B AU - Merrin, Jack AU - Loose, Martin AU - Heisenberg, Carl-Philipp J ID - 14795 IS - 1 JF - Current Biology SN - 0960-9822 TI - Adhesion-induced cortical flows pattern E-cadherin-mediated cell contacts VL - 34 ER - TY - JOUR AB - The epitaxial growth of a strained Ge layer, which is a promising candidate for the channel material of a hole spin qubit, has been demonstrated on 300 mm Si wafers using commercially available Si0.3Ge0.7 strain relaxed buffer (SRB) layers. The assessment of the layer and the interface qualities for a buried strained Ge layer embedded in Si0.3Ge0.7 layers is reported. The XRD reciprocal space mapping confirmed that the reduction of the growth temperature enables the 2-dimensional growth of the Ge layer fully strained with respect to the Si0.3Ge0.7. Nevertheless, dislocations at the top and/or bottom interface of the Ge layer were observed by means of electron channeling contrast imaging, suggesting the importance of the careful dislocation assessment. The interface abruptness does not depend on the selection of the precursor gases, but it is strongly influenced by the growth temperature which affects the coverage of the surface H-passivation. The mobility of 2.7 × 105 cm2/Vs is promising, while the low percolation density of 3 × 1010 /cm2 measured with a Hall-bar device at 7 K illustrates the high quality of the heterostructure thanks to the high Si0.3Ge0.7 SRB quality. AU - Shimura, Yosuke AU - Godfrin, Clement AU - Hikavyy, Andriy AU - Li, Roy AU - Aguilera Servin, Juan L AU - Katsaros, Georgios AU - Favia, Paola AU - Han, Han AU - Wan, Danny AU - de Greve, Kristiaan AU - Loo, Roger ID - 15018 IS - 5 JF - Materials Science in Semiconductor Processing KW - Mechanical Engineering KW - Mechanics of Materials KW - Condensed Matter Physics KW - General Materials Science SN - 1369-8001 TI - Compressively strained epitaxial Ge layers for quantum computing applications VL - 174 ER - TY - JOUR AB - Embryogenesis results from the coordinated activities of different signaling pathways controlling cell fate specification and morphogenesis. In vertebrate gastrulation, both Nodal and BMP signaling play key roles in germ layer specification and morphogenesis, yet their interplay to coordinate embryo patterning with morphogenesis is still insufficiently understood. Here, we took a reductionist approach using zebrafish embryonic explants to study the coordination of Nodal and BMP signaling for embryo patterning and morphogenesis. We show that Nodal signaling triggers explant elongation by inducing mesendodermal progenitors but also suppressing BMP signaling activity at the site of mesendoderm induction. Consistent with this, ectopic BMP signaling in the mesendoderm blocks cell alignment and oriented mesendoderm intercalations, key processes during explant elongation. Translating these ex vivo observations to the intact embryo showed that, similar to explants, Nodal signaling suppresses the effect of BMP signaling on cell intercalations in the dorsal domain, thus allowing robust embryonic axis elongation. These findings suggest a dual function of Nodal signaling in embryonic axis elongation by both inducing mesendoderm and suppressing BMP effects in the dorsal portion of the mesendoderm. AU - Schauer, Alexandra AU - Pranjic-Ferscha, Kornelija AU - Hauschild, Robert AU - Heisenberg, Carl-Philipp J ID - 15048 IS - 4 JF - Development SN - 0950-1991 TI - Robust axis elongation by Nodal-dependent restriction of BMP signaling VL - 151 ER - TY - COMP AU - Hauschild, Robert ID - 14926 TI - Matlab script for analysis of clone dispersal ER - TY - JOUR AB - Poxviruses are among the largest double-stranded DNA viruses, with members such as variola virus, monkeypox virus and the vaccination strain vaccinia virus (VACV). Knowledge about the structural proteins that form the viral core has remained sparse. While major core proteins have been annotated via indirect experimental evidence, their structures have remained elusive and they could not be assigned to individual core features. Hence, which proteins constitute which layers of the core, such as the palisade layer and the inner core wall, has remained enigmatic. Here we show, using a multi-modal cryo-electron microscopy (cryo-EM) approach in combination with AlphaFold molecular modeling, that trimers formed by the cleavage product of VACV protein A10 are the key component of the palisade layer. This allows us to place previously obtained descriptions of protein interactions within the core wall into perspective and to provide a detailed model of poxvirus core architecture. Importantly, we show that interactions within A10 trimers are likely generalizable over members of orthopox- and parapoxviruses. AU - Datler, Julia AU - Hansen, Jesse AU - Thader, Andreas AU - Schlögl, Alois AU - Bauer, Lukas W AU - Hodirnau, Victor-Valentin AU - Schur, Florian KM ID - 14979 JF - Nature Structural & Molecular Biology KW - Molecular Biology KW - Structural Biology SN - 1545-9993 TI - Multi-modal cryo-EM reveals trimers of protein A10 to form the palisade layer in poxvirus cores ER - TY - JOUR AB - Contraction and flow of the actin cell cortex have emerged as a common principle by which cells reorganize their cytoplasm and take shape. However, how these cortical flows interact with adjacent cytoplasmic components, changing their form and localization, and how this affects cytoplasmic organization and cell shape remains unclear. Here we show that in ascidian oocytes, the cooperative activities of cortical actomyosin flows and deformation of the adjacent mitochondria-rich myoplasm drive oocyte cytoplasmic reorganization and shape changes following fertilization. We show that vegetal-directed cortical actomyosin flows, established upon oocyte fertilization, lead to both the accumulation of cortical actin at the vegetal pole of the zygote and compression and local buckling of the adjacent elastic solid-like myoplasm layer due to friction forces generated at their interface. Once cortical flows have ceased, the multiple myoplasm buckles resolve into one larger buckle, which again drives the formation of the contraction pole—a protuberance of the zygote’s vegetal pole where maternal mRNAs accumulate. Thus, our findings reveal a mechanism where cortical actomyosin network flows determine cytoplasmic reorganization and cell shape by deforming adjacent cytoplasmic components through friction forces. AU - Caballero Mancebo, Silvia AU - Shinde, Rushikesh AU - Bolger-Munro, Madison AU - Peruzzo, Matilda AU - Szep, Gregory AU - Steccari, Irene AU - Labrousse Arias, David AU - Zheden, Vanessa AU - Merrin, Jack AU - Callan-Jones, Andrew AU - Voituriez, Raphaël AU - Heisenberg, Carl-Philipp J ID - 14846 JF - Nature Physics SN - 1745-2473 TI - Friction forces determine cytoplasmic reorganization and shape changes of ascidian oocytes upon fertilization ER - TY - JOUR AB - The coupling between Ca2+ channels and release sensors is a key factor defining the signaling properties of a synapse. However, the coupling nanotopography at many synapses remains unknown, and it is unclear how it changes during development. To address these questions, we examined coupling at the cerebellar inhibitory basket cell (BC)-Purkinje cell (PC) synapse. Biophysical analysis of transmission by paired recording and intracellular pipette perfusion revealed that the effects of exogenous Ca2+ chelators decreased during development, despite constant reliance of release on P/Q-type Ca2+ channels. Structural analysis by freeze-fracture replica labeling (FRL) and transmission electron microscopy (EM) indicated that presynaptic P/Q-type Ca2+ channels formed nanoclusters throughout development, whereas docked vesicles were only clustered at later developmental stages. Modeling suggested a developmental transformation from a more random to a more clustered coupling nanotopography. Thus, presynaptic signaling developmentally approaches a point-to-point configuration, optimizing speed, reliability, and energy efficiency of synaptic transmission. AU - Chen, JingJing AU - Kaufmann, Walter AU - Chen, Chong AU - Arai, Itaru AU - Kim, Olena AU - Shigemoto, Ryuichi AU - Jonas, Peter M ID - 14843 JF - Neuron SN - 0896-6273 TI - Developmental transformation of Ca2+ channel-vesicle nanotopography at a central GABAergic synapse ER - TY - JOUR AB - Primary implant stability, which refers to the stability of the implant during the initial healing period is a crucial factor in determining the long-term success of the implant and lays the foundation for secondary implant stability achieved through osseointegration. Factors affecting primary stability include implant design, surgical technique, and patient-specific factors like bone quality and morphology. In vivo, the cyclic nature of anatomical loading puts osteosynthesis locking screws under dynamic loads, which can lead to the formation of micro cracks and defects that slowly degrade the mechanical connection between the bone and screw, thus compromising the initial stability and secondary stability of the implant. Monotonic quasi-static loading used for testing the holding capacity of implanted screws is not well suited to capture this behavior since it cannot capture the progressive deterioration of peri‑implant bone at small displacements. In order to address this issue, this study aims to determine a critical point of loss of primary implant stability in osteosynthesis locking screws under cyclic overloading by investigating the evolution of damage, dissipated energy, and