@article{20187, abstract = {Very long-chain fatty acids (VLCFAs), being constituents of different types of lipids, are critical factors in plant development, presumably due to their impact on the endomembrane system. The VLCFAs are synthesized in the endoplasmic reticulum by a heterotetrameric enzymatic complex including β-ketoacyl CoA reductase 1 (KCR1), whose mutant is lethal. Here, we describe the ectopic shoot meristems (esm) mutant, a viable kcr1 allele presumably affecting surface properties of the KCR1 protein. This kcr1-2 mutant shows reduced fatty acyl elongation that impacts VLCFAs. The kcr1-2 plants show severe defects during different stages of development, which all correlate with defects in polar localization and subcellular trafficking of PIN auxin transporters and resulting asymmetric auxin distribution. Detailed analysis of KCR1 expression and patterning defects in kcr1-2 suggests that KCR1 plays a role in delineating boundaries around meristematic and specialized differentiating tissues, including root and shoot meristems, initiating lateral roots, lateral root primordia, and trichomes. In these contexts, KCR1-produced VLCFAs may act in a non-cell-autonomous manner. Viable kcr1-2 represents a useful tool to study VLCFA roles in plant development and highlights VLCFAs as critical developmental factors at the interface of cell polarity and tissue development.}, author = {Babic, David and Abualia, Rashed and Fiedler, Lukas and Qi, Linlin and Tellier, Frédérique and Smoljan, Adrijana and Rakusova, Hana and Valošek, Petr and Han, Huibin and Benková, Eva and Faure, Jean Denis and Friml, Jiří}, issn = {1365-313X}, journal = {Plant Journal}, number = {3}, publisher = {Wiley}, title = {{Biosynthesis of very long-chain fatty acids is required for Arabidopsis auxin-mediated embryonic and post-embryonic development}}, doi = {10.1111/tpj.70396}, volume = {123}, year = {2025}, } @phdthesis{20362, abstract = {Arabidopsis embryo has long served as a model system for understanding how plant body patterning and axis establishment arise. Previous work established that auxin flows, directed by polar transport and reinforced by local biosynthesis, underpin axes formation, while recent studies suggest that maternal tissue, membrane composition and surface receptor signaling add further regulatory layers. In this thesis, we dissect the layers of auxin-mediated processes which guide plant embryonic and post-embryonic development. We review methodological advances that allow the embryo to be imaged and studied. These tools offer exciting venues for future addressing of outstanding research questions. Furthermore, we dissect the contribution of maternal auxin for early embryo patterning. Targeted auxin scavenging in the integuments lowers the hormone reaching the proembryo, destabilizes PIN7 in the suspensor and weakens the initial apical maximum. The resulting deficit dampens downstream embryonic auxin biosynthesis, as well as auxin flow towards the root pole, indicating that maternally provided auxin controls auxin flows in the embryos on a larger scale than previously understood. Intriguingly, we uncover a prolonged maternal effect mediated by the receptor-like kinases TMK1 and TMK4. Reciprocal crosses and transgene analyses hint that these genes could be preferentially maternally-derived in both embryos and seedlings. Loss of maternal TMK activity arrests embryos, affects seedling development and persists into adult growth, suggesting that maternal effect could shape development beyond fertilization and early embryogenesis. Finally, we describe the role of very-long-chain fatty acids and membrane composition on PIN proteins and their auxin-induced relocalization. Disruption in this lipid metabolism triggers widespread patterning errors. Together, these results reposition early plant patterning as a hierarchical cascade that begins with maternal auxin, gets amplified by auxin transport and biosynthesis, potentially modulated by maternal receptor signaling and could ultimately be gated by the biophysical properties of the membrane. By integrating auxin supply, genetic imprinting and lipid-dependent trafficking into a single narrative, this thesis helps shifting the approach of viewing the embryo as an autonomous unit to regarding it as a dynamic system, continuously sculpted by its maternal context.}, author = {Babic, David}, issn = {2663-337X}, pages = {116}, publisher = {Institute of Science and Technology Austria}, title = {{Mechanisms of auxin-mediated early embryogenesis in Arabidopsis thaliana}}, doi = {10.15479/AT-ISTA-20362}, year = {2025}, } @unpublished{21427, abstract = {While tumor malignancy has been extensively studied under the prism of genetic and epigenetic heterogeneity, tumor cell states also critically depend on reciprocal interactions with the microenvironment. This raises the hitherto untested possibility that heterogeneity of the untransformed tumor stroma can actively fuel malignant progression. As biological heterogeneity is inherently difficult to control, we adopted a reductionist approach and let tumor cells invade micro-engineered environments harboring obstacles with precision-controlled geometry. We find that not only the presence of obstacles, but more surprisingly their spatial disorder, causes a drastic shift from a collective to a single-cell mode of invasion – comparable in strength to cadherin loss. Combining live-imaging and perturbation experiments with minimal biophysical modeling, we demonstrate that cell detachments result both from local geometrical constraints and a global integration of spatial disorder over time. We show that different types of microenvironments map onto different universality classes of invasion dynamics - homogeneous substrates follow Kardar–Parisi–Zhang (KPZ) scaling, while disordered ones exhibit exponents consistent with KPZ with quenched disorder (KPZq). Our findings highlight generic physical principles for how the mode of cancer cell invasion depends on environmental heterogeneity, with potential implications to understand tumor evolution in vivo.