Biology subjects

Schneitz, K.

Publications and source records attributed to Schneitz, K..

14 recordsLinked to original sources

bioRxiv2026

Somatic DNA methylation heterogeneity predicts extreme transgenerational epimutation hotspots in Arabidopsis

Spontaneous epimutations are stochastic and heritable changes in cytosine methylation that arise independently of DNA sequence alterations. In plants, they occur predominantly at CG sites, accumulate across generations at high and clock-like rates, and contribute substantially to constitutive epigenetic diversity. Although these features are well established, the developmental origin of spontaneous epimutations remains poorly understood. One model proposes that they arise during somatic growth in different cell layers of the shoot apical meristem, where lineage bottlenecks enable nascent epimutations to clonally propagate into developing organs. Such layer-specific dynamics are expected to generate spatial mosaics of methylation states within tissues that manifest as heterogeneity in bulk bisulfite sequencing data. Here, we quantified genome-wide CG methylation heterogeneity in single Arabidopsis leaves using read-level methylation discordance metrics adopted from cancer epigenomics. We identified thousands of highly heterogeneous loci and show that they overlap extreme transgenerational epimutation hotspots. Analysis of multiple leaves from the same individuals further revealed that methylation divergence at these loci scales with developmental distance and recapitulates the branching architecture of the shoot. Together, these results support a meristematic origin of spontaneous epimutations and highlight a shared susceptibility to methylation-maintenance errors in both developmental and transgenerational contexts.

genetics↗

bioRxiv2024

The inner integument controls embryo sac development and seed shape in Arabidopsis thaliana

The angiosperm ovule is characterized by the close association of the two generations, with the haploid female gametophyte or embryo sac being encapsulated by the diploid sporophyte, which usually forms two integuments. How the gametophyte and sporophyte coordinate their development has long been of interest. However, the function of the inner integument in embryo sac development has remained elusive. Here, we addressed this question. We applied a genetic ablation strategy to achieve an early block in inner integument outgrowth. We generated plants expressing BARNASE under the control of an early acting endothelium-specific promoter. Corresponding lines carried ovules lacking most of the inner integument. The genetic and cell biological data revealed that in the near absence of an inner integument embryo sac development is blocked at the mono-nuclear embryo sac stage in most pre-fertilization ovules. Approximately 10 percent of the ovules developed a functional embryo sac and underwent fertilization. Subsequent embryo and endosperm development appeared unperturbed and viable seeds were produced albeit of altered shape. Our results show that the inner integument plays an important role in early embryo sac development as well as ovule and seed shape, but is dispensable for embryo and endosperm development. Key words: embryo sac, embryo, endothelium, female gametophyte, integument, seed development HighlightGenetic ablation of the inner integument demonstrates its role in embryo sac development and its irrelevance for embryogenesis.

plant biology↗

bioRxiv2024

A deep learning-based toolkit for 3D nuclei segmentation and quantitative analysis in cellular and tissue context

We present a new set of computational tools that enable accurate and widely applicable 3D segmentation of nuclei in various 3D digital organs. We developed a novel approach for ground truth generation and iterative training of 3D nuclear segmentation models, which we applied to popular CellPose, PlantSeg, and StarDist algorithms. We provide two high-quality models trained on plant nuclei that enable 3D segmentation of nuclei in datasets obtained from fixed or live samples, acquired from different plant and animal tissues, and stained with various nuclear stains or fluorescent protein-based nuclear reporters. We also share a diverse high-quality training dataset of about 10,000 nuclei. Furthermore, we advanced the MorphoGraphX analysis and visualization software by, among other things, providing a method for linking 3D segmented nuclei to their surrounding cells in 3D digital organs. We found that the nuclear-to-cell volume ratio varies between different ovule tissues and during the development of a tissue. Finally, we extended the PlantSeg 3D segmentation pipeline with a proofreading script that uses 3D segmented nuclei as seeds to correct cell segmentation errors in difficult-to-segment tissues. Summary StatementWe present computational tools that allow versatile and accurate 3D nuclear segmentation in plant organs, enable the analysis of cell-nucleus geometric relationships, and improve the accuracy of 3D cell segmentation.