permanent deformation. A custom-made test setup was used to test implanted 2.5 mm locking screws under cyclic overloading test. For each loading cycle, maximum forces and displacement were recorded as well as initial and final cycle displacements and used to calculate damage and energy dissipation evolution. The results of this study demonstrate that for axial, shear, and mixed loading significant damage and energy dissipation can be observed at approximately 20 % of the failure force. Additionally, at this load level, permanent deformations on the screw-bone interface were found to be in the range of 50 to 150 mm which promotes osseointegration and secondary implant stability. This research can assist surgeons in making informed preoperative decisions by providing a better understanding of the critical point of loss of primary implant stability, thus improving the long-term success of the implant and overall patient satisfaction. AU - Silva-Henao, Juan D. AU - Schober, Sophie AU - Pahr, Dieter H. AU - Reisinger, Andreas G. ID - 15164 JF - Medical Engineering and Physics SN - 1350-4533 TI - Critical loss of primary implant stability in osteosynthesis locking screws under cyclic overloading VL - 126 ER - TY - JOUR AB - Thermoelectric materials convert heat into electricity, with a broad range of applications near room temperature (RT). However, the library of RT high-performance materials is limited. Traditional high-temperature synthetic methods constrain the range of materials achievable, hindering the ability to surpass crystal structure limitations and engineer defects. Here, a solution-based synthetic approach is introduced, enabling RT synthesis of powders and exploration of densification at lower temperatures to influence the material's microstructure. The approach is exemplified by Ag2Se, an n-type alternative to bismuth telluride. It is demonstrated that the concentration of Ag interstitials, grain boundaries, and dislocations are directly correlated to the sintering temperature, and achieve a figure of merit of 1.1 from RT to 100 °C after optimization. Moreover, insights into and resolve Ag2Se's challenges are provided, including stoichiometry issues leading to irreproducible performances. This work highlights the potential of RT solution synthesis in expanding the repertoire of high-performance thermoelectric materials for practical applications. AU - Kleinhanns, Tobias AU - Milillo, Francesco AU - Calcabrini, Mariano AU - Fiedler, Christine AU - Horta, Sharona AU - Balazs, Daniel AU - Strumolo, Marissa J. AU - Hasler, Roger AU - Llorca, Jordi AU - Tkadletz, Michael AU - Brutchey, Richard L. AU - Ibáñez, Maria ID - 15182 JF - Advanced Energy Materials SN - 1614-6832 TI - A route to high thermoelectric performance: Solution‐based control of microstructure and composition in Ag2Se ER - TY - JOUR AB - 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. AU - Zens, Bettina AU - Fäßler, Florian AU - Hansen, Jesse AU - Hauschild, Robert AU - Datler, Julia AU - Hodirnau, Victor-Valentin AU - Zheden, Vanessa AU - Alanko, Jonna H AU - Sixt, Michael K AU - Schur, Florian KM ID - 15146 IS - 6 JF - Journal of Cell Biology SN - 0021-9525 TI - Lift-out cryo-FIBSEM and cryo-ET reveal the ultrastructural landscape of extracellular matrix VL - 223 ER - TY - JOUR AB - Post-translational histone modifications modulate chromatin activity to affect gene expression. How chromatin states underlie lineage choice in single cells is relatively unexplored. We develop sort-assisted single-cell chromatin immunocleavage (sortChIC) and map active (H3K4me1 and H3K4me3) and repressive (H3K27me3 and H3K9me3) histone modifications in the mouse bone marrow. During differentiation, hematopoietic stem and progenitor cells (HSPCs) acquire active chromatin states mediated by cell-type-specifying transcription factors, which are unique for each lineage. By contrast, most alterations in repressive marks during differentiation occur independent of the final cell type. Chromatin trajectory analysis shows that lineage choice at the chromatin level occurs at the progenitor stage. Joint profiling of H3K4me1 and H3K9me3 demonstrates that cell types within the myeloid lineage have distinct active chromatin but share similar myeloid-specific heterochromatin states. This implies a hierarchical regulation of chromatin during hematopoiesis: heterochromatin dynamics distinguish differentiation trajectories and lineages, while euchromatin dynamics reflect cell types within lineages. AU - Zeller, Peter AU - Yeung, Jake AU - Viñas Gaza, Helena AU - de Barbanson, Buys Anton AU - Bhardwaj, Vivek AU - Florescu, Maria AU - van der Linden, Reinier AU - van Oudenaarden, Alexander ID - 12158 JF - Nature Genetics KW - Genetics SN - 1061-4036 TI - Single-cell sortChIC identifies hierarchical chromatin dynamics during hematopoiesis VL - 55 ER - TY - GEN AU - Elefante, Stefano AU - Stadlbauer, Stephan AU - Alexander, Michael F AU - Schlögl, Alois ID - 13162 T2 - ASHPC23 - Austrian-Slovenian HPC Meeting 2023 TI - Cryo-EM software packages: A sys-admins point of view ER - TY - GEN AU - Schlögl, Alois AU - Elefante, Stefano AU - Hodirnau, Victor-Valentin ID - 13161 T2 - ASHPC23 - Austrian-Slovenian HPC Meeting 2023 TI - Running Windows-applications on a Linux HPC cluster using WINE ER - TY - JOUR AB - Interstitial fluid (IF) accumulation between embryonic cells is thought to be important for embryo patterning and morphogenesis. Here, we identify a positive mechanical feedback loop between cell migration and IF relocalization and find that it promotes embryonic axis formation during zebrafish gastrulation. We show that anterior axial mesendoderm (prechordal plate [ppl]) cells, moving in between the yolk cell and deep cell tissue to extend the embryonic axis, compress the overlying deep cell layer, thereby causing IF to flow from the deep cell layer to the boundary between the yolk cell and the deep cell layer, directly ahead of the advancing ppl. This IF relocalization, in turn, facilitates ppl cell protrusion formation and migration by opening up the space into which the ppl moves and, thereby, the ability of the ppl to trigger IF relocalization by pushing against the overlying deep cell layer. Thus, embryonic axis formation relies on a hydraulic feedback loop between cell migration and IF relocalization. AU - Huljev, Karla AU - Shamipour, Shayan AU - Nunes Pinheiro, Diana C AU - Preusser, Friedrich AU - Steccari, Irene AU - Sommer, Christoph M AU - Naik, Suyash AU - Heisenberg, Carl-Philipp J ID - 12830 IS - 7 JF - Developmental Cell SN - 1534-5807 TI - A hydraulic feedback loop between mesendoderm cell migration and interstitial fluid relocalization promotes embryonic axis formation in zebrafish VL - 58 ER - TY - JOUR AB - Current methods for assessing cell proliferation in 3D scaffolds rely on changes in metabolic activity or total DNA, however, direct quantification of cell number in 3D scaffolds remains a challenge. To address this issue, we developed an unbiased stereology approach that uses systematic-random sampling and thin focal-plane optical sectioning of the scaffolds followed by estimation of total cell number (StereoCount). This approach was validated against an indirect method for measuring the total DNA (DNA content); and the Bürker counting chamber, the current reference method for quantifying cell number. We assessed the total cell number for cell seeding density (cells per unit volume) across four values and compared the methods in terms of accuracy, ease-of-use and time demands. The accuracy of StereoCount markedly outperformed the DNA content for cases with ~ 10,000 and ~ 125,000 cells/scaffold. For cases with ~ 250,000 and ~ 375,000 cells/scaffold both StereoCount and DNA content showed lower accuracy than the Bürker but did not differ from each other. In terms of ease-of-use, there was a strong advantage for the StereoCount due to output in terms of absolute cell numbers along with the possibility for an overview of cell distribution and future use of automation for high throughput analysis. Taking together, the StereoCount method is an efficient approach for direct cell quantification in 3D collagen scaffolds. Its major benefit is that automated StereoCount could accelerate research using 3D scaffolds focused on drug discovery for a wide variety of human diseases. AU - Zavadakova, Anna AU - Vistejnova, Lucie AU - Belinova, Tereza AU - Tichanek, Filip AU - Bilikova, Dagmar AU - Mouton, Peter R. ID - 13033 IS - 1 JF - Scientific Reports KW - Multidisciplinary SN - 2045-2322 TI - Novel stereological method for estimation of cell counts in 3D collagen scaffolds VL - 13 ER - TY - JOUR AB - Regulation of chromatin states involves the dynamic interplay between different histone modifications to control gene expression. Recent advances have enabled mapping of histone marks in single cells, but most methods are constrained to profile only one histone mark per cell. Here, we present an integrated experimental and computational framework, scChIX-seq (single-cell chromatin immunocleavage and unmixing sequencing), to map several histone marks in single cells. scChIX-seq multiplexes two histone marks together in single cells, then computationally deconvolves the signal using training data from respective histone mark profiles. This framework learns the cell-type-specific correlation structure between histone marks, and therefore does not require a priori assumptions of their genomic distributions. Using scChIX-seq, we demonstrate multimodal