}, author = {Dunajova, Zuzana and Tasciyan, Saren and Majek, Juraj and Merrin, Jack and Sahai, Erik and Sixt, Michael K and Hannezo, Edouard B}, booktitle = {bioRxiv}, title = {{Substrate heterogeneity promotes cancer cell dissemination through interface roughening}}, doi = {10.1101/2025.05.20.655037}, year = {2025}, } @phdthesis{20393, abstract = {During vertebrate development, the morphogen Sonic Hedgehog (Shh) plays a key role in patterning the tissues of the extending body. This requires precise coordination between the size of the source of Shh production and the embryo body. Shh is produced by the notochord, as well as floorplate of the neural tube. In this thesis, we investigated how the sizes of the notochord and floorplate are regulated and how they impact the developing mouse embryo. To measure the size and shape of the developing mouse notochord and the cell behaviours within this organ, we performed high-resolution measurements in cleared embryos between E8.5 and E10.5 of development. Strikingly, our data revealed that the notochord maintains a constant cross-sectional area throughout the trunk over time as it lengthens by cell proliferation. We show that this is achieved by a feedback between the notochord cell proliferation rate and both the cross-sectional area of the trunk notochord and elongation of the embryo. Using live imaging, we further show that notochord cells undergo posteriorly directed active cell migration and that this cell migration contributes to maintaining the structural integrity of the notochord. Our results show a complex coordination of cell behaviours that allow for robust notochord morphogenesis during mouse development and the maintenance of a defined Shh source size. Shh produced by the notochord induces a secondary Shh-producing domain, the floor plate. How the notochord and floor plate contribute to the formation of the Shh gradient in the neural tube and to the regulation of the floor plate size is poorly understood. Our theoretical and experimental analysis indicates that only the initial levels of notochord-derived Shh influence the size of the floor plate, and the floor plate is insensitive to its own production of Shh. After its initial establishment, the floor plate expands by tissue growth, leading to scaling of the Shh gradient amplitude with tissue size. Altogether, we show that the mechanisms regulating the sizes of the notochord and floorplate in mouse development help to achieve size robustness of these tissues. Our data demonstrate that the initial establishment of the Shh gradient in the neural tube is strongly dependent on the notochord and suggest that the robustness of notochord morphogenesis is essential for pattern formation. Later in development, the floor plate size and the Shh gradient are coupled to neural tube growth. Our findings provide new insights on how pattern formation, tissue growth and morphogenesis interact to ensure reproducible developmental outcomes. }, author = {Kishi, Kasumi}, issn = {2663-337X}, pages = {102}, publisher = {Institute of Science and Technology Austria}, title = {{Regulation of notochord and floor plate size during mouse development}}, doi = {10.15479/AT-ISTA-20393}, year = {2025}, } @phdthesis{20364, abstract = {Auxin is a central regulator of plant growth and development, orchestrating processes from embryogenesis to senescence through a complex network of biosynthesis, transport, and signaling pathways. In Arabidopsis thaliana, canonical nuclear auxin signaling is mediated by the TIR1/AFB receptors, which promote degradation of AUX/IAA repressors to enable ARF transcription factors-driven transcriptional reprogramming. Parallel to this well-established nuclear mechanism, auxin also triggers rapid, non-transcriptional responses. In particular, at the cell surface, most notably through ABP1 and TMKs. Despite decades of research, the integration of these pathways and their links to fundamental cellular processes remains not fully understood. In this study, we uncover a new role for TIR1/AFB signaling in rapidly activating macroautophagy during developmental reprogramming. Auxin induces autophagy via TIR1/AFB adenylate cyclase activity, linking auxin perception to cAMP production and transcriptional activation of autophagy machinery. Autophagy-deficient mutants exhibit delayed meristem differentiation, impaired lateral root progression, and reduced shoot organogenesis, demonstrating its necessity for timely developmental transitions. We also revisit the ABP1-TMK1 cell-surface auxin perception complex, providing biochemical and mass photometry evidence for auxin-dependent complex formation. We show that ABP1-TMK1 triggers an ultrafast global phosphorylation response targeting ~1000 proteins, including the auxin transporter PIN2, and Rho-of-plants (ROP) GTPases regulators. This pathway directly regulates PIN2 phosphorylation, for gravitropism and auxin dependent ROP activation. These findings establish an integrated framework in which nuclear TIR1/AFB signaling and cell-surface ABP1-TMK signaling act in parallel to synchronize gene expression, cytoplasmic remodelling, and transporter regulation, thereby ensuring robust developmental plasticity in Arabidopsis thaliana. }, author = {Giannini, Caterina}, issn = {2663-337X}, keywords = {Auxin Signaling, Plant Development}, pages = {151}, publisher = {Institute of Science and Technology Austria}, title = {{Nuclear and cell surface auxin signaling in A. thaliana developmental transitions}}, doi = {10.15479/AT-ISTA-20364}, year = {2025}, } @unpublished{19399, abstract = {Phytohormone auxin and its directional transport mediate much of the remarkably plastic development of higher plants. Positive