@article{17148, abstract = {During neural tube (NT) development, the notochord induces an organizer, the floorplate, which secretes Sonic Hedgehog (SHH) to pattern neural progenitors. Conversely, NT organoids (NTOs) from embryonic stem cells (ESCs) spontaneously form floorplates without the notochord, demonstrating that stem cells can self-organize without embryonic inducers. Here, we investigated floorplate self-organization in clonal mouse NTOs. Expression of the floorplate marker FOXA2 was initially spatially scattered before resolving into multiple clusters, which underwent competition and sorting, resulting in a stable “winning” floorplate. We identified that BMP signaling governed long-range cluster competition. FOXA2+ clusters expressed BMP4, suppressing FOXA2 in receiving cells while simultaneously expressing the BMP-inhibitor NOGGIN, promoting cluster persistence. Noggin mutation perturbed floorplate formation in NTOs and in the NT in vivo at mid/hindbrain regions, demonstrating how the floorplate can form autonomously without the notochord. Identifying the pathways governing organizer self-organization is critical for harnessing the developmental plasticity of stem cells in tissue engineering.}, author = {Krammer, Teresa and Stuart, Hannah T. and Gromberg, Elena and Ishihara, Keisuke and Cislo, Dillon and Melchionda, Manuela and Becerril Perez, Fernando and Wang, Jingkui and Costantini, Elena and Rus, Stefanie and Arbanas, Laura and Hörmann, Alexandra and Neumüller, Ralph A. and Elvassore, Nicola and Siggia, Eric and Briscoe, James and Kicheva, Anna and Tanaka, Elly M.}, issn = {1878-1551}, journal = {Developmental Cell}, number = {15}, pages = {1940--1953.e10}, publisher = {Elsevier}, title = {{Mouse neural tube organoids self-organize floorplate through BMP-mediated cluster competition}}, doi = {10.1016/j.devcel.2024.04.021}, volume = {59}, year = {2024}, } @article{18601, abstract = {Geometrically controlled stem cell differentiation promotes reproducible pattern formation. Here, we present a protocol to fabricate elastomeric stencils for patterned stem cell differentiation. We describe procedures for using photolithography to produce molds, followed by molding polydimethylsiloxane (PDMS) to obtain stencils with through holes. We then provide instructions for culturing cells on stencils and, finally, removing stencils to allow colony growth and cell migration. This approach yields reproducible two-dimensional organoids tailored for quantitative studies of growth and pattern formation. For complete details on the use and execution of this protocol, please refer to Lehr et al.1}, author = {Rus, Stefanie and Merrin, Jack and Kulig, Monika Aleksandra and Minchington, Thomas and Kicheva, Anna}, issn = {2666-1667}, journal = {STAR Protocols}, number = {4}, publisher = {Elsevier}, title = {{Protocol for fabricating elastomeric stencils for patterned stem cell differentiation}}, doi = {10.1016/j.xpro.2024.103187}, volume = {5}, year = {2024}, } @article{17890, abstract = {Our understanding of the molecular pathways that regulate oogenesis and define cellular identity in the Arthropod female reproductive system and the extent of their conservation is currently very limited. This is due to the focus on model systems, including Drosophila and Daphnia, which do not reflect the observed diversity of morphologies, reproductive modes, and sex chromosome systems. We use single-nucleus RNA and ATAC sequencing to produce a comprehensive single nucleus atlas of the adult Artemia franciscana female reproductive system. We map our data to the Fly Cell Atlas single-nucleus dataset of the Drosophila melanogaster ovary, shedding light on the conserved regulatory programs between the two distantly related Arthropod species. We identify the major cell types known to be present in the Artemia ovary, including germ cells, follicle cells, and ovarian muscle cells. Additionally, we use the germ cells to explore gene regulation and expression of the Z chromosome during meiosis, highlighting its unique regulatory dynamics and allowing us to explore the presence of meiotic sex chromosome silencing in this group.}, author = {Elkrewi, Marwan N and Vicoso, Beatriz}, issn = {1553-7404}, journal = {PLoS Genetics}, number = {8}, publisher = {Public Library of Science}, title = {{Single-nucleus atlas of the Artemia female reproductive system suggests germline repression of the Z chromosome}}, doi = {10.1371/journal.pgen.1011376}, volume = {20}, year = {2024}, } @article{18525, abstract = {As their statistical power grows, genome-wide association studies (GWAS) have identified an increasing number of loci underlying quantitative traits of interest. These loci are scattered throughout the genome and are individually responsible only for small fractions of the total heritable trait variance. The recently proposed omnigenic model provides a conceptual framework to explain these observations by postulating that numerous distant loci contribute to each complex trait via effect propagation through intracellular regulatory networks. We formalize this conceptual framework by proposing the “quantitative omnigenic model” (QOM), a statistical model that combines prior knowledge of the regulatory network topology with genomic data. By applying our model to gene expression traits in yeast, we demonstrate that QOM achieves similar gene expression prediction performance to traditional GWAS with hundreds of times less parameters, while simultaneously extracting candidate causal and quantitative chains of effect propagation through the regulatory network for every individual gene. We estimate the fraction of heritable trait variance in cis- and in trans-, break the latter down by effect propagation order, assess the trans- variance not attributable to transcriptional regulation, and show that QOM correctly accounts for the low-dimensional structure of gene expression covariance. We furthermore demonstrate the relevance of QOM for systems biology, by employing it as a statistical test for the quality of regulatory network reconstructions, and linking it to the propagation of nontranscriptional (including environmental) effects.