analysis of histone marks in single cells across a range of mark combinations. Modeling dynamics of in vitro macrophage differentiation enables integrated analysis of chromatin velocity. Overall, scChIX-seq unlocks systematic interrogation of the interplay between histone modifications in single cells. AU - Yeung, Jake AU - Florescu, Maria AU - Zeller, Peter AU - De Barbanson, Buys Anton AU - Wellenstein, Max D. AU - Van Oudenaarden, Alexander ID - 12106 JF - Nature Biotechnology SN - 1087-0156 TI - scChIX-seq infers dynamic relationships between histone modifications in single cells VL - 41 ER - TY - JOUR AB - Treating sick group members is a hallmark of collective disease defence in vertebrates and invertebrates alike. Despite substantial effects on pathogen fitness and epidemiology, it is still largely unknown how pathogens react to the selection pressure imposed by care intervention. Using social insects and pathogenic fungi, we here performed a serial passage experiment in the presence or absence of colony members, which provide social immunity by grooming off infectious spores from exposed individuals. We found specific effects on pathogen diversity, virulence and transmission. Under selection of social immunity, pathogens invested into higher spore production, but spores were less virulent. Notably, they also elicited a lower grooming response in colony members, compared with spores from the individual host selection lines. Chemical spore analysis suggested that the spores from social selection lines escaped the caregivers’ detection by containing lower levels of ergosterol, a key fungal membrane component. Experimental application of chemically pure ergosterol indeed induced sanitary grooming, supporting its role as a microbe-associated cue triggering host social immunity against fungal pathogens. By reducing this detection cue, pathogens were able to evade the otherwise very effective collective disease defences of their social hosts. AU - Stock, Miriam AU - Milutinovic, Barbara AU - Hönigsberger, Michaela AU - Grasse, Anna V AU - Wiesenhofer, Florian AU - Kampleitner, Niklas AU - Narasimhan, Madhumitha AU - Schmitt, Thomas AU - Cremer, Sylvia ID - 12543 JF - Nature Ecology and Evolution TI - Pathogen evasion of social immunity VL - 7 ER - TY - JOUR AB - In the present study, essential and nonessential metal content and biomarker responses were investigated in the intestine of fish collected from the areas polluted by mining. Our objective was to determine metal and biomarker levels in tissue responsible for dietary intake, which is rarely studied in water pollution research. The study was conducted in the Bregalnica River, reference location, and in the Zletovska and Kriva Rivers (the Republic of North Macedonia), which are directly influenced by the active mines Zletovo and Toranica, respectively. Biological responses were analyzed in Vardar chub (Squalius vardarensis; Karaman, 1928), using for the first time intestinal cytosol as a potentially toxic cell fraction, since metal sensitivity is mostly associated with cytosol. Cytosolic metal levels were higher in fish under the influence of mining (Tl, Li, Cs, Mo, Sr, Cd, Rb, and Cu in the Zletovska River and Cr, Pb, and Se in the Kriva River compared to the Bregalnica River in both seasons). The same trend was evident for total proteins, biomarkers of general stress, and metallothioneins, biomarkers of metal exposure, indicating cellular disturbances in the intestine, the primary site of dietary metal uptake. The association of cytosolic Cu and Cd at all locations pointed to similar pathways and homeostasis of these metallothionein-binding metals. Comparison with other indicator tissues showed that metal concentrations were higher in the intestine of fish from mining-affected areas than in the liver and gills. In general, these results indicated the importance of dietary metal pathways, and cytosolic metal fraction in assessing pollution impacts in freshwater ecosystems. AU - Filipović Marijić, Vlatka AU - Krasnici, Nesrete AU - Valić, Damir AU - Kapetanović, Damir AU - Vardić Smrzlić, Irena AU - Jordanova, Maja AU - Rebok, Katerina AU - Ramani, Sheriban AU - Kostov, Vasil AU - Nastova, Rodne AU - Dragun, Zrinka ID - 12863 JF - Environmental Science and Pollution Research SN - 0944-1344 TI - Pollution impact on metal and biomarker responses in intestinal cytosol of freshwater fish VL - 30 ER - TY - JOUR AB - A light-triggered fabrication method extends the functionality of printable nanomaterials AU - Balazs, Daniel AU - Ibáñez, Maria ID - 14404 IS - 6665 JF - Science TI - Widening the use of 3D printing VL - 381 ER - TY - CHAP AB - Imaging of the immunological synapse (IS) between dendritic cells (DCs) and T cells in suspension is hampered by suboptimal alignment of cell-cell contacts along the vertical imaging plane. This requires optical sectioning that often results in unsatisfactory resolution in time and space. Here, we present a workflow where DCs and T cells are confined between a layer of glass and polydimethylsiloxane (PDMS) that orients the cells along one, horizontal imaging plane, allowing for fast en-face-imaging of the DC-T cell IS. AU - Leithner, Alexander F AU - Merrin, Jack AU - Sixt, Michael K ED - Baldari, Cosima ED - Dustin, Michael ID - 13052 SN - 1064-3745 T2 - The Immune Synapse TI - En-Face Imaging of T Cell-Dendritic Cell Immunological Synapses VL - 2654 ER - TY - JOUR AB - Regulation of the Arp2/3 complex is required for productive nucleation of branched actin networks. An emerging aspect of regulation is the incorporation of subunit isoforms into the Arp2/3 complex. Specifically, both ArpC5 subunit isoforms, ArpC5 and ArpC5L, have been reported to fine-tune nucleation activity and branch junction stability. We have combined reverse genetics and cellular structural biology to describe how ArpC5 and ArpC5L differentially affect cell migration. Both define the structural stability of ArpC1 in branch junctions and, in turn, by determining protrusion characteristics, affect protein dynamics and actin network ultrastructure. ArpC5 isoforms also affect the positioning of members of the Ena/Vasodilator-stimulated phosphoprotein (VASP) family of actin filament elongators, which mediate ArpC5 isoform–specific effects on the actin assembly level. Our results suggest that ArpC5 and Ena/VASP proteins are part of a signaling pathway enhancing cell migration. AU - Fäßler, Florian AU - Javoor, Manjunath AU - Datler, Julia AU - Döring, Hermann AU - Hofer, Florian AU - Dimchev, Georgi A AU - Hodirnau, Victor-Valentin AU - Faix, Jan AU - Rottner, Klemens AU - Schur, Florian KM ID - 12334 IS - 3 JF - Science Advances KW - Multidisciplinary SN - 2375-2548 TI - ArpC5 isoforms regulate Arp2/3 complex–dependent protrusion through differential Ena/VASP positioning VL - 9 ER - TY - JOUR AB - Motile cells moving in multicellular organisms encounter microenvironments of locally heterogeneous mechanochemical composition. Individual compositional parameters like chemotactic signals, adhesiveness, and pore sizes are well known to be sensed by motile cells, providing individual guidance cues for cellular pathfinding. However, motile cells encounter diverse mechanochemical signals at the same time, raising the question of how cells respond to locally diverse and potentially competing signals on their migration routes. Here, we reveal that motile amoeboid cells require nuclear repositioning, termed nucleokinesis, for adaptive pathfinding in heterogeneous mechanochemical microenvironments. Using mammalian immune cells and the amoebaDictyostelium discoideum, we discover that frequent, rapid and long-distance nucleokinesis is a basic component of amoeboid pathfinding, enabling cells to reorientate quickly between locally competing cues. Amoeboid nucleokinesis comprises a two-step cell polarity switch and is driven by myosin II-forces, sliding the nucleus from a ‘losing’ to the ‘winning’ leading edge to re-adjust the nuclear to the cellular path. Impaired nucleokinesis distorts fast path adaptions and causes cellular arrest in the microenvironment. Our findings establish that nucleokinesis is required for amoeboid cell navigation. Given that motile single-cell amoebae, many immune cells, and some cancer cells utilize an amoeboid migration strategy, these results suggest that amoeboid nucleokinesis underlies cellular navigation during unicellular biology, immunity, and disease. AU - Kroll, Janina AU - Hauschild, Robert AU - Kuznetcov, Arthur AU - Stefanowski, Kasia AU - Hermann, Monika D. AU - Merrin, Jack AU - Shafeek, Lubuna B AU - Müller-Taubenberger, Annette AU - Renkawitz, Jörg ID - 13342 JF - EMBO Journal SN - 0261-4189 TI - Adaptive pathfinding by nucleokinesis during amoeboid migration ER - TY - JOUR AB - Muscle degeneration is the most prevalent cause for frailty and dependency in inherited diseases and ageing. Elucidation of pathophysiological mechanisms, as well as effective treatments for muscle diseases, represents an important goal in improving human health. Here, we show that the lipid synthesis enzyme phosphatidylethanolamine cytidyltransferase (PCYT2/ECT) is critical to muscle health. Human deficiency in PCYT2 causes a severe disease with failure to thrive and progressive weakness. pcyt2-mutant zebrafish and muscle-specific Pcyt2-knockout mice recapitulate the participant phenotypes, with failure to thrive, progressive muscle weakness and accelerated ageing. Mechanistically, muscle Pcyt2 deficiency affects cellular bioenergetics