feedback between auxin signaling and transport is a key prerequisite for (i) self-organizing processes including vascular tissue formation and (ii) directional growth responses such as gravitropism. Here we identify a mechanism, by which auxin signaling directly targets PIN auxin transporters. Via the cell-surface ABP1-TMK1 receptor module, auxin rapidly induces phosphorylation and thus stabilization of PIN2. Following gravistimulation, initial auxin asymmetry activates autophosphorylation of the TMK1 kinase. This induces TMK1 interaction with and phosphorylation of PIN2, stabilizing PIN2 at the lower root side, thus reinforcing asymmetric auxin flow for root bending. Upstream of TMK1 in this regulation, ABP1 acts redundantly with the root-expressed ABP1-LIKE auxin receptor ABL3. Such positive feedback between cell-surface auxin signaling and PIN-mediated polar auxin transport is fundamental for robust root gravitropism and presumably also for other self-organizing developmental phenomena.}, author = {Rodriguez Solovey, Lesia and Fiedler, Lukas and Zou, Minxia and Giannini, Caterina and Monzer, Aline and Vladimirtsev, Dmitrii and Randuch, Marek and Yu, Yongfan and Gelová, Zuzana and Verstraeten, Inge and Hajny, Jakub and Chen, Meng and Tan, Shutang and Hörmayer, Lukas and Li, Lanxin and Marques-Bueno, Maria Mar and Quddoos, Zainab and Molnar, Gergely and Xu, Tongda and Kulich, Ivan and Jaillais, Yvon and Friml, Jiří}, booktitle = {bioRxiv}, title = {{ABP1/ABL3-TMK1 cell-surface auxin signaling directly targets PIN2-mediated auxin fluxes for root gravitropism}}, doi = {10.1101/2022.11.30.518503}, year = {2025}, } @phdthesis{19763, abstract = {Pattern formation in developing organs is controlled by morphogens. These signalling molecules form concentration gradients across tissues, thereby providing positional information that instructs the pattern of cell differentiation. Morphogen gradients are highly dynamic in space and time. Many factors such as morphogen production, spreading, degradation, cellular rearrangements and others could contribute to changes in the gradient shape, yet how the spatiotemporal signalling dynamics arise in many systems is still unclear. We studied the dynamics of morphogen signalling and tissue patterning in the developing vertebrate neural tube. In this system, neural crest, roof plate and distinct dorsal progenitor subtypes are specified in a spatially and temporally ordered manner in response to dorsal-toventral gradients of BMP and WNT signalling activity. How the BMP and WNT gradients are established and interpreted to ensure ordered cell specification is poorly understood. To address this question, we developed a 2D embryonic stem cell differentiation system that captures key features of dorsal neural tube development. In this system, differentiated colonies display remarkable self-organised pattern formation in response to uniformly applied BMP ligand. We established a method of differentiating the colonies using microfabricated stencils, which allowed us to control the initial size and shape of colonies without confining cell migration and colony growth. This led to highly reproducible pattern formation that facilitates quantification. Using this approach, we observed striking two-phase temporal dynamics of BMP signalling in our colonies: a BMP gradient rapidly forms from the periphery to the centre of colonies, subsequently disappears and is re-established again in the second phase. By combining our quantitative data with a data-driven theoretical model, we uncovered a temporal relay mechanism that underlies this biphasic BMP signalling dynamics. The first signalling phase is controlled by fast tissue-autonomous negative feedback that restricts the duration of the initial response to BMP. The early BMP activity gradient moreover controls the spatial organisation of the cell type pattern: the absence of a first phase results in disordered cell type pattern. The second phase is controlled by slow positive regulation of BMP signalling by the transcription factor LMX1A, a key regulator of roof plate identity. WNT promotes the second phase of BMP signalling via positive feedback on LMX1A. Altogether, the mechanism that we uncovered ensures the coupling of sequential developmental events, making pattern formation spatially and temporally organised. Furthermore, this mechanism allows the BMP signalling pathway to be reused in different contexts – first for the establishment of the neural plate border, and subsequently for dorsal neural progenitor patterning. Our study supports a general developmental principle in which multiple morphogens interact with transcriptional networks resulting in complex spatiotemporal signalling dynamics that ultimately drive organised pattern formation.}, author = {Rus, Stefanie}, issn = {2663-337X}, pages = {129}, publisher = {Institute of Science and Technology Austria}, title = {{Dynamics of morphogen signalling and cell fate decisions in the dorsal neural tube}}, doi = {10.15479/AT-ISTA-19763}, year = {2025}, } @article{18807, abstract = {Developing tissues interpret dynamic changes in morphogen activity to generate cell type diversity. To quantitatively study bone morphogenetic protein (BMP) signaling dynamics in the mouse neural tube, we developed an embryonic stem cell differentiation system tailored for growing tissues. Differentiating cells form striking self-organized patterns of dorsal neural tube cell types driven by sequential phases of BMP signaling that are observed both in vitro and in vivo. Data-driven biophysical modeling showed that these dynamics result from coupling fast negative feedback with slow positive regulation of signaling by the specification of an endogenous BMP source. Thus, in contrast to relays that propagate morphogen signaling in space, we identify a BMP signaling relay that operates in time. This mechanism allows for a rapid initial concentration-sensitive response that is robustly terminated, thereby regulating balanced sequential cell type generation. Our study provides an experimental and theoretical framework to understand how signaling dynamics are exploited in developing tissues.