plant biology↗

bioRxiv2023

Diverse 3D cellular patterns underlie the development of Cardamine hirsuta and Arabidopsis thaliana ovules

A fundamental question in biology is how organ morphogenesis comes about. The ovules of Arabidopsis thaliana have been established as a successful model to study numerous aspects of tissue morphogenesis; however, little is known regarding the relative contributions and dynamics of differential tissue and cellular growth and architecture in establishing ovule morphogenesis in different species. To address this issue, we generated a 3D digital atlas of Cardamine hirsuta ovule development with full cellular resolution. We combined quantitative comparative morphometrics and topological analysis to explore similarities and differences in the 3D cellular architectures underlying ovule development of the two species. We discovered that they show diversity in the way the three radial cell layers of the primordium contribute to its growth, in the formation of a new cell layer in the inner integument and, in certain cases, in the topological properties of the 3D cell architectures of homologous tissues despite their similar shape. Our work demonstrates the power of comparative 3D cellular morphometry and the importance of internal tissues and their cellular architecture in organ morphogenesis. Summary StatementQuantitative morphometric comparison of 3D digital ovules at full cellular resolution reveals diversity in internal 3D cellular architectures between similarly shaped ovules of Cardamine hirsuta and Arabidopsis thaliana.

plant biology↗

bioRxiv2022

The Arabidopsis MCTP member QUIRKY regulates the formation of the STRUBBELIG receptor kinase complex

Intercellular communication plays a central role in organogenesis. Tissue morphogenesis in Arabidopsis thaliana requires signaling mediated by a cell surface complex containing the atypical receptor kinase STRUBBELIG (SUB) and the multiple C2 domains and transmembrane region protein QUIRKY (QKY). QKY is required to stabilize SUB at the plasma membrane. However, it is unclear what the in vivo architecture of the QKY/SUB signaling complex is, how it is controlled, and how it relates to the maintenance of SUB at the cell surface. Using a combination of yeast two-hybrid assays and Forster resonance energy transfer (FRET)/fluorescence lifetime imaging microscopy (FLIM) in epidermal cells of seedling roots we find that QKY promotes the formation of SUB homo-oligomers in vivo, a process that appears to involve an interaction between the extracellular domains of SUB. We also show that QKY and SUB physically interact and form a complex at the cell surface in vivo. In addition, the data show that the N-terminal C2A-B region of QKY interacts with the intracellular domain of SUB. They further reveal that this interaction is essential to maintain SUB levels at the cell surface. Finally, we provide evidence that QKY forms homo-multimers in vivo in a SUB-independent manner. We suggest a model in which the physical interaction of QKY with SUB mediates the oligomerization of SUB and attenuates its internalization, thereby maintaining sufficiently high levels of SUB at the cell surface required for the control of tissue morphogenesis.

plant biology↗

bioRxiv2021

The annotation and analysis of complex 3D plant organs using 3DCoordX

A fundamental question in biology concerns how molecular and cellular processes become integrated during morphogenesis. In plants, characterization of 3D digital representations of organs at single-cell resolution represents a promising approach to addressing this problem. A major challenge is to provide organ-centric spatial context to cells of an organ. We developed several general rules for the annotation of cell position and embodied them in 3DCoordX, a user-interactive computer toolbox implemented in the open-source software MorphoGraphX. It enables rapid spatial annotation of cells even in highly curved biological shapes. With the help of 3DCoordX we obtained new insight by analyzing cellular growth patterns in organs of several species. For example, the data indicated the presence of a basal cell proliferation zone in the ovule primordium of Arabidopsis thaliana. Proof-of-concept analyses suggested a preferential increase in cell length associated with neck elongation in the archegonium of Marchantia polymorpha and variations in cell volume linked to central morphogenetic features of a trap of the carnivorous plant Utricularia gibba. Our work demonstrates the broad applicability of the developed strategies as they provide organ-centric spatial context to cellular features in plant organs of diverse shape complexity.