and membrane lipid bilayer structure and stability. PCYT2 activity declines in ageing muscles of mice and humans, and adeno-associated virus-based delivery of PCYT2 ameliorates muscle weakness in Pcyt2-knockout and old mice, offering a therapy for individuals with a rare disease and muscle ageing. Thus, PCYT2 plays a fundamental and conserved role in vertebrate muscle health, linking PCYT2 and PCYT2-synthesized lipids to severe muscle dystrophy and ageing. AU - Cikes, Domagoj AU - Elsayad, Kareem AU - Sezgin, Erdinc AU - Koitai, Erika AU - Ferenc, Torma AU - Orthofer, Michael AU - Yarwood, Rebecca AU - Heinz, Leonhard X. AU - Sedlyarov, Vitaly AU - Darwish-Miranda, Nasser AU - Taylor, Adrian AU - Grapentine, Sophie AU - al-Murshedi, Fathiya AU - Abot, Anne AU - Weidinger, Adelheid AU - Kutchukian, Candice AU - Sanchez, Colline AU - Cronin, Shane J. F. AU - Novatchkova, Maria AU - Kavirayani, Anoop AU - Schuetz, Thomas AU - Haubner, Bernhard AU - Haas, Lisa AU - Hagelkruys, Astrid AU - Jackowski, Suzanne AU - Kozlov, Andrey AU - Jacquemond, Vincent AU - Knauf, Claude AU - Superti-Furga, Giulio AU - Rullman, Eric AU - Gustafsson, Thomas AU - McDermot, John AU - Lowe, Martin AU - Radak, Zsolt AU - Chamberlain, Jeffrey S. AU - Bakovic, Marica AU - Banka, Siddharth AU - Penninger, Josef M. ID - 12747 JF - Nature Metabolism KW - Cell Biology KW - Physiology (medical) KW - Endocrinology KW - Diabetes and Metabolism KW - Internal Medicine SN - 2522-5812 TI - PCYT2-regulated lipid biosynthesis is critical to muscle health and ageing VL - 5 ER - TY - JOUR AB - Tissue morphogenesis and patterning during development involve the segregation of cell types. Segregation is driven by differential tissue surface tensions generated by cell types through controlling cell-cell contact formation by regulating adhesion and actomyosin contractility-based cellular cortical tensions. We use vertebrate tissue cell types and zebrafish germ layer progenitors as in vitro models of 3-dimensional heterotypic segregation and developed a quantitative analysis of their dynamics based on 3D time-lapse microscopy. We show that general inhibition of actomyosin contractility by the Rho kinase inhibitor Y27632 delays segregation. Cell type-specific inhibition of non-muscle myosin2 activity by overexpression of myosin assembly inhibitor S100A4 reduces tissue surface tension, manifested in decreased compaction during aggregation and inverted geometry observed during segregation. The same is observed when we express a constitutively active Rho kinase isoform to ubiquitously keep actomyosin contractility high at cell-cell and cell-medium interfaces and thus overriding the interface-specific regulation of cortical tensions. Tissue surface tension regulation can become an effective tool in tissue engineering. AU - Méhes, Elod AU - Mones, Enys AU - Varga, Máté AU - Zsigmond, Áron AU - Biri-Kovács, Beáta AU - Nyitray, László AU - Barone, Vanessa AU - Krens, Gabriel AU - Heisenberg, Carl-Philipp J AU - Vicsek, Tamás ID - 14041 JF - Communications Biology TI - 3D cell segregation geometry and dynamics are governed by tissue surface tension regulation VL - 6 ER - TY - JOUR AB - Whether one considers swarming insects, flocking birds, or bacterial colonies, collective motion arises from the coordination of individuals and entails the adjustment of their respective velocities. In particular, in close confinements, such as those encountered by dense cell populations during development or regeneration, collective migration can only arise coordinately. Yet, how individuals unify their velocities is often not understood. Focusing on a finite number of cells in circular confinements, we identify waves of polymerizing actin that function as a pacemaker governing the speed of individual cells. We show that the onset of collective motion coincides with the synchronization of the wave nucleation frequencies across the population. Employing a simpler and more readily accessible mechanical model system of active spheres, we identify the synchronization of the individuals’ internal oscillators as one of the essential requirements to reach the corresponding collective state. The mechanical ‘toy’ experiment illustrates that the global synchronous state is achieved by nearest neighbor coupling. We suggest by analogy that local coupling and the synchronization of actin waves are essential for the emergent, self-organized motion of cell collectives. AU - Riedl, Michael AU - Mayer, Isabelle D AU - Merrin, Jack AU - Sixt, Michael K AU - Hof, Björn ID - 14361 JF - Nature Communications TI - Synchronization in collectively moving inanimate and living active matter VL - 14 ER - TY - JOUR AB - The rapid development of machine learning (ML) techniques has opened up the data-dense field of microbiome research for novel therapeutic, diagnostic, and prognostic applications targeting a wide range of disorders, which could substantially improve healthcare practices in the era of precision medicine. However, several challenges must be addressed to exploit the benefits of ML in this field fully. In particular, there is a need to establish “gold standard” protocols for conducting ML analysis experiments and improve interactions between microbiome researchers and ML experts. The Machine Learning Techniques in Human Microbiome Studies (ML4Microbiome) COST Action CA18131 is a European network established in 2019 to promote collaboration between discovery-oriented microbiome researchers and data-driven ML experts to optimize and standardize ML approaches for microbiome analysis. This perspective paper presents the key achievements of ML4Microbiome, which include identifying predictive and discriminatory ‘omics’ features, improving repeatability and comparability, developing automation procedures, and defining priority areas for the novel development of ML methods targeting the microbiome. The insights gained from ML4Microbiome will help to maximize the potential of ML in microbiome research and pave the way for new and improved healthcare practices. AU - D’Elia, Domenica AU - Truu, Jaak AU - Lahti, Leo AU - Berland, Magali AU - Papoutsoglou, Georgios AU - Ceci, Michelangelo AU - Zomer, Aldert AU - Lopes, Marta B. AU - Ibrahimi, Eliana AU - Gruca, Aleksandra AU - Nechyporenko, Alina AU - Frohme, Marcus AU - Klammsteiner, Thomas AU - Pau, Enrique Carrillo De Santa AU - Marcos-Zambrano, Laura Judith AU - Hron, Karel AU - Pio, Gianvito AU - Simeon, Andrea AU - Suharoschi, Ramona AU - Moreno-Indias, Isabel AU - Temko, Andriy AU - Nedyalkova, Miroslava AU - Apostol, Elena Simona AU - Truică, Ciprian Octavian AU - Shigdel, Rajesh AU - Telalović, Jasminka Hasić AU - Bongcam-Rudloff, Erik AU - Przymus, Piotr AU - Jordamović, Naida Babić AU - Falquet, Laurent AU - Tarazona, Sonia AU - Sampri, Alexia AU - Isola, Gaetano AU - Pérez-Serrano, David AU - Trajkovik, Vladimir AU - Klucar, Lubos AU - Loncar-Turukalo, Tatjana AU - Havulinna, Aki S. AU - Jansen, Christian AU - Bertelsen, Randi J. AU - Claesson, Marcus Joakim ID - 14449 JF - Frontiers in Microbiology TI - Advancing microbiome research with machine learning: Key findings from the ML4Microbiome COST action VL - 14 ER - TY - JOUR AB - Immune responses rely on the rapid and coordinated migration of leukocytes. Whereas it is well established that single-cell migration is often guided by gradients of chemokines and other chemoattractants, it remains poorly understood how these gradients are generated, maintained, and modulated. By combining experimental data with theory on leukocyte chemotaxis guided by the G protein–coupled receptor (GPCR) CCR7, we demonstrate that in addition to its role as the sensory receptor that steers migration, CCR7 also acts as a generator and a modulator of chemotactic gradients. Upon exposure to the CCR7 ligand CCL19, dendritic cells (DCs) effectively internalize the receptor and ligand as part of the canonical GPCR desensitization response. We show that CCR7 internalization also acts as an effective sink for the chemoattractant, dynamically shaping the spatiotemporal distribution of the chemokine. This mechanism drives complex collective migration patterns, enabling DCs to create or sharpen chemotactic gradients. We further show that these self-generated gradients can sustain the long-range guidance of DCs, adapt collective migration patterns to the size and geometry of the environment, and provide a guidance cue for other comigrating cells. Such a dual role of CCR7 as a GPCR that both senses and consumes its ligand can thus provide a novel mode of cellular self-organization. AU - Alanko, Jonna H AU - Ucar, Mehmet C AU - Canigova, Nikola AU - Stopp, Julian A AU - Schwarz, Jan AU - Merrin, Jack AU - Hannezo, Edouard B AU - Sixt, Michael K ID - 14274 IS - 87 JF - Science Immunology KW - General Medicine KW - Immunology SN - 2470-9468 TI - CCR7 acts as both a sensor and a sink for CCL19 to coordinate collective leukocyte migration VL - 8 ER - TY - JOUR AB - Three-dimensional (3D) reconstruction of living brain tissue down to an individual synapse level would create opportunities for decoding the dynamics and structure–function relationships of the brain’s complex and dense information processing network; however, this has been hindered by insufficient 3D resolution, inadequate signal-to-noise ratio and prohibitive light burden in optical imaging, whereas electron microscopy is inherently static. Here we solved these challenges by developing an integrated optical/machine-learning technology, LIONESS (live