Biology subjects

Hejatko, J.

Publications and source records attributed to Hejatko, J..

15 recordsLinked to original sources

bioRxiv2026

PWO1 and TRB proteins coordinate chromatin regulation to prevent premature differentiation and ectopic lignin deposition in Arabidopsis

The Arabidopsis PWWP-DOMAIN INTERACTOR OF POLYCOMBS1 (PWO1) and Telomere Repeat-Binding Proteins 1-3 (TRB1-3, TRBs) associate with distinct and shared protein complexes involved in epigenetic regulation, yet their cooperative roles in chromatin control and plant development remain largely unexplored. Here, we show that the interaction between PWO1 and TRBs is evolutionarily conserved. Both PWO1 and TRBs associate with plant telomeres, interact at these regions, and are co-enriched at subsets of interspersed telo-box motifs across regulatory regions genome-wide. TRBs facilitate PWO1 binding at shared genomic regions, including telo-box motifs. PWO1 and TRBs share a substantial number of genomic targets and preferentially bind chromatin regions associated with transcriptionally active states, whereas TRBs alone associate with repressive marks at thousands of loci. Genetic analyses show that the pwo1 trb1 trb3 triple mutant displays severe developmental defects, including main stem arrest and early maturation associated with aberrant lignin deposition in interfascicular tissues. In the triple mutant, key enzymes in the lignin biosynthesis pathway are upregulated, indicating that PWO1, TRB1, and TRB3 cooperatively regulate secondary cell wall formation. Together, our findings provide new insights into how PWO1 and TRBs cooperate to regulate chromatin states and orchestrate plant development, highlighting their central role in controlling gene expression programs. Significance statementThis study shows that PWO1 and TRB proteins co-occupy telomeres, including interspersed telo-box motifs, to regulate chromatin organization and plant development, particularly ectopic lignin deposition. Our findings reveal how these nuclear protein factors coordinate epigenetic states in Arabidopsis thaliana, providing a framework for understanding the control of developmental programs. O_FIG O_LINKSMALLFIG WIDTH=198 HEIGHT=200 SRC="FIGDIR/small/740627v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@1a3d544org.highwire.dtl.DTLVardef@1067082org.highwire.dtl.DTLVardef@1c49163org.highwire.dtl.DTLVardef@252171_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstract:C_FLOATNO Evolutionarily conserved PWO-TRB interactions and their shared roles in chromatin regulation and plant development. Created with BioRender.com. C_FIG

plant biology↗

bioRxiv2025

Molecular and Phenotypic Characterization of Telomere Repeat Binding (TRBs) Proteins in Moss: Evolutionary and Functional Perspectives

Telomere repeat binding (TRB) proteins are plant-specific proteins with a unique domain structure distinct from telomere-binding proteins in animals and yeast. While extensively studied in seed plants, their role in early-diverging plant lineages remain largely unexplored. Here, we investigate TRB proteins in a model moss, Physcomitrium patens, to assess their evolutionary conservation and functional significance. Functional analysis using single knockout mutants revealed that individual PpTRB genes are essential for normal development, with mutants exhibiting defects in the two-dimensional (protonemal) stage and, more prominently, in the formation of three-dimensional (gametophore) structures. Some double mutants displayed telomere shortening, a phenotype also observed in TRB-deficient seed plants, indicating a conserved role for TRBs in telomere maintenance. Transcriptome profiling of TRB mutants revealed altered expression of genes associated with transcriptional regulation and stimulus response in protonema. Subcellular localization studies across various plant cell types confirmed that PpTRBs, like their seed plant counterparts, localize prevalently to the plant nucleus and mutually interact. In bryophytes, TRBs form a monophyletic group that mirrors the species phylogeny, whereas in seed plants, TRBs have diversified into two distinct monophyletic groups. Our findings provide the first comprehensive characterization of TRB proteins in non-vascular plants and demonstrate their conserved roles in telomere maintenance, with additional implications for plant development and gene regulation across land plant lineages. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=117 SRC="FIGDIR/small/659030v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@1fea36dorg.highwire.dtl.DTLVardef@9b5569org.highwire.dtl.DTLVardef@488d0org.highwire.dtl.DTLVardef@1b22c6b_HPS_FORMAT_FIGEXP M_FIG C_FIG

plant biology↗

bioRxiv2025

SMG7 and eIF4A constitute a homeostatic module controlling P-body condensation and function of Meiotic bodies