}, author = {Ruzickova, Natalia and Hledik, Michal and Tkačik, Gašper}, issn = {1091-6490}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, number = {44}, publisher = {National Academy of Sciences}, title = {{Quantitative omnigenic model discovers interpretable genome-wide associations}}, doi = {10.1073/pnas.2402340121}, volume = {121}, year = {2024}, } @phdthesis{17881, abstract = {This work can be broadly classified into the study of critical phenomena in a one dimensional array of Josephson junctions. While we study quantum criticality when the array is in thermal equilibrium at zero bias, the non-equilibrium study involves understanding the bistability of the array at a critical non-zero bias. This work furthers our knowledge in understanding quantum critical behaviour at finite temperatures in a one dimensional Josephson array, while also establishing relaxation behaviour dual to that observed in a single Josephson junction. Chapter 1 briefly introduces the model to understand superconductor-insulator phase transition in a one dimensional Josephson array and points out the state of the field from where we started our zero-bias experiments. In this context it discusses the phase-charge duality observed in a Josephson array and its dual hysteretic behaviour to that of a single junction, setting the ground for our non-equilibrium study of the array. Chapter 2 shows the experimental setup and the chip layout of the device we measured. In chapter 3 we show that, unlike the typical quantum-critical broadening scenario, in one dimensional Josephson arrays temperature dramatically shifts the critical region. This shift leads to a regime of superconductivity at high temperature, arising from the melted zero-temperature insulator. Our results quantitatively explain the low-temperature onset of superconductivity in nominally insulating regimes, and the transition to the strongly insulating phase. We further present, to our knowledge, the first understanding of the onset of anomalous-metallic resistance saturation [30]. This work demonstrates a non-trivial interplay between thermal effects and quantum criticality. A practical consequence is that, counterintuitively, the coherence of high-impedance quantum circuits is expected to be stabilized by thermal fluctuations. In chapter 4, we show relaxation oscillations in a current-biased one dimensional array of Josephson junctions. These oscillations are well described by a circuit model, dual to the ordinary Josephson relaxation oscillations [72]. Injection locking these oscillations results in current plateaux. The relaxation step is found to obey a characteristic self-consistent relation, suggesting that it is governed by overheating effects. Chapter 5 describes the various checks and analysis we performed to support our conclusions made in chapters 3 and 4. Finally, chapter 6 describes the nanofabrication steps and the finite element electromagnetic simulations we performed to fabricate our devices.}, author = {Mukhopadhyay, Soham}, isbn = {978-3-99078-043-5}, issn = {2663-337X}, pages = {82}, publisher = {Institute of Science and Technology Austria}, title = {{Thermal effects in one dimensional Josephson chains}}, doi = {10.15479/at:ista:17881}, year = {2024}, } @article{15009, abstract = {Since the commercialization of brine shrimp (genus Artemia) in the 1950s, this lineage, and in particular the model species Artemia franciscana, has been the subject of extensive research. However, our understanding of the genetic mechanisms underlying various aspects of their reproductive biology, including sex determination, is still lacking. This is partly due to the scarcity of genomic resources for Artemia species and crustaceans in general. Here, we present a chromosome-level genome assembly of A. franciscana (Kellogg 1906), from the Great Salt Lake, United States. The genome is 1 GB, and the majority of the genome (81%) is scaffolded into 21 linkage groups using a previously published high-density linkage map. We performed coverage and FST analyses using male and female genomic and transcriptomic reads to quantify the extent of differentiation between the Z and W chromosomes. Additionally, we quantified the expression levels in male and female heads and gonads and found further evidence for dosage compensation in this species.}, author = {Bett, Vincent K and Macon, Ariana and Vicoso, Beatriz and Elkrewi, Marwan N}, issn = {1759-6653}, journal = {Genome Biology and Evolution}, number = {1}, publisher = {Oxford University Press}, title = {{Chromosome-level assembly of Artemia franciscana sheds light on sex chromosome differentiation}}, doi = {10.1093/gbe/evae006}, volume = {16}, year = {2024}, } @unpublished{18057, abstract = {We report relaxation oscillations in a one-dimensional array of Josephson junctions. The oscillations are circuit-dual to those ordinarily observed in single junctions. The dual circuit quantitatively accounts for temporal dynamics of the array, including the dependence on biasing conditions. Injection locking the oscillations results in well-developed current plateaux. A thermal model explains the relaxation step of the oscillations.