information-optimized nanoscopy enabling saturated segmentation). This leverages optical modifications to stimulated emission depletion microscopy in comprehensively, extracellularly labeled tissue and previous information on sample structure via machine learning to simultaneously achieve isotropic super-resolution, high signal-to-noise ratio and compatibility with living tissue. This allows dense deep-learning-based instance segmentation and 3D reconstruction at a synapse level, incorporating molecular, activity and morphodynamic information. LIONESS opens up avenues for studying the dynamic functional (nano-)architecture of living brain tissue. AU - Velicky, Philipp AU - Miguel Villalba, Eder AU - Michalska, Julia M AU - Lyudchik, Julia AU - Wei, Donglai AU - Lin, Zudi AU - Watson, Jake AU - Troidl, Jakob AU - Beyer, Johanna AU - Ben Simon, Yoav AU - Sommer, Christoph M AU - Jahr, Wiebke AU - Cenameri, Alban AU - Broichhagen, Johannes AU - Grant, Seth G.N. AU - Jonas, Peter M AU - Novarino, Gaia AU - Pfister, Hanspeter AU - Bickel, Bernd AU - Danzl, Johann G ID - 13267 JF - Nature Methods SN - 1548-7091 TI - Dense 4D nanoscale reconstruction of living brain tissue VL - 20 ER - TY - JOUR AB - Germ granules, condensates of phase-separated RNA and protein, are organelles that are essential for germline development in different organisms. The patterning of the granules and their relevance for germ cell fate are not fully understood. Combining three-dimensional in vivo structural and functional analyses, we study the dynamic spatial organization of molecules within zebrafish germ granules. We find that the localization of RNA molecules to the periphery of the granules, where ribosomes are localized, depends on translational activity at this location. In addition, we find that the vertebrate-specific Dead end (Dnd1) protein is essential for nanos3 RNA localization at the condensates’ periphery. Accordingly, in the absence of Dnd1, or when translation is inhibited, nanos3 RNA translocates into the granule interior, away from the ribosomes, a process that is correlated with the loss of germ cell fate. These findings highlight the relevance of sub-granule compartmentalization for post-transcriptional control and its importance for preserving germ cell totipotency. AU - Westerich, Kim Joana AU - Tarbashevich, Katsiaryna AU - Schick, Jan AU - Gupta, Antra AU - Zhu, Mingzhao AU - Hull, Kenneth AU - Romo, Daniel AU - Zeuschner, Dagmar AU - Goudarzi, Mohammad AU - Gross-Thebing, Theresa AU - Raz, Erez ID - 14781 IS - 17 JF - Developmental Cell KW - Developmental Biology KW - Cell Biology KW - General Biochemistry KW - Genetics and Molecular Biology KW - Molecular Biology SN - 1534-5807 TI - Spatial organization and function of RNA molecules within phase-separated condensates in zebrafish are controlled by Dnd1 VL - 58 ER - TY - JOUR AB - Acanthocephalans, intestinal parasites of vertebrates, are characterised by orders of magnitude higher metal accumulation than free-living organisms, but the mechanism of such effective metal accumulation is still unknown. The aim of our study was to gain new insights into the high-resolution localization of elements in the bodies of acanthocephalans, thus taking an initial step towards elucidating metal uptake and accumulation in organisms under real environmental conditions. For the first time, nanoscale secondary ion mass spectrometry (NanoSIMS) was used for high-resolution mapping of 12 elements (C, Ca, Cu, Fe, N, Na, O, P, Pb, S, Se, and Tl) in three selected body parts (trunk spines, inner part of the proboscis receptacle and inner surface of the tegument) of Dentitruncus truttae, a parasite of brown trout (Salmo trutta) from the Krka River in Croatia. In addition, the same body parts were examined using transmission electron microscopy (TEM) and correlated with NanoSIMS images. Metal concentrations determined using HR ICP-MS confirmed higher accumulation in D. truttae than in the fish intestine. The chemical composition of the acanthocephalan body showed the highest density of C, Ca, N, Na, O, S, as important and constitutive elements in living cells in all studied structures, while Fe was predominant among trace elements. In general, higher element density was found in trunk spines and tegument, as body structures responsible for substance absorption in parasites. The results obtained with NanoSIMS and TEM-NanoSIMS correlative imaging represent pilot data for mapping of elements at nanoscale resolution in the ultrastructure of various body parts of acanthocephalans and generally provide a contribution for further application of this technique in all parasite species. AU - Filipović Marijić, Vlatka AU - Subirana, Maria Angels AU - Schaumlöffel, Dirk AU - Barišić, Josip AU - Gontier, Etienne AU - Krasnici, Nesrete AU - Mijošek, Tatjana AU - Hernández-Orts, Jesús S. AU - Scholz, Tomáš AU - Erk, Marijana ID - 14786 JF - Science of The Total Environment KW - Pollution KW - Waste Management and Disposal KW - Environmental Chemistry KW - Environmental Engineering SN - 0048-9697 TI - First insight in element localisation in different body parts of the acanthocephalan Dentitruncus truttae using TEM and NanoSIMS VL - 887 ER - TY - JOUR AB - A round-robin study has been carried out to estimate the impact of the human element in small-angle scattering data analysis. Four corrected datasets were provided to participants ready for analysis. All datasets were measured on samples containing spherical scatterers, with two datasets in dilute dispersions and two from powders. Most of the 46 participants correctly identified the number of populations in the dilute dispersions, with half of the population mean entries within 1.5% and half of the population width entries within 40%. Due to the added complexity of the structure factor, far fewer people submitted answers on the powder datasets. For those that did, half of the entries for the means and widths were within 44 and 86%, respectively. This round-robin experiment highlights several causes for the discrepancies, for which solutions are proposed. AU - Pauw, Brian R. AU - Smales, Glen J. AU - Anker, Andy S. AU - Annadurai, Venkatasamy AU - Balazs, Daniel AU - Bienert, Ralf AU - Bouwman, Wim G. AU - Breßler, Ingo AU - Breternitz, Joachim AU - Brok, Erik S. AU - Bryant, Gary AU - Clulow, Andrew J. AU - Crater, Erin R. AU - De Geuser, Frédéric AU - Giudice, Alessandra Del AU - Deumer, Jérôme AU - Disch, Sabrina AU - Dutt, Shankar AU - Frank, Kilian AU - Fratini, Emiliano AU - Garcia, Paulo R.A.F. AU - Gilbert, Elliot P. AU - Hahn, Marc B. AU - Hallett, James AU - Hohenschutz, Max AU - Hollamby, Martin AU - Huband, Steven AU - Ilavsky, Jan AU - Jochum, Johanna K. AU - Juelsholt, Mikkel AU - Mansel, Bradley W. AU - Penttilä, Paavo AU - Pittkowski, Rebecca K. AU - Portale, Giuseppe AU - Pozzo, Lilo D. AU - Rochels, Leonhard AU - Rosalie, Julian M. AU - Saloga, Patrick E.J. AU - Seibt, Susanne AU - Smith, Andrew J. AU - Smith, Gregory N. AU - Spiering, Glenn A. AU - Stawski, Tomasz M. AU - Taché, Olivier AU - Thünemann, Andreas F. AU - Toth, Kristof AU - Whitten, Andrew E. AU - Wuttke, Joachim ID - 14799 IS - 6 JF - Journal of Applied Crystallography SN - 0021-8898 TI - The human factor: Results of a small-angle scattering data analysis round robin VL - 56 ER - TY - GEN AB - Superconductor/semiconductor hybrid devices have attracted increasing interest in the past years. Superconducting electronics aims to complement semiconductor technology, while hybrid architectures are at the forefront of new ideas such as topological superconductivity and protected qubits. In this work, we engineer the induced superconductivity in two-dimensional germanium hole gas by varying the distance between the quantum well and the aluminum. We demonstrate a hard superconducting gap and realize an electrically and flux tunable superconducting diode using a superconducting quantum interference device (SQUID). This allows to tune the current phase relation (CPR), to a regime where single Cooper pair tunneling is suppressed, creating a $ \sin \left( 2 \varphi \right)$ CPR. Shapiro experiments complement this interpretation and the microwave drive allows to create a diode with $ \approx 100 \%$ efficiency. The reported results open up the path towards monolithic integration of spin qubit devices, microwave resonators and (protected) superconducting qubits on a silicon technology compatible platform. AU - Valentini, Marco AU - Sagi, Oliver AU - Baghumyan, Levon AU - Gijsel, Thijs de AU - Jung, Jason AU - Calcaterra, Stefano AU - Ballabio, Andrea AU - Servin, Juan Aguilera AU - Aggarwal, Kushagra AU - Janik, Marian AU - Adletzberger, Thomas AU - Souto, Rubén Seoane AU - Leijnse, Martin AU - Danon, Jeroen AU - Schrade, Constantin AU - Bakkers, Erik AU - Chrastina, Daniel AU - Isella, Giovanni AU - Katsaros, Georgios ID - 13312 KW - Mesoscale and Nanoscale Physics T2 - arXiv TI - Radio frequency driven superconducting diode and parity conserving Cooper pair transport in a two-dimensional germanium hole gas ER - TY - DATA AB - Mapping the complex and dense arrangement of cells and their connectivity in brain tissue demands nanoscale spatial resolution imaging. Super-resolution optical microscopy excels at visualizing specific molecules and individual cells but fails to provide tissue context. Here, we developed Comprehensive Analysis of Tissues across Scales (CATS), a technology to densely map brain tissue architecture from millimeter regional to nanometer synaptic scales in diverse