Processing bodies (P-bodies) are ribonucleoprotein condensates that regulate RNA processing and storage. Although present constitutively in most cells, their size and composition dynamically change in response to developmental and environmental cues. However, mechanisms governing P-body assembly and remodelling remain poorly understood. Here we show that in Arabidopsis, SMG7 interacts with the eIF4A helicases and recruits them to P-bodies. eIF4As limit P-bodies condensation and also restrict stress granule (SG) formation under heat stress. We further identify meiotic (M-)bodies as composite RNP granules with a P-body core surrounded by a SG-like shell. The SMG7-eIF4A module regulates the recruitment of the meiosis-specific protein TDM1 into M-bodies, influencing meiotic exit and plant reproduction. Our findings suggest that SMG7 functions as an adaptor protein that recruits client proteins into P-bodies and, together with eIF4A, forms a regulatory module that regulates P-body composition and maintains their size homeostasis.

cell biology↗

bioRxiv2025

PWO proteins are associated with PRC2 since their emergence in vascular plants

PWWP-Domain Interactor of Polycombs 1 (PWO1), also known as PWWP1, interacts with the catalytic subunits of the Polycomb Repressive Complex 2 (PRC2) and, together with PWO2/3 proteins, plays a critical role in the development of Arabidopsis thaliana (At). PWOs are unique to plants and impact chromatin structure by enabling crosstalk between active and repressive epigenetic marks through mechanisms that are not yet fully understood. We aimed to understand the evolution of PWO proteins and whether their interaction with PRC2 has been conserved through evolution. Our study reveals that PWO proteins are present in vascular plants, but absent in bryophytes and green algae. The ancestral clade of PWO proteins includes the Selaginella moellendorffii (Sm) PWO orthologs SmPWOa and SmPWOb. Transient expression assays showed that both AtPWO1 and SmPWOa form nuclear speckles where they tether At, Sm, but also Physcomitrium patens (Pp) PRC2 catalytic subunits, despite the absence of PWO proteins in Pp. The PWO-PRC2 interactions were confirmed by protein-protein analyses. A newly identified evolutionarily conserved short C-terminal alpha-helix (c-motif) in PWO proteins contributes to an interaction interface for PWO-PRC2 binding. SmPWOs partially rescue the pwo1;pwo2 mutant phenotype in Arabidopsis, highlighting the functional conservation of PWOs in vascular plants.

plant biology↗

bioRxiv2024

Cytokinin acts as a systemic signal coordinating reproductive effort in Arabidopsis

The duration of the flowering phase is a key adaptive trait related to reproductive effort and plant longevity. In plants with indeterminate inflorescences, the number of flowers produced is determined by the activity of the inflorescence meristem. However, the integration of inflorescence meristem activity with the physiological and developmental status of a plant is not well understood. We combined quantitative imaging of the inflorescence meristem with the manipulation of cytokinin levels and distribution to investigate how meristem activity is regulated. Our data support the hypothesis that emerging fruits drain the acropetal flow of cytokinin away from the meristem inducing its proliferative arrest. We further show that the duration of flowering and cytokinin signaling in the inflorescence meristem is influenced by nitrate availability. Taken together, these data suggest that cytokinin functions as a systemic signal that integrates information about ongoing reproductive effort and plant nutrient status to control flower production.

plant biology↗

bioRxiv2024

Cytokinin-inducible DIRIGENT13 involved in lignan synthesis and ROS accumulation promotes root growth and abiotic stress tolerance in Arabidopsis

Dirigent (DIR) proteins mediate regio- and stereoselectivity of phenoxy radical coupling of monolignols for (neo)lignan or lignin synthesis. However, the role of DIR proteins and lignans in plant development and response to environmental stresses remains elusive. Here, we provide a functional characterization of the cytokinin-responsive DIRIGENT13 (DIR13) in Arabidopsis thaliana. DIR13 was localized to the root endodermis and at the margins of lateral root primordia and lateral roots. While not necessary for Casparian strip formation, DIR13 promoted both main and lateral root growth. Untargeted metabolomics and imaging mass spectrometry analyses unveiled the role of DIR13 in facilitating (neo)lignan synthesis in primary and lateral roots. Interestingly, DIR13 activated the production of putative oxomatairesinol and matairesinol-cysteine which are oxidative derivatives of the lignan matairesinol. Our data also identified DIR13 as an enhancer of salt and drought tolerance. Particularly, DIR13 attenuated salt stress-mediated inhibition of germination and root growth. Moreover, DIR13 activated reactive oxygen species (ROS) accumulation both under control and salt stress conditions, and cytokinin further enhanced the salt-induced ROS production specifically in the DIR13 overexpressing line. Our results uncover a role for DIR13-produced lignans in the priming of ROS accumulation, mediating both abiotic stress tolerance and the control of root architecture.