}, author = {Rus, Stefanie and Brückner, David and Minchington, Thomas and Greunz, Martina and Merrin, Jack and Hannezo, Edouard B and Kicheva, Anna}, issn = {1534-5807}, journal = {Developmental Cell}, number = {4}, pages = {567--580}, publisher = {Cell Press}, title = {{Self-organized pattern formation in the developing mouse neural tube by a temporal relay of BMP signaling}}, doi = {10.1016/j.devcel.2024.10.024}, volume = {60}, year = {2025}, } @phdthesis{20694, abstract = {Understanding the mechanisms underlying speciation is a central aim of evolutionary biology. A persistent challenge in the field is to identify loci that contribute to reproductive isolation, while disentangling signals of selection from demography, linkage and intrinsic genomic features. Traditional population genomic approaches that rely on site-based statistics in arbitrary fixed windows face inherent limitations, as they conflate historical and contemporary processes of divergence and overlook haplotype structure. Recent advances in whole-genome sequencing and methods to infer ancestral recombination graphs (ARGs) now offer the opportunity to study genealogical relationships explicitly, revealing how lineages coalesce and recombine through time. By directly analysing haplotype clustering by species or phenotype and their patterns of coalescence, ARG-based methods show promise for diagnosing sweeps, identifying barrier loci maintained under divergent selection amid gene flow, and tracing their evolutionary history. In this thesis, I explore the utility of genealogical approaches for studying species divergence. In chapter 2, I propose a conceptual framework for defining haplotype blocks through the structure of the ARG, using simulations and empirical data to highlight how genealogical processes generate rich and often overlooked haplotypic patterns. In chapter 3, I examine the genomic basis of a key evolutionary innovation in marine snails Littorina. These snails offer a unique opportunity to study an innovation because they include a very recent transition from egg-laying to live bearing, yet snails with the different reproductive modes are not reciprocally monophyletic. I exploited this by using topology clustering in ARG-derived local genealogical trees to pinpoint narrow genomic regions or haplotype blocks that carry swept alleles, thus revealing that the transition from egg-laying to live-bearing involves multiple, live-bearer-specific sweeps. Chapter 4 establishes a population-scale, phased genomic resource for Antirrhinum majus, using cost-effective haplotagging, then optimizes imputation from low-coverage data against high-accuracy KASP sequencing to maximize sequence completeness with modest accuracy trade-offs against a traditional short-read sequence pipeline. A hybrid phasing strategy combines molecular phasing with statistical phasing to generate phased whole genome sequences of 1084 Antirrhinum individuals at a fraction of long-read sequencing costs. In chapter 5, I analyse hybridising populations from two replicate hybrid zones to find a parallel genetic basis of flower colour, amidst the noise in genomic differentiation landscape driven by variation in demographic history. While outlier genome scans of FST failed to dissect the causes of differentiation, ARG-based topology clustering revealed a reuse of colour associated haplotypes across hybrid zones. In addition to the biological insight, this chapter also presents a comparison of the latest ARG inference tools, showing that signals of Abstract viii topological clustering qualitatively agree between methods, despite differences in the tree sequences. Next, in chapter 6, by leveraging ~1000 individuals in one of the hybrid zones, I integrated genome-wide association studies of floral pigmentation with genealogical inference, to test for additional colour loci, and confirm the effect of previously described loci. This work demonstrates that flower colour variation is driven by a small number of large effect loci, while also hinting at the presence of a new candidate regulatory factor. Finally in chapter 7, in a preliminary analysis, I begin to dissect the genomic island of speciation around Rosea/Eluta to understand its evolutionary origins. My results show that it consists of 5 highly divergent loci, each of which is associated with flower colour. Using patterns of coalescence in genealogical trees, I find evidence of staggered selective sweeps and a persistent localized barrier to gene flow within an otherwise permeable genome. Together, these chapters add to the increasing pool of studies using genealogical approaches to complement and extend site-based statistics to use haplotype structures in speciation research. By tracking haplotypes directly and connecting genealogical clustering to population processes, ARG-based inference promises to provide new insights into how local selective pressures, demographic history, and long-term barriers interact to shape the genomic architecture of divergence. By underscoring the value of ARGs in revealing the finescale origins and maintenance of biodiversity, this thesis presents cautious optimism about the benefits of using genealogical inference to learn more than what site-based statistics could tell us.