}, author = {Mukhopadhyay, Soham and Lancheros Naranjo, Diego A and Senior, Jorden L and Higginbotham, Andrew P}, booktitle = {arXiv}, title = {{Dual relaxation oscillations in a Josephson junction array}}, doi = {10.48550/arXiv.2408.07829}, year = {2024}, } @phdthesis{18531, abstract = {Sex chromosomes and autosomes exhibit very different evolutionary dynamics. The Y chromosome usually degenerates, leaving many X-linked loci hemizygous in males. Since recessive X-linked mutations are always exposed to selection in males, selection is more efficient on the X chromosome than on autosomes on recessive mutations, leading to faster adaptation on the X chromosome than other genomic regions, if beneficial mutations are on average recessive (known as the Faster-X effect). In the presence of the functional, but non-recombining gametolog on the Y (as is often the case in young non-recombining regions), recessive mutations are sheltered from selection on the X chromosome. We model this scenario and show that the efficiency of selection is reduced on diploid X loci due to sheltering by the Y chromosome. Reduced efficiency of selection leads to slower adaptation and increased accumulation of deleterious mutations (Slower-X effect). We extended this model to explore the effect of sex-specific selection on degeneration of sex chromosomes, showing theoretically that male-limited genes degenerate on the X chromosome and female-biased genes degenerate on the Y chromosome. This prediction depends on the effective population size and the mutation rate, explaining the variety of sex chromosome degeneration patterns observed in nature. To test for direct evidence of a Slower-X (or Slower-Z) effect, we analyzed the ZW sex chromosomes of the flatworm Schistosoma japonicum, which have a very young non-recombining region with non-degenerated W. Diploid Z-linked genes have higher ratios of non-synonymous to synonymous polymorphisms than autosomal genes, supporting reduced efficiency of selection on the diploid Z region. These results provide evidence of sheltering by the W chromosome, a mechanism that could contribute to Z (X) chromosome degeneration, and illustrate contrasting evolutionary patterns in old and young sex chromosome regions. In addition, genes with sexspecific patterns of expression show opposite patterns of selection in the young (diploid) and old (hemizygous) Z, showing the complex manner in which sex-specific selection shapes the evolutionary patterns of sex chromosomes. }, author = {Mrnjavac, Andrea}, issn = {2663-337X}, keywords = {Sex chromosomes, evolution, selection, sheltering}, pages = {181}, publisher = {Institute of Science and Technology Austria}, title = {{Early stages of sex chromosome evolution}}, doi = {10.15479/at:ista:18531}, year = {2024}, } @phdthesis{18661, abstract = {Across the tree of life, distinct designs of cellular membranes have evolved that are both stable and flexible. In bacteria and eukaryotes this trade-off is accomplished by single-headed lipids that self-assemble into flexible bilayer membranes. By contrast, archaea in many cases possess both bilayer and double-headed, monolayer spanning bolalipids. This composition is believed to enable extremophile archaea to survive harsh environments. Here, through the creation of a minimal computational model for bolalipid membranes, we discover trade-offs when forming membranes using lipids of a single type. Similar to living archaea, we can tune the stiffness of bolalipid molecules. We find that membranes made out of flexible bolalipid molecules resemble bilayer membranes as they can adopt U-shaped conformations to enable higher curvatures. Conversely, rigid bolalipid molecules, like those found in archaea at higher temperatures, preferentially take on a straight conformation to self-assemble into liquid membranes that are stable, stiff, prone to pore formation, and which tear during membrane reshaping. Strikingly, however, our analysis reveals that it is possible to achieve the best of both worlds – membranes that are fluid, stable at high temperatures and flexible enough to be reshaped without leaking – through the inclusion of a small fraction of bilayer lipids into a bolalipid membrane. Additionally, the curvature-dependent softening of bolalipid membranes made of lipids with tension-sensitive conformation can also enable high rigidity at low curvatures while softening at high curvatures, making the membrane effectively a plastic material. Taken together, our study compares the different membrane designs across the tree of life and indicates how combining lipids can be used to resolve trade-offs when generating membranes for (bio)technological applications. }, author = {Santana de Freitas Amaral, Miguel}, isbn = {978-3-99078-046-6}, issn = {2663-337X}, pages = {57}, publisher = {Institute of Science and Technology Austria}, title = {{Archaeal membranes: In silico modelling and design}}, doi = {10.15479/at:ista:18661}, year = {2024}, } @phdthesis{18104, abstract = {We introduce a new all-electric platform, that strong couples light to mechanical motion by ensuring that the external environmental coupling dominates over internal mechanical dissipation. The system only has three everyday components: AC, DC, and a fip-chip, in which a metallized silicon nitride membrane is fipped on top of the device under test. This everyday electromechanical device can be operated at low or room temperature and has 10000× lower insertion loss than a comparable commercial quartz crystal, achieves a position imprecision matching state-of-the-art optical interferometer, and enables remote cooling of mechanical motion. The spatial properties of higher order mechanical modes are a promising feature for reconstructing unknown charge distributions. }, author = {Puglia, Denise}, issn = {2663-337X}, pages = {63}, publisher = {Institute of Science and Technology Austria}, title = {{Everyday electromechanics: Capacitive strong coupling to mechanical motion}}, doi = {10.15479/at:ista:18104}, year = {2024}, } @unpublished{18143, abstract = {Strong optomechanical coupling -- a regime where mechanical motion is damped by environmental radiation -- has traditionally required demanding experimental ingredients such as superconducting resonators, high-quality optical cavities, or large magnetic fields. Here we demonstrate a room temperature, cavity-free, all-electric device reaching this regime at radio frequencies, enabled by a mechanically compliant parallel-plate capacitor with a nanoscale plate separation and an aspect ratio exceeding 1,000. The device has four orders of magnitude lower insertion loss than a comparable commercial quartz crystal, and achieves a position imprecision rivaling an optical interferometer. With the help of a back-action isolation scheme, we observe radiative cooling of mechanical motion by a remote cryogenic load. This work provides a technologically accessible route to high-precision sensing, transduction, and signal processing.}, author = {Puglia, Denise and Odessey, Rachel H and Burns, Peter S. and Luhmann, Niklas and Schmid, Silvan and Higginbotham, Andrew P}, booktitle = {arXiv}, title = {{Room temperature, cavity-free capacitive strong coupling to mechanical motion}}, doi = {10.48550/arXiv.2407.15314}, year = {2024}, } @unpublished{18549, abstract = {Sex-linked and autosomal loci experience different selective pressures and evolutionary dynamics. X (or Z) chromosomes are often hemizygous, as Y (or W) chromosomes often degenerate. Such hemizygous regions can be under greater efficacy of selection, as recessive mutations are immediately exposed to selection in the heterogametic sex (the so-called Faster-X or Faster-Z effect). However, in young non-recombining regions, Y/W chromosomes often have many functional genes, and many X/Z-linked loci are therefore diploid. The sheltering of recessive mutations on the X/Z by the Y/W homolog is expected to drive a Slower-X (Slower-Z) effect for diploid X/Z loci, i.e. a reduction in the efficacy of selection. While the Faster-X effect has been studied extensively, much less is known empirically about the evolutionary dynamics of diploid X or Z chromosomes. Here, we took advantage of published population genomic data in the female-heterogametic human parasite Schistosoma japonicum to characterize the gene content and diversity levels of the diploid and hemizygous regions of the Z chromosome. We used different metrics of selective pressures acting on genes to test for differences in the efficacy of selection in hemizygous and diploid Z regions, relative to autosomes. We found consistent patterns suggesting reduced Ne, and reduced efficacy of purifying selection, on both hemizygous and diploid Z regions. Moreover, relaxed selection was particularly pronounced for female-biased genes on the diploid Z, as predicted by Slower-Z theory. }, author = {Mrnjavac, Andrea and Vicoso, Beatriz}, booktitle = {bioRxiv}, title = {{Evidence of a Slower-Z effect in Schistosoma japonicum}}, doi = {10.1101/2024.07.02.601697}, year = {2024}, } @unpublished{18670, abstract = {Across the tree of life, distinct designs of cellular membranes have evolved. In bacteria and eukaryotes single-headed lipids self-assemble into flexible bilayer membranes. By contrast, archaea often possess double-headed, monolayer spanning bolalipids, mixed with bilayer lipids, enabling them to survive in harsh environments. Here, using a minimal computational model for bolalipid membranes, we discover trade-offs when forming membranes. We find that membranes made out of flexible bolalipids resemble bilayer membranes as bolalipids exhibit conformational switch into U-shaped conformations to enable higher curvatures. Conversely, stiffer bolalipids, resembling those in extremophile archaea, take on straight conformations and form liquid membranes that are stiff, and prone to pore formation during membrane reshaping. Strikingly, we show how to achieve fluid bolalipid membranes that are both stable and flexible – by including small amounts of bilayer lipids, as archaea do. Our study explains how different organisms resolve trade-offs when generating membranes of desired material properties.}, author = {Santana de Freitas Amaral, Miguel and Frey, Felix F and Jiang, Xiuyun and Baum, Buzz and Šarić, Anđela}, booktitle = {bioRxiv}, title = {{Stability vs flexibility: Reshaping archaeal membranes in silico}}, doi = {10.1101/2024.10.18.619072}, year = {2024}, } @phdthesis{17465, abstract = {In the modern age of machine learning, artificial neural networks have become an integral part of many practical systems. One of the key ingredients of the success of the deep learning approach is recent computational advances which allowed the training of models with billions of parameters on large-scale data. Such over-parameterized and data-hungry regimes pose a challenge for the theoretical analysis of modern models since “classical” statistical wisdom is no longer applicable. In this view, it is paramount to extend or develop new machinery that will allow tackling the neural network analysis under new challenging asymptotic regimes, which is the focus of this thesis. Large neural network systems are usually optimized via “local” search algorithms, such as stochastic gradient descent (SGD). However, given the high-dimensional nature of the parameter space, it is a priori not clear why such a crude “local” approach works so remarkably well in practice. We take a step towards