chemically fixed brain preparations, including rodent and human. CATS uses fixation-compatible extracellular labeling and optical imaging, including stimulated emission depletion or expansion microscopy, to comprehensively delineate cellular structures. It enables three-dimensional reconstruction of single synapses and mapping of synaptic connectivity by identification and analysis of putative synaptic cleft regions. Applying CATS to the mouse hippocampal mossy fiber circuitry, we reconstructed and quantified the synaptic input and output structure of identified neurons. We furthermore demonstrate applicability to clinically derived human tissue samples, including formalin-fixed paraffin-embedded routine diagnostic specimens, for visualizing the cellular architecture of brain tissue in health and disease. AU - Danzl, Johann G ID - 13126 TI - Research data for the publication "Imaging brain tissue architecture across millimeter to nanometer scales" ER - TY - JOUR AB - Mapping the complex and dense arrangement of cells and their connectivity in brain tissue demands nanoscale spatial resolution imaging. Super-resolution optical microscopy excels at visualizing specific molecules and individual cells but fails to provide tissue context. Here we developed Comprehensive Analysis of Tissues across Scales (CATS), a technology to densely map brain tissue architecture from millimeter regional to nanometer synaptic scales in diverse chemically fixed brain preparations, including rodent and human. CATS uses fixation-compatible extracellular labeling and optical imaging, including stimulated emission depletion or expansion microscopy, to comprehensively delineate cellular structures. It enables three-dimensional reconstruction of single synapses and mapping of synaptic connectivity by identification and analysis of putative synaptic cleft regions. Applying CATS to the mouse hippocampal mossy fiber circuitry, we reconstructed and quantified the synaptic input and output structure of identified neurons. We furthermore demonstrate applicability to clinically derived human tissue samples, including formalin-fixed paraffin-embedded routine diagnostic specimens, for visualizing the cellular architecture of brain tissue in health and disease. AU - Michalska, Julia M AU - Lyudchik, Julia AU - Velicky, Philipp AU - Korinkova, Hana AU - Watson, Jake AU - Cenameri, Alban AU - Sommer, Christoph M AU - Amberg, Nicole AU - Venturino, Alessandro AU - Roessler, Karl AU - Czech, Thomas AU - Höftberger, Romana AU - Siegert, Sandra AU - Novarino, Gaia AU - Jonas, Peter M AU - Danzl, Johann G ID - 14257 JF - Nature Biotechnology SN - 1087-0156 TI - Imaging brain tissue architecture across millimeter to nanometer scales ER - TY - JOUR AB - Singlet oxygen (1O2) formation is now recognised as a key aspect of non-aqueous oxygen redox chemistry. For identifying 1O2, chemical trapping via 9,10-dimethylanthracene (DMA) to form the endoperoxide (DMA-O2) has become the mainstay method due to its sensitivity, selectivity, and ease of use. While DMA has been shown to be selective for 1O2, rather than forming DMA-O2 with a wide variety of potentially reactive O-containing species, false positives might hypothetically be obtained in the presence of previously overlooked species. Here, we first give unequivocal direct spectroscopic proof by the 1O2-specific near infrared (NIR) emission at 1270 nm for the previously proposed 1O2 formation pathways, which centre around superoxide disproportionation. We then show that peroxocarbonates, common intermediates in metal-O2 and metal carbonate electrochemistry, do not produce false-positive DMA-O2. Moreover, we identify a previously unreported 1O2-forming pathway through the reaction of CO2 with superoxide. Overall, we give unequivocal proof for 1O2 formation in non-aqueous oxygen redox and show that chemical trapping with DMA is a reliable method to assess 1O2 formation. AU - Mondal, Soumyadip AU - Jethwa, Rajesh B AU - Pant, Bhargavi AU - Hauschild, Robert AU - Freunberger, Stefan Alexander ID - 13044 JF - Faraday Discussions KW - Physical and Theoretical Chemistry SN - 1359-6640 TI - Singlet oxygen in non-aqueous oxygen redox: Direct spectroscopic evidence for formation pathways and reliability of chemical probes ER - TY - JOUR AB - 5-Carboxycytosine (5caC) is a rare epigenetic modification found in nucleic acids of all domains of life. Despite its sparse genomic abundance, 5caC is presumed to play essential regulatory roles in transcription, maintenance and base-excision processes in DNA. In this work, we utilize nuclear magnetic resonance (NMR) spectroscopy to address the effects of 5caC incorporation into canonical DNA strands at multiple pH and temperature conditions. Our results demonstrate that 5caC has a pH-dependent global destabilizing and a base-pair mobility enhancing local impact on dsDNA, albeit without any detectable influence on the ground-state B-DNA structure. Measurement of hybridization thermodynamics and kinetics of 5caC-bearing DNA duplexes highlighted how acidic environment (pH 5.8 and 4.7) destabilizes the double-stranded structure by ∼10–20 kJ mol–1 at 37 °C when compared to the same sample at neutral pH. Protonation of 5caC results in a lower activation energy for the dissociation process and a higher barrier for annealing. Studies on conformational exchange on the microsecond time scale regime revealed a sharply localized base-pair motion involving exclusively the modified site and its immediate surroundings. By direct comparison with canonical and 5-formylcytosine (5fC)-edited strands, we were able to address the impact of the two most oxidized naturally occurring cytosine derivatives in the genome. These insights on 5caC’s subtle sensitivity to acidic pH contribute to the long-standing questions of its capacity as a substrate in base excision repair processes and its purpose as an independent, stable epigenetic mark. AU - Dubini, Romeo C. A. AU - Korytiaková, Eva AU - Schinkel, Thea AU - Heinrichs, Pia AU - Carell, Thomas AU - Rovo, Petra ID - 10758 IS - 3 JF - ACS Physical Chemistry Au TI - 1H NMR chemical exchange techniques reveal local and global effects of oxidized cytosine derivatives VL - 2 ER - TY - JOUR AB - Immune cells are constantly on the move through multicellular organisms to explore and respond to pathogens and other harmful insults. While moving, immune cells efficiently traverse microenvironments composed of tissue cells and extracellular fibers, which together form complex environments of various porosity, stiffness, topography, and chemical composition. In this protocol we describe experimental procedures to investigate immune cell migration through microenvironments of heterogeneous porosity. In particular, we describe micro-channels, micro-pillars, and collagen networks as cell migration paths with alternative pore size choices. Employing micro-channels or micro-pillars that divide at junctions into alternative paths with initially differentially sized pores allows us to precisely (1) measure the cellular translocation time through these porous path junctions, (2) quantify the cellular preference for individual pore sizes, and (3) image cellular components like the nucleus and the cytoskeleton. This reductionistic experimental setup thus can elucidate how immune cells perform decisions in complex microenvironments of various porosity like the interstitium. The setup further allows investigation of the underlying forces of cellular squeezing and the consequences of cellular deformation on the integrity of the cell and its organelles. As a complementary approach that does not require any micro-engineering expertise, we describe the usage of three-dimensional collagen networks with different pore sizes. Whereas we here focus on dendritic cells as a model for motile immune cells, the described protocols are versatile as they are also applicable for other immune cell types like neutrophils and non-immune cell types such as mesenchymal and cancer cells. In summary, we here describe protocols to identify the mechanisms and principles of cellular probing, decision making, and squeezing during cellular movement through microenvironments of heterogeneous porosity. AU - Kroll, Janina AU - Ruiz-Fernandez, Mauricio J.A. AU - Braun, Malte B. AU - Merrin, Jack AU - Renkawitz, Jörg ID - 11182 IS - 4 JF - Current Protocols TI - Quantifying the probing and selection of microenvironmental pores by motile immune cells VL - 2 ER - TY - JOUR AB - This article investigates library-related documents written by Gerard van Swieten (1700–72) during his tenure as Library Prefect in the Imperial Library of Vienna (1745–72). Van Swieten’s time as Library Prefect is considered through a textual analysis. Handwritten letters were deconstructed in terms of their appearance, layout, and tone in order to mine them for meaning. Furthermore, the contents were examined for library matters such as censorship, catalogues, and collection development. The Imperial Court Library held a prominent role as a repository for rare and valuable works, later becoming the National Library of Austria. Gerard van Swieten’s work as a librarian tends to be overlooked, perhaps because he is better known as the private physician of Maria Theresia, as well as a medical reformer. Nevertheless, he was a hard-working chief librarian deeply involved in all aspects of librarianship. Van Swieten endorsed modern scientific works, which were otherwise banned officially by the censorship commission, for the use