}, author = {Pal, Arka}, issn = {2663-337X}, pages = {268}, publisher = {Institute of Science and Technology Austria}, title = {{Using genealogies to study the genomic basis of species divergence}}, doi = {10.15479/AT-ISTA-20694}, year = {2025}, } @article{20190, abstract = {A major goal of speciation research is identifying loci that underpin barriers to gene flow. Population genomics takes a ‘bottom-up’ approach, scanning the genome for molecular signatures of processes that drive or maintain divergence. However, interpreting the ‘genomic landscape’ of speciation is complicated, because genome scans conflate multiple processes, most of which are not informative about gene flow. However, studying replicated population contrasts, including multiple incidences of secondary contact, can strengthen inferences. In this paper, we use linked-read sequencing (haplotagging), FST scans and genealogical methods to characterise the genomic landscape associated with replicate hybrid zone formation. We studied two flower colour varieties of the common snapdragon, Antirrhinum majus subspecies majus, that form secondary hybrid zones in multiple independent valleys in the Pyrenees. Consistent with past work, we found very low differentiation at one well-studied zone (Planoles). However, at a second zone (Avellanet), we found stronger differentiation and greater heterogeneity, which we argue is due to differences in the amount of introgression following secondary contact. Topology weighting of genealogical trees identified loci where haplotype diversity was associated with the two snapdragon varieties. Two of the strongest associations were at previously identified flower colour loci: Flavia, that affects yellow pigmentation, and Rosea/Eluta, two linked loci that affect magenta pigmentation. Preliminary analysis of coalescence times provides additional evidence for selective sweeps at these loci and barriers to gene flow. Our study highlights the impact of demographic history on the differentiation landscape, emphasising the need to distinguish between historical divergence and recent introgression.}, author = {Pal, Arka and Shipilina, Daria and Le Moan, Alan and Mcnairn, Adrian J. and Grenier, Jennifer K. and Kucka, Marek and Coop, Graham and Chan, Yingguang Frank and Barton, Nicholas H and Field, David and Stankowski, Sean}, issn = {1365-294X}, journal = {Molecular Ecology}, number = {22}, publisher = {Wiley}, title = {{Genealogical analysis of replicate flower colour hybrid zones in Antirrhinum}}, doi = {10.1111/mec.70067}, volume = {34}, year = {2025}, } @phdthesis{19271, abstract = {The medial habenula (MHb) is implicated in regulating emotional responses to aversive events. Studies in zebrafish have identified a remarkable morphological left-right asymmetry in the dorsal habenula (zebrafish equivalent of mammalian MHb)-to-interpeduncular nucleus (IPN) pathway and its left-side specific role in modulating fear responses. However, there is little evidence for structural or functional lateralization in the mammalian MHb-IPN pathway. Here, I investigated the synaptic properties of the left and right MHb afferents to the IPN in mice and addressed whether these synaptic connections selectively influence the expression of conditioned fear in mice. My findings reveal that each individual IPN neuron receives inputs from both left and right MHb. Electrophysiological recordings from the same postsynaptic IPN neurons demonstrate that the left MHb-originating synapses exhibit lower release probability and higher 𝛾-aminobutyric acid type B receptor (GABABR)-mediated potentiation compared to the right MHb-originating synapses. Interestingly, chemogenetic inhibition of cholinergic neurons in the left but not the right MHb significantly attenuated cue-dependent fear recall. Furthermore, conditional deletion of GABABR in the left MHb interfered with the recall of cued fear memory, whereas that in the right MHb neurons spared fear memory expression. Collectively, I demonstrate a functional asymmetry of the MHb in mice, revealing a predominant role for GABABR-mediated signaling in the left MHb-IPN pathway in the modulation of fear memories. These findings suggest that lateralized pathways could represent a fundamental principle in the neural regulation of emotion across species.}, author = {Önal, Hüseyin C}, issn = {2663-337X}, publisher = {Institute of Science and Technology Austria}, title = {{Asymmetrical modulation of fear expression via GABAB receptors in the mouse medial habenula}}, doi = {10.15479/AT-ISTA-19271}, year = {2025}, } @phdthesis{19478, abstract = {Auxin is one of the most important phytohormones, playing a crucial role in plant growth and development. Genetic studies in Arabidopsis have identified core components of auxin signalling and established the classical transcriptional regulation mechanism. This classical paradigm recognizes TRANSPORT INHIBITOR RESPONSE 1 (TIR1)/AUXIN SIGNALLING F-BOX (AFB) proteins as auxin receptors possessing E3 ubiquitin ligase activity. Auxin functions as a "molecular glue" enhancing TIR1/AFB-Aux/IAA (AUXIN/INDOLE-3-ACETIC ACID) inhibitor interactions, leading to Aux/IAAs’ ubiquitination and degradation. This process releases AUXIN RESPONSE FACTORs (ARFs) from repression, enabling transcriptional activation of auxin-responsive genes. Notably, the discovery of adenylate cyclase (AC) activity in TIR1/AFBs, catalysing cAMP production from ATP, presents novel transcriptional regulatory possibilities, though cAMP's mechanistic role in auxin signalling remains unclear. Beyond transcriptional regulation (slow response) a non transcriptional pathway (rapid response) mediated by AFB1 also exists, as auxin triggers rapid root growth inhibition within 30 seconds. How TIR1 and AFB1, with their distinct subcellular localizations, differentially regulate transcriptional and non-transcriptional processes remains unclear. To address these two key questions, this study conducted the following investigations. TIR1 and AFB1 display distinct subcellular localization patterns, with AFB1 specifically mediating auxin-induced rapid root growth inhibition. To examine whether subcellular localization differences contribute to functional divergence between TIR1 and AFB1, we utilized microfluidic systems