demystifying this phenomenon by showing that the landscape of the SGD training dynamics exhibits a few beneficial properties for the optimization. First, we show that along the SGD trajectory an over-parameterized network is dropout stable. The emergence of dropout stability allows to conclude that the minima found by SGD are connected via a continuous path of small loss. This in turn means that the high-dimensional landscape of the neural network optimization problem is provably not so unfavourable to gradient-based training, due to mode connectivity. Next, we show that SGD for an over-parameterized network tends to find solutions that are functionally more “simple”. This in turn means that the SGD minima are more robust, since a less complicated solution will less likely overfit the data. More formally, for a prototypical example of a wide two-layer ReLU network on a 1d regression task we show that the SGD algorithm is implicitly selective in its choice of an interpolating solution. Namely, at convergence the neural network implements a piece-wise linear function with the number of linear regions depending only on the amount of training data. This is in contrast to a “smooth”-like behaviour which one would expect given such a severe over-parameterization of the model. Diverging from the generic supervised setting of classification and regression problems, we analyze an auto-encoder model that is commonly used for representation learning and data compression. Despite the wide applicability of the auto-encoding paradigm, the theoretical understanding of their behaviour is limited even in the simplistic shallow case. The related work is restricted to extreme asymptotic regimes in which the auto-encoder is either severely over-parameterized or under-parameterized. In contrast, we provide a tight characterization for the 1-bit compression of Gaussian signals in the challenging proportional regime, i.e., the input dimension and the size of the compressed representation obey the same asymptotics. We also show that gradient-based methods are able to find a globally optimal solution and that the predictions made for Gaussian data extrapolate beyond - to the case of compression of natural images. Next, we relax the Gaussian assumption and study more structured input sources. We show that the shallow model is sometimes agnostic to the structure of the data vii which results in a Gaussian-like behaviour. We prove that making the decoding component slightly less shallow is already enough to escape the “curse” of Gaussian performance. }, author = {Shevchenko, Aleksandr}, issn = {2663-337X}, pages = {232}, publisher = {Institute of Science and Technology Austria}, title = {{High-dimensional limits in artificial neural networks}}, doi = {10.15479/at:ista:17465}, year = {2024}, } @phdthesis{17119, abstract = {Genomes are shaped by natural selection at the level of the organism, as genomic variants that have a beneficial effect on the viability or fecundity of their carriers are on average expected to be passed on to more offspring than less beneficial alleles. However, selection also favors genomic variants that drive their own transmission to the next generation above the mendelian expectation of 50 percent in heterozygotes, even if these self-promoting variants are less beneficial to the organism than other variants at the same locus. Such variants, called meiotic drivers, are found in diverse taxa, and often impose fitness costs on their host organisms. As meiotic drivers often require multiple genes and sequences for transmission ratio distortion, they are often found in regions of low recombination, such as inversions, which prevent their recombination with the non-driving homologous regions. Reduced recombination rates are expected to lead to the accumulation of deleterious mutations, which may affect hundreds of genes trapped in the inversions of meiotic drivers. Although the observed fitness costs of self-promoting haplotypes are thought to possibly reflect sequence degeneration, no study has systematically investigated the level of degeneration on a meiotic driver. Further, the low rates of recombination between driving and non-driving haplotypes have limited the power of traditional genetic studies in uncovering the gene content of meiotic drivers, and made the the identification of the genes causing transmission ratio distortion difficult. After an introduction to meiotic drivers in Chapter 1, this thesis presents three studies that make use of next generation sequencing data to characterize the sequence and expression evolution of genes on the t-haplotype, a large and ancient meiotic driver in house mice that is transmitted to up to 100% of the offspring in males heterozygous for it. Chapter 2 presents a comprehensive assessment of the t-haplotype’s sequence evolution, which shows signs of sequence degeneration counteracted by occasional recombination with the non-driving homolog over large parts of the meiotic driver, proposing an explanation for its long-term survival. Chapter 3 investigates the sequence and expression evolution of genes on the t-haplotype, and finds widespread expression and copy number changes and signs of less efficient purifying selection compared to the genes on the non-driving homolog. Further, this chapter finds candidates for involvment in drive: two positively selected genes on the t-haplotype, and the discovery of a t-specific gene duplicate, which was gained from another chromosome, and which acquired novel sequence and testis-specific expression on the t-haplotype. Finally, Chapter 4 provides unprecedented insights into the gene expression landscape in testes of t-carrier mice, using single nucleus sequencing. Cell-resolved RNA-sequencing allows the comparison of expression in spermatids carrying or not carrying the t-haplotype as well as the timing of t-haplotype-induced expression changes along spermatogenesis. This study shows the timing of previously found drive-associated genes, and uncovers novel candidate genes and biological processes that may underlie the complex biology of transmission ratio distortion of the t-haplotype. Chapter 5 synthesizes the findings of the three studies, and discusses them in the context of the current state of meiotic drive research.