of scholars in the library, expanded the collection by acquiring books through his network of scholars and publishers, and reissued library catalogues. He also provided for the comfort of users in the library reading room, at a time when such considerations were unusual. In conclusion, a proposal is made that van Swieten viewed his role as librarian with some importance and pride. AU - Chlebak, Clara A AU - Reid, Peter H. ID - 11444 IS - 1 JF - Library and Information History SN - 1758-3489 TI - From the prefect’s desk: Gerard van Swieten’s library correspondence VL - 38 ER - TY - GEN AU - Schlögl, Alois AU - Hornoiu, Andrei AU - Elefante, Stefano AU - Stadlbauer, Stephan ID - 12894 SN - 978-3-200-08499-5 T2 - ASHPC22 - Austrian-Slovenian HPC Meeting 2022 TI - Where is the sweet spot? A procurement story of general purpose compute nodes ER - TY - JOUR AB - Lymph nodes (LNs) comprise two main structural elements: fibroblastic reticular cells that form dedicated niches for immune cell interaction and capsular fibroblasts that build a shell around the organ. Immunological challenge causes LNs to increase more than tenfold in size within a few days. Here, we characterized the biomechanics of LN swelling on the cellular and organ scale. We identified lymphocyte trapping by influx and proliferation as drivers of an outward pressure force, causing fibroblastic reticular cells of the T-zone (TRCs) and their associated conduits to stretch. After an initial phase of relaxation, TRCs sensed the resulting strain through cell matrix adhesions, which coordinated local growth and remodeling of the stromal network. While the expanded TRC network readopted its typical configuration, a massive fibrotic reaction of the organ capsule set in and countered further organ expansion. Thus, different fibroblast populations mechanically control LN swelling in a multitier fashion. AU - Assen, Frank P AU - Abe, Jun AU - Hons, Miroslav AU - Hauschild, Robert AU - Shamipour, Shayan AU - Kaufmann, Walter AU - Costanzo, Tommaso AU - Krens, Gabriel AU - Brown, Markus AU - Ludewig, Burkhard AU - Hippenmeyer, Simon AU - Heisenberg, Carl-Philipp J AU - Weninger, Wolfgang AU - Hannezo, Edouard B AU - Luther, Sanjiv A. AU - Stein, Jens V. AU - Sixt, Michael K ID - 9794 JF - Nature Immunology SN - 1529-2908 TI - Multitier mechanics control stromal adaptations in swelling lymph nodes VL - 23 ER - TY - JOUR AB - Tension of the actomyosin cell cortex plays a key role in determining cell–cell contact growth and size. The level of cortical tension outside of the cell–cell contact, when pulling at the contact edge, scales with the total size to which a cell–cell contact can grow [J.-L. Maître et al., Science 338, 253–256 (2012)]. Here, we show in zebrafish primary germ-layer progenitor cells that this monotonic relationship only applies to a narrow range of cortical tension increase and that above a critical threshold, contact size inversely scales with cortical tension. This switch from cortical tension increasing to decreasing progenitor cell–cell contact size is caused by cortical tension promoting E-cadherin anchoring to the actomyosin cytoskeleton, thereby increasing clustering and stability of E-cadherin at the contact. After tension-mediated E-cadherin stabilization at the contact exceeds a critical threshold level, the rate by which the contact expands in response to pulling forces from the cortex sharply drops, leading to smaller contacts at physiologically relevant timescales of contact formation. Thus, the activity of cortical tension in expanding cell–cell contact size is limited by tension-stabilizing E-cadherin–actin complexes at the contact. AU - Slovakova, Jana AU - Sikora, Mateusz K AU - Arslan, Feyza N AU - Caballero Mancebo, Silvia AU - Krens, Gabriel AU - Kaufmann, Walter AU - Merrin, Jack AU - Heisenberg, Carl-Philipp J ID - 10766 IS - 8 JF - Proceedings of the National Academy of Sciences of the United States of America TI - Tension-dependent stabilization of E-cadherin limits cell-cell contact expansion in zebrafish germ-layer progenitor cells VL - 119 ER - TY - JOUR AB - In eukaryotes, clathrin-coated vesicles (CCVs) facilitate the internalization of material from the cell surface as well as the movement of cargo in post-Golgi trafficking pathways. This diversity of functions is partially provided by multiple monomeric and multimeric clathrin adaptor complexes that provide compartment and cargo selectivity. The adaptor-protein assembly polypeptide-1 (AP-1) complex operates as part of the secretory pathway at the trans-Golgi network (TGN), while the AP-2 complex and the TPLATE complex jointly operate at the plasma membrane to execute clathrin-mediated endocytosis. Key to our further understanding of clathrin-mediated trafficking in plants will be the comprehensive identification and characterization of the network of evolutionarily conserved and plant-specific core and accessory machinery involved in the formation and targeting of CCVs. To facilitate these studies, we have analyzed the proteome of enriched TGN/early endosome-derived and endocytic CCVs isolated from dividing and expanding suspension-cultured Arabidopsis (Arabidopsis thaliana) cells. Tandem mass spectrometry analysis results were validated by differential chemical labeling experiments to identify proteins co-enriching with CCVs. Proteins enriched in CCVs included previously characterized CCV components and cargos such as the vacuolar sorting receptors in addition to conserved and plant-specific components whose function in clathrin-mediated trafficking has not been previously defined. Notably, in addition to AP-1 and AP-2, all subunits of the AP-4 complex, but not AP-3 or AP-5, were found to be in high abundance in the CCV proteome. The association of AP-4 with suspension-cultured Arabidopsis CCVs is further supported via additional biochemical data. AU - Dahhan, DA AU - Reynolds, GD AU - Cárdenas, JJ AU - Eeckhout, D AU - Johnson, Alexander J AU - Yperman, K AU - Kaufmann, Walter AU - Vang, N AU - Yan, X AU - Hwang, I AU - Heese, A AU - De Jaeger, G AU - Friml, Jiří AU - Van Damme, D AU - Pan, J AU - Bednarek, SY ID - 10841 IS - 6 JF - Plant Cell SN - 1040-4651 TI - Proteomic characterization of isolated Arabidopsis clathrin-coated vesicles reveals evolutionarily conserved and plant-specific components VL - 34 ER - TY - JOUR AB - The broad implementation of thermoelectricity requires high-performance and low-cost materials. One possibility is employing surfactant-free solution synthesis to produce nanopowders. We propose the strategy of functionalizing “naked” particles’ surface by inorganic molecules to control the nanostructure and, consequently, thermoelectric performance. In particular, we use bismuth thiolates to functionalize surfactant-free SnTe particles’ surfaces. Upon thermal processing, bismuth thiolates decomposition renders SnTe-Bi2S3 nanocomposites with synergistic functions: 1) carrier concentration optimization by Bi doping; 2) Seebeck coefficient enhancement and bipolar effect suppression by energy filtering; and 3) lattice thermal conductivity reduction by small grain domains, grain boundaries and nanostructuration. Overall, the SnTe-Bi2S3 nanocomposites exhibit peak z T up to 1.3 at 873 K and an average z T of ≈0.6 at 300–873 K, which is among the highest reported for solution-processed SnTe. AU - Chang, Cheng AU - Liu, Yu AU - Lee, Seungho AU - Spadaro, Maria AU - Koskela, Kristopher M. AU - Kleinhanns, Tobias AU - Costanzo, Tommaso AU - Arbiol, Jordi AU - Brutchey, Richard L. AU - Ibáñez, Maria ID - 11705 IS - 35 JF - Angewandte Chemie - International Edition SN - 1433-7851 TI - Surface functionalization of surfactant-free particles: A strategy to tailor the properties of nanocomposites for enhanced thermoelectric performance VL - 61 ER - TY - JOUR AB - Capacity, rate performance, and cycle life of aprotic Li–O2 batteries critically depend on reversible electrodeposition of Li2O2. Current understanding states surface-adsorbed versus solvated LiO2 controls Li2O2 growth as surface film or as large particles. Herein, we show that Li2O2 forms across a wide range of electrolytes, carbons, and current densities as particles via solution-mediated LiO2 disproportionation, bringing into question the prevalence of any surface growth under practical conditions. We describe a unified O2 reduction mechanism, which can explain all found capacity relations and Li2O2 morphologies with exclusive solution discharge. Determining particle morphology and achievable capacities are species mobilities, true areal rate, and the degree of LiO2 association in solution. Capacity is conclusively limited by mass transport through the tortuous Li2O2 rather than electron transport through a passivating Li2O2 film. Provided that species mobilities and surface growth are high, high capacities are also achieved with weakly solvating electrolytes, which were previously considered prototypical for low capacity via surface growth. AU - Prehal, Christian AU - Mondal, Soumyadip AU - Lovicar, Ludek AU - Freunberger, Stefan Alexander ID - 12065 IS - 9 JF - ACS Energy Letters TI - Exclusive solution discharge in Li-O₂ batteries? VL - 7 ER - TY - JOUR AB - Kelvin probe force microscopy (KPFM) is a powerful tool for studying contact electrification (CE) at the nanoscale, but converting KPFM voltage maps to charge density maps is nontrivial due to long-range forces and complex system geometry. Here we present a strategy using finite-element method (FEM) simulations to determine