plant biology↗

bioRxiv2024

The molecular core of transcriptome responses to abiotic stress in plants: a machine learning-driven meta-analysis

Understanding how plants adapt their physiology to overcome severe stress conditions is vital in light of the current climate crisis. This remains a challenge given the complex nature of the underlying molecular mechanisms. To provide a full picture of stress mitigation mechanisms, an exhaustive analysis of publicly available stress-related transcriptomic data was conducted. We combined a meta-analysis with an unsupervised machine learning algorithm to identify a core of stress-related genes. To ensure robustness and biological significance of the output, often lacking in meta-analyses, a three-layered biovalidation was incorporated. Our results present a stress gene core, a set of key genes involved in plant tolerance to a multitude of adverse environmental conditions rather than specific ones. In addition, we provide a biologically validated database to assist in design of multi-stress resilience. Taken together, our results pave the way towards future-proof sustainable agriculture. TeaserUsing a machine learning-driven meta-analysis, a plant stress gene core was identified as a hub mediating multi-stress regulation

plant biology↗

bioRxiv2023

Primary multistep phosphorelay activation comprises both cytokinin and abiotic stress responses in Brassicaceae

Multistep phosphorelay (MSP) signaling integrates hormonal and environmental signals to control plant development and adaptive responses. The type-A RESPONSE REGULATORs (RRAs), the downstream members of the MSP cascade and cytokinin primary response genes, are supposed to mediate primarily the negative feedback regulation of (cytokinin-induced) MSP signaling. However, the transcriptional data suggest the involvement of RRAs in stress-related responses as well. By employing evolutionary conservation with the well-characterized Arabidopsis thaliana RRAs, we identified 5 and 38 novel putative RRAs in Brassica oleracea and Brassica napus, respectively. Our phylogenetic analysis suggests the existence of gene-specific selective pressure, maintaining the homologs of ARR3, ARR6, and ARR16 as singletons during the evolution of Brassica oleracea and Brassica rapa. We categorized RRAs based on the kinetics of their cytokinin-mediated upregulation and observed both similarities and specificities in this type of response across Brassicaceae. Using bioinformatic analysis and experimental data demonstrating the cytokinin responsiveness of Arabidopsis-derived TCSv2 reporter, we unveil the mechanistic conservation of cytokinin-mediated upregulation of RRAs in Brassica rapa and Brassica napus. Notably, we identify partial cytokinin dependency of cold stress-induced RRA transcription, thus corroborating the role of cytokinin signaling in the crop adaptive responses. HighlightsWe identified Brassica homologs of Arabidopsis type-A response regulators (RRAs), demonstrate existence of selective pressure preventing several RRAs multiplication during Brassicaceae evolution and describe cytokinin dependency of cold-induced RRAs upregulation.

plant biology↗

bioRxiv2023

Cytokinins control secondary cell wall formation in the inflorescence stem of Arabidopsis

Spatiotemporal control over developmental programs is vital to all organisms. Here we show that cytokinin (signaling) deficiency leads to early secondary cell wall (SCW) formation in Arabidopsis inflorescence stem that associates with precocious upregulation of a SCW transcriptional cascade controlled by NAC TFs (NSTs). We demonstrate that cytokinin signaling through the AHK2/3 and the ARR1/10/12 suppresses the expression of several NSTs and SCW formation in the apical portions of stems. Exogenous cytokinin application reconstituted both proper development and apical-basal gradient of NST1 and NST3 in a cytokinin biosynthesis-deficient mutant. We show that AHK2 and AHK3 required functional NST1 or NST3 to control SCW initiation in the interfascicular fibers, further evidencing that cytokinins act upstream of NSTs transcription factors. The premature onset of a rigid SCW biosynthesis and altered expression of NST1/3 and VND6/7 due to cytokinin deficiency led to the formation of smaller tracheary elements (TEs) and impaired hydraulic conductivity. We conclude that cytokinins downregulate NSTs to inhibit premature SCW formation in the apical part of the inflorescence stem, facilitating thus the development of fully functional TEs and interfascicular fibers. Summary statementCytokinins attenuate premature secondary cell wall (SCW) formation via downregulating the expression of NAC TFs, the master switches of SCW transcriptional cascade, thus affecting the tracheary elements size and conductivity.