}, author = {Kelemen, Réka K}, isbn = {978-3-99078-039-8}, issn = {2663-337X}, keywords = {meiotic driver, neofunctionalization, single nucleus sequencing}, pages = {105}, publisher = {Institute of Science and Technology Austria}, title = {{Characterizing the sequence and expression evolution of the t-haplotype, a model meiotic driver}}, doi = {10.15479/at:ista:17119}, year = {2024}, } @phdthesis{18477, abstract = {ADAR1 is broadly expressed across various tissues and is vital in regulating pathways associated with innate immune responses. ADAR1 marks double-stranded RNA as "self" through its A-to-I editing activity, effectively repressing autoimmunity and maintaining immune tolerance. This editing process has been detected at millions of sites across the human genome. However, the mechanism underlying ADAR1's substrate selectivity properties remains largely unclear, with much of the current knowledge derived from comparisons to its more extensively studied homolog, ADAR2. By studying ADAR1 in complex with its RNA substrates and applying a combination of biochemical techniques and structural studies using CryoEM, we aim to gain a more comprehensive understanding of the substrate selectivity characteristics of ADAR1. In this thesis, the purification protocol for ADAR1 was successfully optimized, resulting in the first report in the literature to achieve high protein purity and activity. This advancement enabled the investigation of complex formation between ADAR1 and various RNA substrates, leading to the identification of optimal conditions for preparing the cryoEM sample. However, despite comprehensive optimization of the cryo-EM conditions, the resulting data lacked the desired quality, highlighting the need for similar rigorous optimization of the RNA substrates to facilitate structural studies of the ADAR1-RNA complex. The study was complemented by AlphaFold predictions, which provided some insights into this mechanism. Moreover, during this project I established a collaboration with a research group focused on studying ADAR homologs. Notably ADAR homologs were identified in bivalve species, and it was further demonstrated that ADAR and its A-to-I editing activity are upregulated in Pacific oysters during infections with Ostreid herpesvirus-1—a highly infectious virus that leads to significant losses in oyster populations globally. I successfully purified oyster ADAR and prepared in vitro edited RNA for nanopore sequencing—a direct sequencing technology capable of detecting modified nucleotides without the need for reverse transcription. The collaborators initiated optimization of this nanopore-based approach. However, current technological limitations still constrain the reliable detection of modified nucleotides. The project also examined the impact of RNA editing on RNA binding and filament formation by MDA5, a key cytosolic dsRNA sensor that triggers an interferon response. A primary target of ADAR1's editing activity is RNA derived from repetitive elements present in the genome, particularly Alu elements forming double-stranded RNA. When unedited, these RNA sequences are recognized by MDA5. However, the mechanisms by which MDA5 interacts with Alu RNAs, as well as the role of A-to-I editing in influencing this binding, are still not well understood. The interaction between MDA5 and Alu elements, was successfully established. This was achieved through the testing of different RNA variants and the evaluation of filament formation using binding techniques and electron microscopy imaging. This groundwork has set the conditions for further evaluation using CryoEM. Furthermore, the effects of A-to-I editing on the binding properties of MDA5 with Alu RNA were investigated. Given the recent research that has provided new insights into MDA5's interaction with dsRNA, it is essential to revise the experimental setup to integrate these findings before moving forward with the CryoEM sample analysis.}, author = {Kaczmarek, Beata M}, isbn = {978-3-99078-045-9}, issn = {2663-337X}, pages = {124}, publisher = {Institute of Science and Technology Austria}, title = {{Biochemical and structural insights into ADAR1 RNA editing}}, doi = {10.15479/at:ista:18477}, year = {2024}, } @inproceedings{17469, abstract = {Autoencoders are a prominent model in many empirical branches of machine learning and lossy data compression. However, basic theoretical questions remain unanswered even in a shallow two-layer setting. In particular, to what degree does a shallow autoencoder capture the structure of the underlying data distribution? For the prototypical case of the 1-bit compression of sparse Gaussian data, we prove that gradient descent converges to a solution that completely disregards the sparse structure of the input. Namely, the performance of the algorithm is the same as if it was compressing a Gaussian source - with no sparsity. For general data distributions, we give evidence of a phase transition phenomenon in the shape of the gradient descent minimizer, as a function of the data sparsity: below the critical sparsity level, the minimizer is a rotation taken uniformly at random (just like in the compression of non-sparse data); above the critical sparsity, the minimizer is the identity (up to a permutation). Finally, by exploiting a connection with approximate message passing algorithms, we show how to improve upon Gaussian performance for the compression of sparse data: adding a denoising function to a shallow architecture already reduces the loss provably, and a suitable multi-layer decoder leads to a further improvement. We validate our findings on image datasets, such as CIFAR-10 and MNIST.