plant biology↗

bioRxiv2021

MorphoGraphX 2.0: Providing context for biological image analysis with positional information

Positional information is a central concept in developmental biology. In developing organs, positional information can be idealized as a local coordinate system that arises from morphogen gradients controlled by organizers at key locations. This offers a plausible mechanism for the integration of the molecular networks operating in individual cells into the spatially-coordinated multicellular responses necessary for the organization of emergent forms. Understanding how positional cues guide morphogenesis requires the quantification of gene expression and growth dynamics in the context of their underlying coordinate systems. Here we present recent advances in the MorphoGraphX software (Barbier de Reuille et al. eLife 2015;4:e05864) that implement a generalized framework to annotate developing organs with local coordinate systems. These coordinate systems introduce an organ-centric spatial context to microscopy data, allowing gene expression and growth to be quantified and compared in the context of the positional information thought to control them.

developmental biology↗

bioRxiv2020

Cell wall damage attenuates root hair cell patterning and tissue morphogenesis mediated by the Arabidopsis receptor kinase STRUBBELIG.

Cell wall remodeling is essential for the control of growth and development as well as the regulation of stress responses. However, the underlying cell wall monitoring mechanisms remain poorly understood. Regulation of root hair fate and flower development in Arabidopsis thaliana requires signaling mediated by the atypical receptor kinase STRUBBELIG (SUB). Furthermore, SUB is involved in cell wall integrity signaling and regulates the cellular response to reduced levels of cellulose, a central component of the cell wall. Here, we show that continuous exposure to sub-lethal doses of the cellulose biosynthesis inhibitor isoxaben results in altered root hair patterning and floral morphogenesis. Genetically impairing cellulose biosynthesis also results in root hair patterning defects. We further show that isoxaben exerts its developmental effects through the attenuation of SUB signaling. Our evidence indicates that down-regulation of SUB is a multi-step process and involves changes in SUB complex architecture at the plasma membrane, enhanced removal of SUB from the cell surface, and downregulation of SUB transcript levels. The results provide molecular insight into how the cell wall regulates cell fate and tissue morphogenesis.

plant biology↗

bioRxiv2020

A digital 3D reference atlas reveals cellular growth patterns shaping the Arabidopsis ovule

A fundamental question in biology is how morphogenesis integrates the multitude of distinct processes that act at different scales, ranging from the molecular control of gene expression to cellular coordination in a tissue. Investigating morphogenesis of complex organs strongly benefits from three-dimensional representations of the organ under study. Here, we present a digital analysis of ovule development from Arabidopsis thaliana as a paradigm for a complex morphogenetic process. Using machine-learning-based image analysis we generated a three-dimensional atlas of ovule development with cellular resolution. It allows quantitative stage- and tissue-specific analysis of cellular patterns. Exploiting a fluorescent reporter enabled precise spatial determination of gene expression patterns, revealing subepidermal expression of WUSCHEL. Underlying the power of our approach, we found that primordium outgrowth progresses evenly, discovered a novel mode of forming a new cell layer, and detected a new function of INNER NO OUTER in restricting cell proliferation in the nucellus. Moreover, we identified two distinct subepidermal cell populations that make crucial contributions to ovule curvature. Our work demonstrates the expedience of a three-dimensional digital representation when studying the morphogenesis of an organ of complex cellular architecture and shape that eventually consists of 1,900 cells.

plant biology↗

bioRxiv2020

Accurate And Versatile 3D Segmentation Of Plant Tissues At Cellular Resolution

Quantitative analysis of plant and animal morphogenesis requires accurate segmentation of individual cells in volumetric images of growing organs. In the last years, deep learning has provided robust automated algorithms that approach human performance, with applications to bio-image analysis now starting to emerge. Here, we present PlantSeg, a pipeline for volumetric segmentation of plant tissues into cells. PlantSeg employs a convolutional neural network to predict cell boundaries and graph partitioning to segment cells based on the neural network predictions. PlantSeg was trained on fixed and live plant organs imaged with confocal and light sheet microscopes. PlantSeg delivers accurate results and generalizes well across different tissues, scales, and acquisition settings. We present results of PlantSeg applications in diverse developmental contexts. PlantSeg is free and open-source, with both a command line and a user-friendly graphical interface.

