Biology subjects

Krueger, S.

Publications and source records attributed to Krueger, S..

14 recordsLinked to original sources

bioRxiv2026

Functional specialization of the gibberellin receptor GIBBERELLIN-INSENSITIVE DWARF 1C in plant neighbour detection

Plants detect neighbours through a reduced red-to-far-red ratio (R:FR), triggering elongation growth that reduces crop yield. Although Gibberellin (GA) is required for the neighbour-proximity (NP) elongation response, bioactive GA levels do not increase sufficiently to account for elongation magnitude, suggesting GA sensitivity as an additional regulated variable. Here, we show that GID1C, one of three Arabidopsis GA receptors, is the primary GA receptor involved in NP-induced elongation. GID1C protein accumulates selectively in hypocotyls and root tips under low R:FR without an increase in bioactive GA. The gid1c mutant shows a reduced elongation response that exogenous GA treatment cannot rescue. Transcriptome profiling reveals that GID1C controls 86% of the NP-responsive transcriptome, including genes for cell growth, division, and transcriptional regulation. Hub analysis identifies ICE1 as a GID1C-repressed transcriptional brake. ICE1 transcript is suppressed under low R:FR in a GID1C-dependent manner, and a phosphorylation-resistant ICE1 allele blocks NP-induced elongation. Together, these findings establish GA perception as an additional regulatory layer in NP, with subfunctionalisation among GID1 paralogs shaping the response to neighbouring plants.

plant biology↗

bioRxiv2026

Endosomal mRNA transport coordinates local mitochondrial bioenergetics during polar fungal growth

Mitochondrial function relies on the precise spatial coordination of protein synthesis and import. Most mitochondrial proteins are nuclear-encoded and must be supplied across varying intracellular distances. In highly polarized cells such as fungal hyphae and neurons, active long-distance mRNA transport is thought to sustain distal mitochondrial function, but its mechanistic coupling to protein import and organelle physiology is unclear. Here, we demonstrate that endosomal transport of mRNAs encoding mitochondrial proteins orchestrates local bioenergetics in infectious hyphae of Ustilago maydis. Using the subunit Atp3 of electron transport chain Complex V as a model, we uncover that the endosomal mRNA transporter Rrm4 is required for efficient mitochondrial protein import, particularly at growth poles. Loss of Rrm4 leads to defects in mitochondrial import, resulting in altered physiology. We propose that endosome-coupled mRNA transport constitutes a fundamental layer of subcellular mitochondrial homeostasis, with implications extending from fungal pathogenicity to neuronal disease. Significance StatementFungal pathogens depend on efficient polar growth to execute their infection programs. Consequently, their growing cell poles face a massive local demand of energy, which is supplied by mitochondria. Currently, it is unclear how involved proteins of the mitochondrial electron transport chain (ETC) reach these distant organelles. Here, we combine fungal genetics, metabolomics, transcriptomics and minimal invasive live-cell imaging to resolve this spatial challenge in the corn pathogen Ustilago maydis. We discover that long-distance endosomal hitchhiking of mRNAs encoding mitochondrial ETC components is essential to sustain active mitochondria at the expanding pole. Ultimately, this membrane-coupled mRNA trafficking precisely orchestrates subcellular mitochondrial function, disclosing a previously unrecognized Achilles heel for the development of novel fungicides.

microbiology↗

bioRxiv2026

Context-dependent siderophore exploitability shapes microbial community structure

1)Siderophores are classically viewed as shared iron-scavenging public goods, yet their ecological roles in multispecies communities remain poorly defined. Here, we establish a synthetic microbial community to dissect how different siderophores, their uptake compatibility and spatial structure shape iron competition. Using Corynebacterium glutamicum as a model, we show that this siderophore non-producer accesses diverse xenosiderophores, including enterobactin secreted by Escherichia coli. However, exploitation was constrained and co-cultures converged to stable compositions. Dose-response experiments combined with mathematical modelling indicated that the producer retains more effective access to enterobactin than the exploiter. Presence of Pseudomonas putida altered this interaction, as it exploited enterobactin while producing pyoverdine, a siderophore inaccessible to the other community members that restricted their iron access. Across different cultivation scales, community dynamics was strongly influenced by spatial organization and initial composition. These findings identify siderophores as context-dependent iron-allocation agents that can promote microbial coexistence or exclusion.