}, author = {Kögler, Kevin and Shevchenko, Aleksandr and Hassani, Hamed and Mondelli, Marco}, booktitle = {Proceedings of the 41st International Conference on Machine Learning}, location = {Vienna, Austria}, pages = {24964--25015}, publisher = {ML Research Press}, title = {{Compression of structured data with autoencoders: Provable benefit of nonlinearities and depth}}, volume = {235}, year = {2024}, } @phdthesis{18101, abstract = {The Retroviridae family consists of two sub-families, the Orthoretrovirinae and the Spumaretrovirinae. The Orthoretroviruses contain important human pathogens, such as the human immunodeficiency virus 1 (HIV-1). They also harbor other retrovirus species which are regularly used as model systems to study the retroviral life cycle. The main structural component of the retroviruses, is the Gag protein and its truncation derivatives occurring during viral maturation. Orthoretroviral Gag assemblies have been extensively studied to understand the interactions that confer stability and morphology to viral particles. The Spumaretrovirinae subfamily represent an early diverging branch of the Retroviridae. Its members, the Foamy viruses (FV), share most of the conventional features found in retroviruses. However, they also possess multiple characteristics that make them unique. In particular, FV Gag does not get extensively cleaved as in orthoretroviruses. Hence, the Gag architecture deviates from the canonical domain arrangement in FV. They also exhibit a peculiar particle morphology, having no apparent immature state and a seemingly icosahedral mature particle. Due to this, many fundamental questions on FV structural assembly mechanisms remain open. To answer these questions, was the main focus of this thesis. Mainly, it is not known how FV assemble their core in a virus particle and what are the important assembly interaction sites within said core. What is the minimum assembly competent domain of FV Gag? Is there a morphological change in the assembly type of FVGag lattices? If so, what is defining these morphological shifts? Finally, it would be interesting to know what is the evolutionary relationship between FV and the rest of the retrotranscribing elements, from a structural point of view? To answer these questions, membrane-enveloped mammalian cell-derived FV virus-like particles (VLPs) were produced. Cryo-electron tomography (cryo-ET) analysis suggested these FV VLPs do not form a canonical retroviral Gag lattice structure, which is in line with earlier observations. To further evaluate FV Gag assembly competence and morphology, the first bacterial cell-derived in vitro VLP assembly system was designed and optimized. Using this system with different truncation variants, the minimum assembly competent domain of FV Gag was found to be the putative CA300-477 domain. Varying VLP morphologies were also observed and strongly suggested residues upstream of CA300-477 play a role in morphology determination. Finally, a combined cryo-electron microscopy (cryoEM) and cryo-ET approach was taken to analyze tubular assemblies from the minimal assembly competent domain. This revealed an unexpectedly unique non-canonical assembly architecture. Three novel lattice stabilizing interfaces were described which proved to be as unique as the lattice arrangement. Comparison to a newly published FV CA core structure revealed the CA-CA interactions in the atypical assembly do not recapitulate what is described for the FV core lattice. However, the new in vitro VLP assembly system obtained in this thesis also provides an exciting opportunity to study still unresolved FV assembly features in a potentially facilitated approach compared to conventional methods. In summary, this work provided a deeper understanding of the basic FV Gag assembly unit, as well as presenting the first FV Gag-derived in vitro VLP assembly system. This system reveals a novel and unique assembly architecture among retroviral in vitro assemblies.}, author = {Porley, Dario J}, isbn = {978-3-99078-041-1}, issn = {2663-337X}, pages = {131}, publisher = {Institute of Science and Technology Austria}, title = {{Structural characterization of spumavirus capsid assemblies}}, doi = {10.15479/at:ista:18101}, year = {2024}, } @phdthesis{17346, abstract = {Acquiring, retaining, and retrieving information over a wide range of timescales are crucial functions of the brain. The successful processing of memories affects many aspects of our lives and enables us and many other organisms to operate in a complex environment and to interact with it. In this context, the hippocampus and functionally connected brain areas, such as the prefrontal cortex, are central and have been subject to intensive research in the past decades. Storage of memories is believed to rely on distributed neural activity within these neural circuits. Additionally, neural memory traces of recent experience are reinstated during periods of rest or sleep. These reactivations are thought to play an outstanding role in the consolidation of memories and potentially facilitate the transfer of information from the hippocampus to cortical areas for long-term storage and integration into