plant biology↗

bioRxiv2019

The Arabidopsis receptor kinase STRUBBELIG regulates the response to cellulose deficiency

Plant cells are encased in a semi-rigid cell wall of complex build. As a consequence, cell wall remodeling is essential for the control of growth and development as well as the regulation of abiotic and biotic stress responses. Plant cells actively sense physico-chemical changes in the cell wall and initiate corresponding cellular responses. However, the underlying cell wall monitoring mechanisms remain poorly understood. In Arabidopsis the atypical receptor kinase STRUBBELIG (SUB) mediates tissue morphogenesis. Here, we show that SUB-mediated signal transduction also regulates the cellular response to a reduction in the biosynthesis of cellulose, a central carbohydrate component of the cell wall. SUB signaling affects early increase of intracellular reactive oxygen species, stress gene induction as well as ectopic lignin and callose accumulation upon exogenous application of the cellulose biosynthesis inhibitor isoxaben. Moreover, our data reveal that SUB signaling is required for maintaining cell size and shape of root epidermal cells and the recovery of root growth after transient exposure to isoxaben. SUB is also required for root growth arrest in mutants with defective cellulose biosynthesis. Genetic data further indicate that SUB controls the isoxaben-induced cell wall stress response independently from other known receptor kinase genes mediating this response, such as THESEUS1 or MIK2. We propose that SUB functions in a least two distinct biological processes: the control of tissue morphogenesis and the response to cell wall damage. Taken together, our results reveal a novel signal transduction pathway that contributes to the molecular framework underlying cell wall integrity signaling.\n\nAuthor SummaryPlant cells are encapsulated by a semi-rigid and biochemically complex cell wall. This particular feature has consequences for multiple biologically important processes, such as cell and organ growth or various stress responses. For a plant cell to grow the cell wall has to be modified to allow cell expansion, which is driven by outward-directed turgor pressure generated inside the cell. In return, changes in cell wall architecture need to be monitored by individual cells, and to be coordinated across cells in a growing tissue, for an organ to attain its regular size and shape. Cell wall surveillance also comes also into play in the reaction against certain stresses, including for example infection by plant pathogens, many of which break through the cell wall during infection, thereby generating wall-derived factors that can induce defense responses. There is only limited knowledge regarding the molecular system that monitors the composition and status of the cell wall. Here we provide further insight into the mechanism. We show that the cell surface receptor STRUBBELIG, previously known to control organ development in Arabidopsis, also promotes the cells response to reduced amounts of cellulose, a main component of the cell wall.

plant biology↗

bioRxiv2018

The AGC protein kinase UNICORN controls planar growth by attenuating PDK1 in Arabidopsis thaliana

Tissue morphogenesis critically depends on the coordination of cellular growth patterns. In plants, many organs consist of clonally distinct cell layers, such as the epidermis, whose cells undergo divisions that are oriented along the plane of the layer. The developmental control of such planar growth is poorly understood. We have previously identified the Arabidopsis AGCVIII-class protein kinase UNICORN (UCN) as a central regulator of this process. Plants lacking UCN activity show spontaneous formation of ectopic multicellular protrusions in integuments and malformed petals indicating that UCN suppresses uncontrolled growth in those tissues. In the current model UCN regulates planar growth of integuments in part by directly repressing the putative transcription factor ABERRANT TESTA SHAPE (ATS). Here we report on the identification of 3-PHOSPHOINOSITIDE-DEPENDENT PROTEIN KINASE 1 (PDK1) as a novel factor involved in UCN-mediated growth control. PDK1 constitutes a basic component of signaling mediated by AGC protein kinases throughout eukaryotes. Arabidopsis PDK1 is implied in stress responses and growth promotion. Here we show that loss-of-function mutations in PDK1 suppress aberrant growth in integuments and petals of ucn mutants. Additional genetic, in vitro, and cell biological data support the view that UCN functions by repressing PDK1. Furthermore, our data indicate that PDK1 is indirectly required for deregulated growth caused by ATS overexpression. Our findings support a model proposing that UCN suppresses ectopic growth in integuments through two independent processes: the attenuation of the protein kinase PDK1 in the cytoplasm and the repression of the transcription factor ATS in the nucleus.\n\nAuthor SummaryPlant organs, such as petals or roots, are composites of distinct cell layers. As a rule, cells making up a layer, for example the epidermis, the outermost layer of a tissue, divide \"within the plane\" of the layer. This cellular behavior results in the two-dimensional sheet-like or planar growth of the cell layer. The mechanism orchestrating such a growth pattern is poorly understood. In particular, it is unclear how uncontrolled and \"out-of-plane\" growth is avoided. Here we provide insight into this process. Our data indicate that higher than normal activity of a central regulator of growth and stress responses results in wavy and malformed petals and in protrusion-like aberrant outgrowths in the tissue that will develop into the seed coat. It is therefore important to keep this factor in check to allow proper formation of those tissues. We further show that a protein called UNICORN attenuates the activity of this regulator thereby ensuring the sheet-like growth of young petals or the developing seed coat.