plant biology↗

bioRxiv2023

Cytokinins regulate spatially-specific ethylene production to control root growth in Arabidopsis

The two principal growth regulators cytokinins and ethylene are known to interact in the regulation of plant growth. However, information about underlying molecular mechanism and positional specificity of the cytokinin/ethylene crosstalk in root growth control is scarce. We have identified spatial specificity of cytokinin-regulated root elongation and root apical meristem (RAM) size, both of which we demonstrate to be ethylene biosynthesis-dependent. Upregulation of the cytokinin biosynthetic gene ISOPENTENYLTRANSFERASE (IPT) in proximal and peripheral tissues leads to both root and RAM shortening. In contrast, IPT activation in distal and inner tissues reduces RAM size while leaving the root length comparable to mock-treated controls. We show that cytokinins regulate two steps specific to ethylene biosynthesis, the production of ACC by ACC SYNTHASEs (ACSs), and its conversion to ethylene by ACC OXIDASEs (ACOs). We describe cytokinin- and ethylene-specific regulation controlling the activity of ACSs and ACOs that are spatially discrete along both proximo/distal and radial root axes. Using direct ethylene measurements, we identify ACO2, ACO3 and ACO4 as being responsible for ethylene biosynthesis and the ethylene-regulated root and RAM shortening in cytokinin-treated Arabidopsis. Finally, we describe the tight cooperation between cytokinin and ethylene signaling in cytokinin-induced, ethylene-regulated control of ACO4 due to the direct interaction between ARABIDOPSIS RESPONSE REGULATOR 2 (ARR2), a member of the multistep phosphorelay cascade and the C-terminal portion of ETHYLENE INSENSITIVE 2 (EIN2-C), a key regulator of canonical ethylene signaling.

plant biology↗

bioRxiv2021

AHK5 mediates ETR1-initiated multistep phosphorelay in Arabidopsis

Plants, like other sessile organisms, need to sense many different signals, and in response to them, modify their developmental programs to be able to survive in a highly changing environment. The multistep phosphorelay (MSP) in plants is a good candidate for a response mechanism that integrates multiple signal types both environmental and intrinsic in origin. Recently, ethylene was shown to control MSP activity via the histidine kinase (HK) activity of ETHYLENE RESPONSE 1 (ETR1)1,2, but the underlying molecular mechanism still remains unclear. Here we show that although ETR1 is an active HK, its receiver domain (ETR1RD) is structurally and functionally unable to accept the phosphate from the phosphorylated His in the ETR1 HK domain (ETR1HK) to initiate the phosphorelay to ARABIDOPSIS HISTIDINE-CONTAINING PHOSPHOTRANSMITTERs (AHPs), the next link downstream members in MSP signaling. Instead, ETR1 interacts with another HK ARABIDOPSIS HISTIDINE KINASE 5 (AHK5) and transfers the phosphate from ETR1HK through the receiver domain of AHK5 (AHK5RD), and subsequently to AHP1, AHP2 and AHP3, independently of the HK activity of AHK5. We show that AHK5 is necessary for ethylene-initiated, but not cytokinin-initiated, MSP signaling in planta and that it thus mediates hormonal control of root growth.