microbiology↗

bioRxiv2026

Glycogen deficiency impairs diurnal energy metabolism and cell division in Synechocystis

Diurnal changes in light availability are a defining feature of life on Earth. Photoautotrophic organisms therefore store reduced carbon during the day to sustain energy metabolism at night. In cyanobacteria, glycogen is the primary carbon storage compound and supports both energy homeostasis and stress responses. Although glycogen-deficient Synechocystis strains have been studied previously, how these mutants cope with the loss of the major daytime carbon sink and can sustain themselves during the night remains unclear. Using single-cell microfluidics, transcriptomics, and metabolomics, we show that {Delta}glgC mutants exhibit pronounced light sensitivity. At sub-lethal light intensities, daytime transcriptional responses are dominated by downregulation of photosynthesis-related genes, likely preventing NADPH overaccumulation in the absence of a carbon sink. During the night, mutants display severe energy limitation, characterized by reduced ATP levels, altered redox balance, and depletion of central carbon intermediates. In contrast, fumarate and malate accumulate, indicating enhanced respiratory flux through succinate dehydrogenase. These metabolic constraints lead to extended lag phases and delayed cell divisions after the onset of light, demonstrating that glycogen-deficient cells fail to efficiently reinitiate growth after dawn. Overall, our results as a snapshot of the initial response to diurnal regimes highlight glycogen as a central integrator of diurnal physiology in Synechocystis, coordinating energy metabolism, redox balance, and cell division, with implications for metabolic robustness and the evolutionary constraints shaping (endo)symbiosis. Short summaryGlycogen deficiency disrupts day-night energy and redox homeostasis in Synechocystis, revealing constraints on growth, division, and symbiotic potential.

plant biology↗

bioRxiv2025

Widespread low-affinity motifs enhance chromatin accessibility and regulatory potential in mESCs

Low-affinity transcription factor (TF) motifs are an important element of the cis-regulatory code, yet they are notoriously difficult to map and mechanistically incompletely understood, limiting our ability to interpret non-coding variation in development, evolution, and disease. Here we investigate their role in pioneering and leverage sequence-to-profile models of chromatin accessibility in mouse embryonic stem cells to reliably map and interpret low-affinity motifs across the genome. We find that low-affinity motifs have outsized effects by cooperating with nearby motifs through intra-nucleosomal soft syntax. By modeling nucleosome-mediated cooperativity with a kinetic model, we discover and validate that pioneer cooperativity makes a motif operate at higher pioneering ranges across changing TF concentrations, thereby raising the regulatory potential. These results show that low-affinity motifs can be accurately mapped, shape the properties of developmental enhancers and likely play a widespread role in fine-tuning enhancers during evolution.

genomics↗

bioRxiv2025

Root growth in Arabidopsis depends on the amount of glutathione and not the glutathione redox potential

Cell cycle activity and hence plant growth strictly depends on glutathione homeostasis. Despite compelling evidence for this dependency based on glutathione depletion, the cause for the cell cycle arrest had remained unclear. Cell cycle control may depend on either the absolute amount of glutathione or on the glutathione redox potential (EGSH). To unambiguously distinguish those two options, we characterized an allelic series of six Arabidopsis mutants affected in glutamate-cysteine ligase, which catalyses the first step for the biosynthesis of reduced glutathione (GSH). When grown under the same conditions, even mutants with 20% and 50% of wild-type GSH amounts are slightly stunted. The most severely compromised mutants, zir1 and rml1, were crossed with either gr1, which lacks cyto-nuclear glutathione disulfide reductase and was used to induce a pronounced shift in EGSH, or with bir6, which has a diminished glutathione consumption and thus exhibits slightly increased levels of GSH. Based on theoretical considerations, these levels are not expected to shift the EGSH to any significant extent. Our study shows that deleting GR1 in the zir1 or rml1 background does not result in an obvious phenotypic change. By contrast, deleting BIR6 was sufficient to suppress the growth arrest in rml1 and to attenuate the growth in zir1. These findings demonstrate that root growth is dependent on the availability of sufficient amounts of GSH, and not affected by pronounced changes in EGSH. This insight provides a decisive step towards understanding the mechanisms underpinning the proposed role of glutathione in cell cycle and growth control. HighlightCrosses of GSH-deficient Arabidopsis mutants with mutants affected in glutathione reduction or GSH consumption pinpoint the GSH amount rather than the glutathione redox potential as causative for changes in root growth.