combined with genetic complementation assays. Nuclear export signal-fused TIR1 failed to rescue the rapid response defects observed in afb1 mutants. In contrast, nuclear localization signal-fused AFB1 exhibited dual functional deficiencies: it neither restored the rapid response impairment in afb1 mutants nor compensated for transcriptional regulation defects in tir1 mutants which limitation stemmed from AFB1's intrinsic incapacity to assemble functional SCF complexes. These results collectively establish that the functional specialization of TIR1 and AFB1 is governed by both their subcellular compartmentalization and inherent biochemical characteristics. The mechanism by which cAMP derived from the auxin receptor TIR1/AFB modulates transcriptional regulation remains undefined. To address this, we genetically dissociated TIR1 AC activity from its E3 ubiquitin ligase function to investigate cAMP's signalling role. AC inactivating mutations preserved TIR1-mediated Aux/IAAs’ degradation but abolished ARF i dependent transcriptional activation. tir1afb2 mutants expressing AC-deficient TIR1 variants failed to restore transcriptional responses. Crucially, compartmentalized cAMP accumulation within Aux/IAA - ARF complexes vicinity triggered auxin signalling independently of both auxin perception and Aux/IAAs’ degradation, confirming its secondary messenger identity. Further mechanistic analysis demonstrated cAMP directly interact with ARF. cAMP directly promotes ARF dissociation from Aux/IAA repressors and enhances ARF homodimerization. Genetic complementation assays revealed that ARF mutants lacking conserved cAMP-binding motifs exhibited partial functional recovery, providing direct evidence for cAMP-ARF physical interaction during signal propagation. These findings collectively challenge the established model of auxin signalling and define a novel TIR1-cAMP-ARF regulatory axis, substantially advancing our comprehension of auxin-mediated physiological regulation. }, author = {Chen, Huihuang}, issn = {2663-337X}, pages = {118}, publisher = {Institute of Science and Technology Austria}, title = {{The cAMP second messenger in auxin signalling}}, doi = {10.15479/AT-ISTA-19478}, year = {2025}, } @phdthesis{20467, abstract = {Prostaglandins play a central role in the inflammatory response. Prostaglandins are synthesized in a rate-limiting manner by the enzyme cyclooxygenase (Cox), which exists in two isoforms: the Cox-2 and Cox-1 isoforms. Traditionally, Cox-1 is thought to mediate homeostatic functions due to its constitutive expression due to its constitutive expression, whereas Cox-2 is temporary induced in response to inflammatory stimuli and is associated with the inflammatory response. Interestingly, in the central nervous system, Cox-1 is predominantly expressed in microglia. As the resident immune cells of the brain, microglia play a key role in inflammation. Cox-1 expressing microglia are increased in neurodegenerative diseases, and both global knockout of Cox-1 and selective Cox-1 inhibition has been shown to ameliorate inflammatory effects in neurodegenerative diseases. However, direct evidence elucidating how microglial Cox-1 expression influences their response to insults remains limited and warrants further investigation. First, I utilize the optic nerve crush (ONC) model to study the microglial response to injury and inflammation, confirming a reactive microglial response in both the crushed and uninjured contralateral retina using CD68 expression and morphological analysis. While previous studies reported retinal ganglion cell (RGC) loss in the contralateral eye, I did not observe this, suggesting that unilateral ONC induces microglial activation in the contralateral eye without RGC degeneration. To explore the role of Cox-1 in this response, I generated a mouse model with microglia-specific deletion of Cox-1. After ONC, these mice exhibited an attenuated microglial response in both the ipsilateral and contralateral retinas, as well as in visual processing areas of the brain. Notably, Cox-2 does not show any expression after ONC in both wild-type and Cox-1 knockout mice Additionally, Cox-1KO mice show a reduction in phagocytic cups and a reduction in microglia proliferation after ONC. However, due to reproducibility problems of the observed phenotype, even after the generation of two new mouse models, I cannot draw any definitive conclusions based on the data. Next, due to the ambiguity of the mouse model, I explored an alternative route to investigate functional consequences of Cox-1 in microglia. I generated a COX1 knock-out human induced pluripotent cell (hIPSC) line using CRISPR/Cas9. These cells are were differentiated in microglia-like cells and integrated in dissociated retinal organoids according to protocols developed in our lab. In response to LPS and POLY(I:C) stimulation, the release and expression of cytokines and chemokines was decreased in COX1 knockout cells. However, the prostaglandin PGE2 was not increased in microglia-like cells in both control and COX1 knockout cells. Overall, the results from the COX1 knock-out microglia-like cells suggests that COX1 expression in microglia play a role in neuroinflammation by affecting the release of inflammatory cytokines and chemokines. Hereby further challenging that the dogma of the Cox-1 isoform involved in housekeeping functions and the Cox-2 isoform involved in inflammation applies for the central nervous system. }, author = {Miteva, Florianne E}, issn = {2663-337X}, pages = {99}, publisher = {Institute of Science and Technology Austria}, title = {{The role of cyclooxygenase 1 on microglial response to inflammatory stressors}}, doi = {10.15479/AT-ISTA-20467}, year = {2025}, } @article{19566, abstract = {Purpose: Optic nerve crush (ONC) is a model for studying optic nerve trauma. Unilateral ONC induces massive retinal ganglion cell (RGC) degeneration in the