plant biology↗

bioRxiv2018

Asymmetric redundancy of ZERZAUST and ZERZAUST HOMOLOG in different accessions of Arabidopsis thaliana

Divergence among duplicate genes is one of the important sources of evolutionary innovation. But, the contribution of duplicate divergence to variation in Arabidopsis accessions is sparsely known. Recently, we studied the role of a cell wall localized protein, ZERZAUST (ZET), in Landsberg erecta (Ler) accession. Here, we present the study of ZET in Columbia (Col) accession, which not only showed differential expression patterns in comparison to Ler, but also revealed its close homolog, ZERZAUST HOMOLOG (ZETH). Although, genetic analysis implied redundancy, expression analysis revealed divergence, with ZETH showing minimal expression in both Col and Ler. In addition, ZETH shows relatively higher expression levels in Col compared to Ler. Our data also reveal compensatory up-regulation of ZETH in Col, but not in Ler, implying it is perhaps dispensable in Ler. However, a novel CRISPR/Cas9-induced zeth allele confirmed that ZETH has residual activity in Ler. The results provide genetic evidence for accession-specific differences in compensation mechanism and asymmetric gene contribution. Thus, our work reveals a novel example for how weakly expressed homologs contribute to diversity among accessions.

genetics↗

bioRxiv2018

Tissue morphogenesis mediated by the Arabidopsis receptor kinase STRUBBELIG involves a clathrin-dependent process

HighlightThe Arabidopsis receptor kinase STRUBBELIG is internalized by clathrin-mediated endocytosis and affects clathrin-dependent processes in a tissue-dependent manner.\n\nAbstractSignaling mediated by cell surface receptor kinases is central to the coordination of growth patterns during organogenesis. Receptor kinase signaling is in part controlled through endocytosis and subcellular distribution of the respective receptor kinase. For the majority of plant cell surface receptors the underlying trafficking mechanisms are not characterized. In Arabidopsis, tissue morphogenesis relies on the atypical receptor kinase STRUBBELIG (SUB). Here, we approach the endocytic mechanism of SUB. Our data reveal that a functional SUB:EGFP fusion is ubiquitinated in vivo. We further show that plasma membrane-bound SUB:EGFP becomes internalized in a clathrin-dependent fashion. We also find that SUB:EGFP associates with the trans-Golgi network and accumulates in multivesicular bodies and the vacuole. Coimmunoprecipitation experiments reveal that SUB:EGFP and clathrin are present within the same protein complex. Our genetic analysis shows that SUB and CLATHRIN HEAVY CHAIN 2 promote root hair patterning. By contrast, SUB behaves as a negative regulator of a clathrin-dependent process during floral development. Taken together, the data indicate that SUB undergoes clathrin-mediated endocytosis, that this process does not dependent on stimulation of SUB signaling by an exogenous agent, and that SUB genetically interacts with clathrin-dependent pathways in a tissue-specific manner.

plant biology↗