plant biology↗

bioRxiv2025

Solution conformational differences between conventional and CENP-A nucleosomes are accentuated by reversible deformation under high pressure

Solution-based interrogation of the physical nature of nucleosomes has its roots in X-ray and neutron scattering experiments, including those that provided the initial observation that DNA wraps around core histones. In this study, we performed a comprehensive small-angle scattering study to compare canonical nucleosomes with variant centromeric nucleosomes harboring the histone variant, CENP-A. We used nucleosome core particles (NCPs) assembled on an artificial positioning sequence (Widom 601) and compared these to those assembled on a natural -satellite DNA cloned from human centromeres. We establish the native solution properties of octameric H3 and CENP-A NCPs using analytical ultracentrifugation (AUC), small-angle X-ray scattering (SAXS), and contrast variation small-angle neutron scattering (CV-SANS). Using high-pressure SAXS (HP-SAXS), we discovered that both histone identity and DNA sequence have an impact on the stability of octameric nucleosomes in solution under high pressure (300 MPa), with evidence of reversible unwrapping in these experimental conditions. Both canonical nucleosomes harboring conventional histone H3 and their centromeric counterparts harboring CENP-A have a substantial increase in their radius of gyration, but this increase is much less prominent for centromeric nucleosomes. More broadly for chromosome-related research, we note that as HP-SAXS methodologies expand in their utility, we anticipate this will provide a powerful solution-based approach to study nucleosomes and higher-order chromatin complexes.

biophysics↗

bioRxiv2024

The plant specific cohesin subunit SYN4 contributes to 3D genome organization and activity

O_LIChromatin architecture in the cells of animals and fungi influences gene expression. The molecular factors that influence higher genome architecture in plants and their effects on gene expression remain unknown. Cohesin complexes, conserved in eukaryotes, are essential factors in genome structuring. Here, we investigated the relevance of the plant-specific somatic cohesin subunit SYN4 for chromatin organisation in Arabidopsis thaliana. C_LIO_LIPlants mutated in SYN4 were studied using HRM, Hi-C, RNA sequencing, untargeted and targeted metabolomics and physiological assays to understand the role of this plant-specific cohesin. C_LIO_LIWe show that syn4 mutants exhibit altered intra- and interchromosomal interactions, expressed as sharply reduced contacts between telomeres and chromosome arms but not between centromeres, and differences in the placement and number of topologically associated domains (TADs)-like structures. By transcriptome sequencing, we also show that syn4 mutants have altered gene expression, including numerous genes that control abiotic stress responses. C_LIO_LIThe response to drought stress in Arabidopsis is strongly influenced by the genome structure in syn4 mutants, potentially due to altered expression of CYP707A3, an ABA 8-hydroxylase. C_LI In briefThe 3D architecture of the genome extensively influences gene expression in animals and fungi. We show that the plant-specific cohesin subunit SYN4 affects intra- and interchromosomal interactions including telomere clustering, with consequences for the expression of genes of transient and induced biological pathways and the biosynthesis of bioactive compounds. Highly condensed genome structures at the CYP707A3 locus positively affects the stress response to water deprivation regulated by abscisic acid.

genetics↗

bioRxiv2024

Interpretable deep learning reveals the sequence rules of Hippo signaling

The response to signaling pathways is highly context-specific, and identifying the transcription factors and mechanisms that are responsible is very challenging. Using the Hippo pathway in mouse trophoblast stem cells as a model, we show here that this information is encoded in cis-regulatory sequences and can be learned from high-resolution binding data of signaling transcription factors. Using interpretable deep learning, we show that the binding levels of TEAD4 and YAP1 are enhanced in a distance-dependent manner by cell type-specific transcription factors, including TFAP2C. We also discovered that strictly spaced Tead double motifs are widespread highly active canonical response elements that mediate cooperativity by promoting labile TEAD4 protein-protein interactions on DNA. These syntax rules and mechanisms apply genome-wide and allow us to predict how small sequence changes alter the activity of enhancers in vivo. This illustrates the power of interpretable deep learning to decode canonical and cell type-specific sequence rules of signaling pathways. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=183 SRC="FIGDIR/small/580842v1_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@1ee22e9org.highwire.dtl.DTLVardef@1362c82org.highwire.dtl.DTLVardef@1a42512org.highwire.dtl.DTLVardef@17dd619_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

bioRxiv2023

Binding-site resolution structure and dynamics of Cebpa regulatory regions during macrophage-neutrophil differentiation