affected eye, leading to vision loss within a month. A common assumption has been that the non-injured contralateral eye is unaffected due to the minimal retino-retinal projections of the RGCs at the chiasm. Yet, recently, microglia, the brain-resident macrophages, have shown a responsive phenotype in the contralateral eye after ONC. Whether RGC loss accompanies this phenotype is still controversial. Methods: Using the available RGCode algorithm and developing our own RGC-Quant deep-learning-based tool, we quantify RGC's total number and density across the entire retina after ONC. Results: We confirm a short-term microglia response in the contralateral eye after ONC, but this did not affect the microglia number. Furthermore, we cannot confirm the previously reported RGC loss between naïve and contralateral retinas 5 weeks after ONC induction across the commonly used Cx3cr1creERT2 and C57BL6/J mouse models. Neither sex nor the direct comparison of the RGC markers Brn3a and RBPMS, with Brn3a co-labeling, on average, 89% of the RBPMS+-cells, explained this discrepancy, suggesting that the early microglia-responsive phenotype does not have immediate consequences on the RGC number. Conclusions: Our results corroborate that unilateral optic nerve injury elicits a microglial response in the uninjured contralateral eye but without RGC loss. Therefore, the contralateral eye should be treated separately and not as an ONC control.}, author = {Schoot Uiterkamp, Florianne E and Maes, Margaret E and Alamalhoda, Mohammad and Firoozi, Arsalan and Colombo, Gloria and Siegert, Sandra}, issn = {1552-5783}, journal = {Investigative Ophthalmology & Visual Science}, number = {3}, publisher = {Association for Research in Vision and Ophthalmology}, title = {{Optic nerve crush does not induce retinal ganglion cell loss in the contralateral eye}}, doi = {10.1167/iovs.66.3.49}, volume = {66}, year = {2025}, } @article{19421, abstract = {The phytohormone auxin (Aux) is a principal endogenous developmental signal in plants. It mediates transcriptional reprogramming by a well-established canonical signalling mechanism. TIR1/AFB auxin receptors are F-box subunits of an ubiquitin ligase complex; after auxin perception, they associate with Aux/IAA transcriptional repressors and ubiquitinate them for degradation, thus enabling the activation of auxin response factor (ARF) transcription factors1,2,3. Here we revise this paradigm by showing that without TIR1 adenylate cyclase (AC) activity4, auxin-induced degradation of Aux/IAAs is not sufficient to mediate the transcriptional auxin response. Abolishing the TIR1 AC activity does not affect auxin-induced degradation of Aux/IAAs but renders TIR1 non-functional in mediating transcriptional reprogramming and auxin-regulated development, including shoot, root, root hair growth and lateral root formation. Transgenic plants show that local cAMP production in the vicinity of the Aux/IAA–ARF complex by unrelated AC enzymes bypasses the need for auxin perception and is sufficient to induce ARF-mediated transcription. These discoveries revise the canonical model of auxin signalling and establish TIR1/AFB-produced cAMP as a second messenger essential for transcriptional reprograming.}, author = {Chen, Huihuang and Qi, Linlin and Zou, Minxia and Lu, Mengting and Kwiatkowski, M and Pei, Yuanrong and Jaworski, K and Friml, Jiří}, issn = {1476-4687}, journal = {Nature}, pages = {1011--1016}, publisher = {Springer Nature}, title = {{TIR1-produced cAMP as a second messenger in transcriptional auxin signalling}}, doi = {10.1038/s41586-025-08669-w}, volume = {640}, year = {2025}, } @phdthesis{20415, abstract = {Well-defined powder precursors are important for the rational design of crystalline inorganic materials with controlled defects and functional properties. Such powders can be prepared from dried nanoparticles synthesized using a surfactant-assisted/colloidal method that provides precise control over size, shape, composition, and surface chemistry of nanoparticles. These tailored nanoparticles further act as building blocks for three-dimensional assemblies, exhibiting collective nanoscale properties and enabling tailored microstructures of crystalline inorganic materials. However, the programmable engineering of such materials using nanoparticle-based precursors remains limited, due to a lack of fundamental understanding of both the synthesis of nanoparticles and their conversion into macroscopic solids. Furthermore, the formation of three-dimensional assemblies often requires multi-step post-synthetic treatments, which can limit reproducibility and the production of the bulk inorganic materials. In this thesis, I demonstrate a key synthetic parameter governing the formation of desired nanostructures, especially metal/semiconductor core–shell nanoparticles which are potential precursors for engineering composition, crystal structure, and defects of final materials. Then, I investigate their transformative pathways to unveil structural and chemical changes that occur during heat treatment of the nanoparticle-based precursors. Building on understanding of atomic-scale interactions during nanoparticle-based precursor formation, I further demonstrate the possibility of synthesizing the nanoparticle assembly directly in the nanoparticle synthesis reaction and rationalize the process of in situ self-assembly of nanoparticles. Finally, the impact of NP-based precursors on the formation and properties of crystalline inorganic materials is evaluated, demonstrating the potential of employing NP-based precursors for the rational design of functional inorganic materials. }, author = {Lee, Seungho}, issn = {2663-337X}, pages = {144}, publisher = {Institute of Science and Technology Austria}, title = {{Nanoparticle-based precursors toward advanced crystalline inorganic solids}}, doi = {10.15479/AT-ISTA-20415}, year = {2025}, } @article{22026, abstract = {We address two pressing questions in the theory of the Korteweg–de Vries (KdV) equation. First, we show the uniqueness of solutions to KdV that are merely bounded, without any further decay, regularity, periodicity, or almost periodicity assumptions. The second question, emphasized by Deift, regards whether almost periodic initial data leads to almost periodic solutions to KdV. Building on the new observation that this is false for the Airy equation, we construct an example of almost periodic initial data whose KdV evolution remains bounded, but fails to be almost periodic at a later time. Our uniqueness result ensures that the solution constructed is the unique development of this initial data.