existing knowledge. However, there is growing evidence that memory-related neural representations in the hippocampus are not as stable as initially thought and that they change even in the absence of learning. It has been suggested that these changes reflect the accumulation of experience, but the influence of interspersed consolidation periods has not been considered. Previous studies have analyzed consolidation periods by detecting activity that strongly resembled neural activity during the acquisition of memory. Besides being often limited to only non-rapid eye movement (NREM) sleep, the used approaches were not capable of tracking changes in neural representations over extended temporal periods. More fluid representations do not only challenge our understanding of how information is stored, but they also affect the transfer of information between brain areas during the consolidation process. For this thesis, I investigated the evolution of memory-related activity during sleep periods expected to be involved in consolidation in the hippocampus and between the hippocampus and prefrontal cortex. I found that reactivated activity in the hippocampus gradually transformed during prolonged periods of sleep and inactivity. In the beginning, neural activity strongly resembled acquisition activity, whereas, with the progression of time, it became more similar to the subsequent recall activity. NREM periods drove this process, while rapid-eye movement (REM) periods showed a resetting effect. This reactivation drift was due to firing rate changes of a subset of cells and mirrored the representational changes from the acquisition to the recall. A stable subset of cells withstood the drift and maintained their activity. Therefore, my results indicate that memory-related representations undergo spontaneous modifications during consolidation periods and that these changes are predictive of representational drift. Furthermore, I found that the amount of change in the neural activity during subsequent sleep periods was biased by prior behavioral performance. Observed changes in the hippocampus and the prefrontal cortex were synchronized and increased after poor performance, highlighting a potential role in the exchange of information. Low-variance vii periods with distinct, more stable activity from a subset of cells significantly contributed to the heightened synchrony between both areas. Hence, interleaved phases of more stable neural activity could facilitate the information transfer between brain areas. In conclusion, my investigations underline the fluidity of memory-related representations and assign a prominent role to sleep reactivation periods in their evolution. In addition, I identified a potential mechanism of stable activity phases that might facilitate the synchronization across hippocampal-prefrontal activity despite ongoing changes. Reconciling and integrating findings from both spontaneous and behaviorally-related representational changes in functionally related brain areas will help to broaden our understanding of how knowledge is stored, maintained, updated, and transferred between brain areas.}, author = {Bollmann, Lars}, issn = {2663-337X}, keywords = {Memory, Hippocampus, Consolidation}, pages = {103}, publisher = {Institute of Science and Technology Austria}, title = {{Stability and change in the memory system during rest}}, doi = {10.15479/at:ista:17346}, year = {2024}, } @phdthesis{18129, abstract = {State-of-the-art quantum computers, with roughly a thousand qubits, face a crucial technological challenge of scaling up. Spins confined in quantum dots (QDs) are a promising candidate for qubits due to their long coherence, tunability, control, and readout. However, their natural coupling is the short-ranged (∼ 100 nm) exchange interaction, limited to nearest neighbours. Long-ranged (∼ 1 mm) qubit interactions mediated by a photon could be engineered through a coherent spin-photon coupling. Achieving a strong coupling to a photon is inherently challenging in QDs due to the small dipole moment of the confined charge. However, the potential of high-impedance resonators to compensate for this has gained significant attention in the past decade. Nevertheless, previous QD circuit quantum electrodynamics implementations have not exceeded the impedance of ∼ 3.8 kΩ, leaving opportunities for significant improvement. The large kinetic inductance of granular aluminium (grAl) could provide an order-of-magnitude enhancement. However, fully exploiting the potential of disordered or granular superconductors is challenging as their impedances close to the superconductor-to-insulator transition are difficult to control reproducibly. We report on the realization of a wireless ohmmeter which allows in situ resistance measurements during film deposition and, therefore, indirect control of the kinetic inductance of grAl films. This allows us to reproducibly fabricate resonators with characteristic impedance exceeding the resistance quantum, even reaching 22.3 kW, due to the large sheet kinetic inductance of up to 3 nH □−1 . By integrating an 8 kW resonator with a germanium double QD, we demonstrate a strong charge-photon coupling with the highest rate reported, 566 MHz. The demonstrated method and grAl properties make these resonators suitable for boosting the spin-photon coupling strength, a crucial requirement for fast, high-fidelity, long-distance two-qubit gates. }, author = {Janik, Marian}, issn = {2663-337X}, pages = {164}, publisher = {Institute of Science and Technology Austria}, title = {{Strong charge-photon coupling in Germanium enabled by granular aluminium superinductors}}, doi = {10.15479/at:ista:18129}, year = {2024}, }