The upregulation of gene expression by enhancers depends upon the interplay between the binding of sequence-specific transcription factors (TFs) and DNA accessibility. DNA accessibility is thought to limit the ability of TFs to bind to their sites, while TFs can increase accessibility to recruit additional factors that upregulate gene expression. Given this interplay, the causative regulatory events underlying the modulation of gene expression during cellular differentiation remain unknown for the vast majority of genes. We investigated the binding-site resolution dynamics of DNA accessibility and the expression dynamics of the enhancers of an important neutrophil gene, Cebpa, during macrophage-neutrophil differentiation. Reporter genes were integrated in a site-specific manner in PUER cells, which are progenitors that can be differentiated into neutrophils or macrophages in vitro by activating the pan-leukocyte TF PU.1. Time series data show that two enhancers upregulate reporter expression during the first 48 hours of neutrophil differentiation. Surprisingly, there is little or no increase in the total accessibility, measured by ATAC-Seq, of the enhancers during the same time period. Conversely, total accessibility peaks 96 hrs after PU.1 activation--consistent with its role as a pioneer--but the enhancers do not upregulate gene expression. Combining deeply sequenced ATAC-Seq data with a new bias-correction method allowed the profiling of acces-sibility at single-nucleotide resolution and revealed protected regions in the enhancers that match all previously characterized TF binding sites and ChIP-Seq data. Although the accessibility of most positions does not change during early differentiation, that of positions neighboring TF binding sites, an indicator of TF occupancy, did in-crease significantly. The localized accessibility changes are limited to nucleotides neighboring C/EBP-family TF binding sites, showing that the upregulation of enhancer activity during early differentiation is driven by C/EBP-family TF binding. These results show that increasing the total accessibility of enhancers is not sufficient for upregulating their activity and other events such as TF binding are necessary for upregulation. Also, TF binding can cause upregulation without a perceptible increase in total accessibility. Finally, this study demonstrates the feasibility of comprehensively mapping individual TF binding sites as footprints using high coverage ATAC-Seq and inferring the sequence of events in gene regulation by combining with time-series gene expression data.

genomics↗

bioRxiv2022

Chromatin accessibility is a two-tier process regulated by transcription factor pioneering and enhancer activation

Chromatin accessibility is integral to the process by which transcription factors (TFs) read out cis-regulatory DNA sequences, but it is difficult to differentiate between TFs that drive accessibility and those that do not. Deep learning models that learn complex sequence rules provide an unprecedented opportunity to dissect this problem. Using zygotic genome activation in the Drosophila embryo as a model, we generated high-resolution TF binding and chromatin accessibility data, analyzed the data with interpretable deep learning, and performed genetic experiments for validation. We uncover a clear hierarchical relationship between the pioneer TF Zelda and the TFs involved in axis patterning. Zelda consistently pioneers chromatin accessibility proportional to motif affinity, while patterning TFs augment chromatin accessibility in sequence contexts in which they mediate enhancer activation. We conclude that chromatin accessibility occurs in two phases: one through pioneering, which makes enhancers accessible but not necessarily active, and a second when the correct combination of transcription factors leads to enhancer activation.