}, author = {Chapouto, Andreia and Killip, Rowan and Visan, Monica}, issn = {0012-7094}, journal = {Duke Mathematical Journal}, keywords = {Almost-periodic solutions, Korteweg–de Vries, unconditional uniqueness}, number = {7}, pages = {1227--1267}, publisher = {Duke University Press}, title = {{Bounded solutions of KdV: Uniqueness and the loss of almost periodicity}}, doi = {10.1215/00127094-2023-0035}, volume = {173}, year = {2024}, } @article{20838, abstract = {For any 1 ≤ r ≤ ∞, we show that every diffeomorphism of a manifold of the form R/Z × M is a total renormalization of a Cr-close to identity map. In other words, for every diffeomorphism f of R/Z×M, there exists a map g arbitrarily close to identity such that the first return map of g to a domain is conjugate to f and moreover the orbit of this domain is equal to R/Z×M. This enables us to localize near the identity the existence of many properties in dynamical systems, such as being Bernoulli for a smooth volume form.}, author = {Berger, Pierre and Gourmelon, Nicolaz and Helfter, Mathieu}, issn = {1432-1297}, journal = {Inventiones mathematicae}, number = {2}, pages = {431--468}, publisher = {Springer Nature}, title = {{Every diffeomorphism is a total renormalization of a close to identity map}}, doi = {10.1007/s00222-024-01305-w}, volume = {239}, year = {2024}, } @article{20961, abstract = {Self-replicating molecules and well-defined folded macromolecules are of great significance in the emergence and evolution of life. How they may interconnect and affect each other remains largely elusive. Here, we demonstrate an abiotic system where a single building block can oligomerize to yield either a self-replicating molecule or a foldamer. Specifically, agitation of a disulfide-based dynamic combinatorial library at moderately elevated pH channels it selectively into a self-replicating hexamer assembled into fibers, after passing through a period where a 15-subunit macrocyclic foldamer existed transiently. Without mechanoagitation or at lower pH, the formation of hexamer fiber is suppressed, resulting in the accumulation of the 15mer foldamer. Foldamer and self-replicator can be interconverted in response to external stimuli, including agitation and a change in pH. Furthermore, upon the addition of a photoacid, the pH of the medium can be controlled by irradiation, driving the switching between replicator and foldamer and allowing a dissipative out-of-equilibrium state to be accessed, using light as a source of energy.}, author = {Jin, Yulong and Mandal, Pradeep K and Wu, Juntian and Kiani, Armin and Zhao, Rui and Huc, Ivan and Otto, Sijbren}, issn = {1520-5126}, journal = {Journal of the American Chemical Society}, number = {49}, pages = {33395--33402}, publisher = {American Chemical Society}, title = {{Light-mediated interconversion between a foldamer and a self-replicator}}, doi = {10.1021/jacs.4c09114}, volume = {146}, year = {2024}, } @article{20962, abstract = {Systems chemistry has emerged as a useful paradigm to access structures and phenomena typically exhibited by living systems, including complex molecular systems such as self-replicators and foldamers. As we progress further toward the noncovalent synthesis of life-like systems, and eventually life itself, it is necessary to gain control over assembly pathways. Dissipative chemical fueling has enabled access to stable populations of (self-assembled) structures that would normally form only transiently. Here, we report a synthetic dynamic combinatorial library, made from a single structurally simple building block, from which a self-replicator and a foldamer can emerge along two distinct and competing pathways through an inter- or intramolecular assembly process, respectively. A fueled chemical reaction cycle is then set up to generate the foldamer transiently, in the presence of the self-replicator. The partitioning of the building block between the folding and self-replication pathways and the duration of the fueled reaction cycles are controlled by adjusting the amount of the chemical fuel. An out-of-equilibrium steady state involving the two assemblies could also be achieved by using a continuous stirred tank reactor with inflow and outflow of material. This work connects the domains of folding and self-replication in synthetic systems through dissipative out-of-equilibrium chemistry. It demonstrates that foldamers and self-replicators, formed from the same building block, can stably coexist if the system is continuously supplied with energy, while at equilibrium, the Gibbs phase rule prohibits such coexistence.}, author = {Sood, Ankush and Mandal, Pradeep K and Ottelé, Jim and Wu, Juntian and Eleveld, Marcel and Hatai, Joydev and Pappas, Charalampos G. and Huc, Ivan and Otto, Sijbren}, issn = {1520-5126}, journal = {Journal of the American Chemical Society}, keywords = {Fibers, Foldamers, Macrocycles, Monomers, Peptides, Proteins}, number = {49}, pages = {33386--33394}, publisher = {American Chemical Society}, title = {{Simultaneous formation of a foldamer and a self-replicator by out-of-equilibrium dynamic covalent chemistry}}, doi = {10.1021/jacs.4c09111}, volume = {146}, year = {2024}, }