the Green's function of the KPFM probe/insulator/ground system, which allows us to quantitatively extract surface charge. Testing our approach with synthetic data, we find that accounting for the atomic force microscope (AFM) tip, cone, and cantilever is necessary to recover a known input and that existing methods lead to gross miscalculation or even the incorrect sign of the underlying charge. Applying it to experimental data, we demonstrate its capacity to extract realistic surface charge densities and fine details from contact-charged surfaces. Our method gives a straightforward recipe to convert qualitative KPFM voltage data into quantitative charge data over a range of experimental conditions, enabling quantitative CE at the nanoscale. AU - Pertl, Felix AU - Sobarzo Ponce, Juan Carlos A AU - Shafeek, Lubuna B AU - Cramer, Tobias AU - Waitukaitis, Scott R ID - 12109 IS - 12 JF - Physical Review Materials TI - Quantifying nanoscale charge density features of contact-charged surfaces with an FEM/KPFM-hybrid approach VL - 6 ER - TY - JOUR AB - Muskelin (Mkln1) is implicated in neuronal function, regulating plasma membrane receptor trafficking. However, its influence on intrinsic brain activity and corresponding behavioral processes remains unclear. Here we show that murine Mkln1 knockout causes non-habituating locomotor activity, increased exploratory drive, and decreased locomotor response to amphetamine. Muskelin deficiency impairs social novelty detection while promoting the retention of spatial reference memory and fear extinction recall. This is strongly mirrored in either weaker or stronger resting-state functional connectivity between critical circuits mediating locomotor exploration and cognition. We show that Mkln1 deletion alters dendrite branching and spine structure, coinciding with enhanced AMPAR-mediated synaptic transmission but selective impairment in synaptic potentiation maintenance. We identify muskelin at excitatory synapses and highlight its role in regulating dendritic spine actin stability. Our findings point to aberrant spine actin modulation and changes in glutamatergic synaptic function as critical mechanisms that contribute to the neurobehavioral phenotype arising from Mkln1 ablation. AU - Muhia, Mary W AU - YuanXiang, PingAn AU - Sedlacik, Jan AU - Schwarz, Jürgen R. AU - Heisler, Frank F. AU - Gromova, Kira V. AU - Thies, Edda AU - Breiden, Petra AU - Pechmann, Yvonne AU - Kreutz, Michael R. AU - Kneussel, Matthias ID - 12224 JF - Communications Biology KW - General Agricultural and Biological Sciences KW - General Biochemistry KW - Genetics and Molecular Biology KW - Medicine (miscellaneous) SN - 2399-3642 TI - Muskelin regulates actin-dependent synaptic changes and intrinsic brain activity relevant to behavioral and cognitive processes VL - 5 ER - TY - JOUR AB - The question of how RNA, as the principal carrier of genetic information evolved is fundamentally important for our understanding of the origin of life. The RNA molecule is far too complex to have formed in one evolutionary step, suggesting that ancestral proto-RNAs (first ancestor of RNA) may have existed, which evolved over time into the RNA of today. Here we show that isoxazole nucleosides, which are quickly formed from hydroxylamine, cyanoacetylene, urea and ribose, are plausible precursors for RNA. The isoxazole nucleoside can rearrange within an RNA-strand to give cytidine, which leads to an increase of pairing stability. If the proto-RNA contains a canonical seed-nucleoside with defined stereochemistry, the seed-nucleoside can control the configuration of the anomeric center that forms during the in-RNA transformation. The results demonstrate that RNA could have emerged from evolutionarily primitive precursor isoxazole ribosides after strand formation. AU - Xu, Felix AU - Crisp, Antony AU - Schinkel, Thea AU - Dubini, Romeo C. A. AU - Hübner, Sarah AU - Becker, Sidney AU - Schelter, Florian AU - Rovo, Petra AU - Carell, Thomas ID - 12228 IS - 45 JF - Angewandte Chemie International Edition KW - General Chemistry KW - Catalysis SN - 1433-7851 TI - Isoxazole nucleosides as building blocks for a plausible proto‐RNA VL - 61 ER - TY - JOUR AB - Biological systems are the sum of their dynamic three-dimensional (3D) parts. Therefore, it is critical to study biological structures in 3D and at high resolution to gain insights into their physiological functions. Electron microscopy of metal replicas of unroofed cells and isolated organelles has been a key technique to visualize intracellular structures at nanometer resolution. However, many of these methods require specialized equipment and personnel to complete them. Here, we present novel accessible methods to analyze biological structures in unroofed cells and biochemically isolated organelles in 3D and at nanometer resolution, focusing on Arabidopsis clathrin-coated vesicles (CCVs). While CCVs are essential trafficking organelles, their detailed structural information is lacking due to their poor preservation when observed via classical electron microscopy protocols experiments. First, we establish a method to visualize CCVs in unroofed cells using scanning transmission electron microscopy tomography, providing sufficient resolution to define the clathrin coat arrangements. Critically, the samples are prepared directly on electron microscopy grids, removing the requirement to use extremely corrosive acids, thereby enabling the use of this method in any electron microscopy lab. Secondly, we demonstrate that this standardized sample preparation allows the direct comparison of isolated CCV samples with those visualized in cells. Finally, to facilitate the high-throughput and robust screening of metal replicated samples, we provide a deep learning analysis method to screen the “pseudo 3D” morphologies of CCVs imaged with 2D modalities. Collectively, our work establishes accessible ways to examine the 3D structure of biological samples and provide novel insights into the structure of plant CCVs. AU - Johnson, Alexander J AU - Kaufmann, Walter AU - Sommer, Christoph M AU - Costanzo, Tommaso AU - Dahhan, Dana A. AU - Bednarek, Sebastian Y. AU - Friml, Jiří ID - 12239 IS - 10 JF - Molecular Plant KW - Plant Science KW - Molecular Biology SN - 1674-2052 TI - Three-dimensional visualization of planta clathrin-coated vesicles at ultrastructural resolution VL - 15 ER - TY - JOUR AB - Theoretical foundations of chaos have been predominantly laid out for finite-dimensional dynamical systems, such as the three-body problem in classical mechanics and the Lorenz model in dissipative systems. In contrast, many real-world chaotic phenomena, e.g., weather, arise in systems with many (formally infinite) degrees of freedom, which limits direct quantitative analysis of such systems using chaos theory. In the present work, we demonstrate that the hydrodynamic pilot-wave systems offer a bridge between low- and high-dimensional chaotic phenomena by allowing for a systematic study of how the former connects to the latter. Specifically, we present experimental results, which show the formation of low-dimensional chaotic attractors upon destabilization of regular dynamics and a final transition to high-dimensional chaos via the merging of distinct chaotic regions through a crisis bifurcation. Moreover, we show that the post-crisis dynamics of the system can be rationalized as consecutive scatterings from the nonattracting chaotic sets with lifetimes following exponential distributions. AU - Choueiri, George H AU - Suri, Balachandra AU - Merrin, Jack AU - Serbyn, Maksym AU - Hof, Björn AU - Budanur, Nazmi B ID - 12259 IS - 9 JF - Chaos: An Interdisciplinary Journal of Nonlinear Science KW - Applied Mathematics KW - General Physics and Astronomy KW - Mathematical Physics KW - Statistical and Nonlinear Physics SN - 1054-1500 TI - Crises and chaotic scattering in hydrodynamic pilot-wave experiments VL - 32 ER - TY - JOUR AB - The AAA-ATPase Drg1 is a key factor in eukaryotic ribosome biogenesis that initiates cytoplasmic maturation of the large ribosomal subunit. Drg1 releases the shuttling maturation factor Rlp24 from pre-60S particles shortly after nuclear export, a strict requirement for downstream maturation. The molecular mechanism of release remained elusive. Here, we report a series of cryo-EM structures that captured the extraction of Rlp24 from pre-60S particles by Saccharomyces cerevisiae Drg1. These structures reveal that Arx1 and the eukaryote-specific rRNA expansion segment ES27 form a joint docking platform that positions Drg1 for efficient extraction of Rlp24 from the pre-ribosome. The tips of the Drg1 N domains thereby guide the Rlp24 C terminus into the central pore of the Drg1 hexamer, enabling extraction by a hand-over-hand translocation mechanism. Our results uncover substrate recognition and processing by Drg1 step by step and provide a comprehensive mechanistic picture of the conserved modus operandi of AAA-ATPases. AU - Prattes, Michael AU - Grishkovskaya, Irina AU - Hodirnau, Victor-Valentin AU - Hetzmannseder, Christina AU - Zisser, Gertrude AU - Sailer, Carolin AU - Kargas, Vasileios AU - Loibl, Mathias AU - Gerhalter, Magdalena AU - Kofler, Lisa AU - Warren, Alan J. AU - Stengel, Florian AU - Haselbach, David AU - Bergler, Helmut ID - 12262 IS - 9 JF - Nature Structural & Molecular Biology KW - Molecular Biology KW - Structural Biology SN - 1545-9993 TI - Visualizing maturation factor extraction from the nascent ribosome by the AAA-ATPase Drg1 VL - 29 ER -