genomics↗

bioRxiv2019

SurA is a "Groove-y" Chaperone That Expands Unfolded Outer Membrane Proteins

The periplasmic chaperone network ensures the biogenesis of bacterial outer membrane proteins (OMPs) and has recently been identified as a promising target for antibiotics. SurA is the most important member of this network both due to its genetic interaction with the {beta}-barrel assembly machinery complex as well as its ability to prevent unfolded OMP (uOMP) aggregation. Using only binding energy, the mechanism by which SurA carries out these two functions is not well understood. Here we use a combination of photo-crosslinking, mass spectrometry, solution scattering, and molecular modeling techniques to elucidate the key structural features that define how SurA solubilizes uOMPs. Our experimental data support a model in which SurA binds uOMPs in a groove formed between the core and P1 domains. This binding event results in a drastic expansion of the rest of the uOMP, which has many biological implications. Using these experimental data as restraints, we adopted an integrative modeling approach to create a sparse ensemble of models of a SurA*uOMP complex. We validated key structural features of the SurA*uOMP ensemble using independent scattering and chemical crosslinking data. Our data suggest that SurA utilizes three distinct binding modes to interact with uOMPs and that more than one SurA can bind a uOMP at a time. This work demonstrates that SurA operates in a distinct fashion compared to other chaperones in the OMP biogenesis network. Significance StatementOuter membrane proteins play critical roles in bacterial physiology and increasingly are exploited as antibiotic targets. SurA is the most important chaperone in the OMP biogenesis network and is thought to initiate their folding through an interaction with the BAM complex. We observe an unprecedented expansion of unfolded outer membrane proteins when bound to SurA. This expansion suggests a potential mechanism by which SurA can deliver uOMPs to the BAM complex. In addition, this study highlights the use of an integrative/hybrid structural biology approach and emerging methods to map highly heterogeneous structural ensembles such as that of an unfolded protein bound to a chaperone.

biophysics↗

bioRxiv2019

Deep learning at base-resolution reveals motif syntax of the cis-regulatory code

The arrangement of transcription factor (TF) binding motifs (syntax) is an important part of the cis-regulatory code, yet remains elusive. We introduce a deep learning model, BPNet, that uses DNA sequence to predict base-resolution ChIP-nexus binding profiles of pluripotency TFs. We develop interpretation tools to learn predictive motif representations and identify soft syntax rules for cooperative TF binding interactions. Strikingly, Nanog preferentially binds with helical periodicity, and TFs often cooperate in a directional manner, which we validate using CRISPR-induced point mutations. Our model represents a powerful general approach to uncover the motifs and syntax of cis-regulatory sequences in genomics data. HighlightsO_LIThe neural network BPNet accurately predicts TF binding data at base-resolution. C_LIO_LIModel interpretation discovers TF motifs and TF interactions dependent on soft syntax. C_LIO_LIMotifs for Nanog and partners are preferentially spaced at [~]10.5 bp periodicity. C_LIO_LIDirectional cooperativity is validated: Sox2 enhances Nanog binding, but not vice versa. C_LI

genomics↗

bioRxiv2018

The regulatory control of Cebpa enhancers and silencers in the myeloid and red-blood cell lineages

During development, cell identity is determined by lineage-specific transcriptional programs en-coded in the cis-regulatory DNA sequence of developmental genes. The sequence-level regulatory logic--identities of bound transcription factors (TFs), TF binding sites, and TF regulatory roles--of most developmental cis-regulatory modules (CRMs) is yet to be determined. We had previously developed an approach for inferring regulatory logic de novo by training sequence-based thermo-dynamic models on comprehensive reporter activity and gene expression datasets and applied it to Cebpa, an important hematopoietic gene. Here, we experimentally test thermodynamic models to decode the cis-regulatory logic of 4 enhancers and 3 silencers neighboring Cebpa at the resolution of individual binding sites. Cebpa is expressed at high and intermediate levels in neutrophils and macrophages respectively and downregulated in non-myeloid lineages. We tested the binding sites and functional roles of inferred TFs by designing and constructing mutated CRMs and comparing theoretical predictions of their activity against empirical measurements. Reporter activity was measured in PUER cells, which can be induced to differentiate into macrophages or neutrophils. All four enhancers were found to be simultaneously active in undifferentiated PUER cells and early-stage macrophages and neutrophils, and activated by combinations of PU.1, C/EBP family TFs, Egr1, and Gfi1. We show that silencers repress the activity of the proximal promoter in a dominant manner in G1ME cells, which are derived from the red-blood cell lineage. Dominant repression in G1ME cells can be traced to binding sites for GATA and Myb, a motif shared by all of the silencers. Finally, we demonstrate that GATA and Myb act redundantly to silence the proximal promoter. The result that silencers quench the promoter selectively in non-myeloid cells indicates that dominant repression is a novel mechanism for resolving hematopoietic lineages. Furthermore, Cebpa has a fail-safe cis-regulatory architecture, featuring several functionally similar CRMs, each of which contains redundant binding sites for multiple TFs. Lastly, by experimentally demonstrating the predictive ability of our sequence-based thermodynamic models, this work highlights the utility of this computational approach for decoding the logic of mammalian gene regulation.

developmental biology↗