plant biology↗

bioRxiv2020

Expansin-controlled cell wall stiffness regulates root growth in Arabidopsis

Expansins facilitate cell expansion via mediating pH-dependent cell wall (CW) loosening. However, the role of expansins in the control of biomechanical CW properties in the tissue and organ context remains elusive. We determined hormonal responsiveness and specificity of expression and localization of expansins predicted to be direct targets of cytokinin signalling. We found EXPA1 homogenously distributed throughout the CW of columella/ lateral root cap, while EXPA10 and EXPA14 localized predominantly at the three-cell boundaries of epidermis/cortex in various root zones. Cell type-specific localization of EXPA15 overlaps with higher CW stiffness measured via Brillouin light scattering microscopy. As indicated by both Brillouin frequency shift and AFM-measured Youngs modulus, EXPA1 overexpression upregulated CW stiffness, associated with shortening of the root apical meristem and root growth arrest. We propose that root growth in Arabidopsis requires delicate orchestration of biomechanical CW properties via tight regulation of various expansins localization to specific cell types and extracellular domains.

plant biology↗

bioRxiv2020

Mutually opposing activity of PIN7 splicing isoforms is required for auxin-mediated tropic responses in Arabidopsis thaliana

Advanced transcriptome sequencing has uncovered that the majority of eukaryotic genes undergo alternative splicing (AS). Nonetheless, little effort has been dedicated to investigating the functional relevance of particular splicing events, even those in the key developmental and hormonal regulators. Here we reveal, in the plant model Arabidopsis thaliana, that the PIN7 gene, which encodes a polarly localized transporter for the phytohormone auxin, produces two evolutionary-conserved transcripts. These isoforms PIN7a and PIN7b, differing in a 4 amino acid motif, are present at nearly equal levels in most cells. Although both variants do not differ in the subcellular localization and transport auxin with similar capacity, they closely associate and mutually influence their stability within the plasma membrane. Phenotypic complementation tests reveal that the functional contribution of PIN7b per se is minor but it markedly attenuates the prominent PIN7a activity, which is required for correct seedling apical hook formation and auxin-mediated tropic responses. These results establish alternative splicing of the PIN family as a conserved, functionally-relevant mechanism, unveiling an additional regulatory level of auxin-mediated plant development.

plant biology↗

bioRxiv2019

Cell-surface receptors enable perception of extracellular cytokinins

Cytokinins are mobile multifunctional plant hormones with roles in development and stress resilience 1,2. Although cytokinin receptors are substantially localised to the endoplasmic reticulum 3-5, the cellular sites of cytokinin perception continue to be debated 1,6,7. Several cytokinin types display bioactivity 8,9 and recently a cell-specific cytokinin gradient was reported in roots 10. Yet, the importance of spatially heterogeneous cytokinin distribution and the specific cytokinin(s) that account for the different responses remain unclear. Here we show that cytokinin perception by plasma membrane receptors is an effective path for cytokinin response in root cells. Readout from a Two Component Signalling cytokinin-specific reporter (TCSn::GFP;11) is closely matched to intracellular cytokinin content, yet a proportion of bioactive cytokinins are detected in the extracellular fluid. Using cytokinins covalently linked to beads that could not pass the plasma membrane, we demonstrate that strong TCSn activation still occurs and that this response is greatly diminished in cytokinin receptor mutants. Although intracellular receptors play significant roles, we argue for a revision of concepts of cytokinin perception to include the spatial dimensions. In particular, selective ligand-receptor affinities, cellular localisation and tissue distribution of bioactive cytokinins, their receptors, transporters and inactivation enzymes appear all to be components of the signalling regulatory mechanisms.

plant biology↗

bioRxiv2018

Cellulose biosynthesis inhibition reduces cell cycle activity in a nitrate reductase- and cytokinin-dependent manner

O_LIDuring growth, development and defense, cell wall integrity needs to be coordinated with cell cycle activity. In Saccharomyces cerevisiae, coordination is mediated by the cell wall integrity maintenance mechanism. In plants, little is known how coordination is achieved.\nC_LIO_LIHere we investigated coordination between plant cell wall and cell cycle activity in Arabidopsis thaliana seedlings by studying the impact of cell wall damage (CWD, caused by cellulose biosynthesis inhibition) on cell cycle gene expression, growth, phytohormone (jasmonic acid, salicylic acid, cytokinins) and lignin accumulation.\nC_LIO_LIWe found root growth and cell cycle gene expression are reduced by CWD in an osmo-sensitive manner. trans-zeatin application suppressed the CWD effect on gene expression. Quantification of cytokinins revealed CWD-induced, osmo-sensitive changes in several cytokinins. Expression of CYTOKININ OXIDASE2/DEHYDROGENASE (CKX2) and CKX3, encoding cytokinin-degrading enzymes, was elevated in CWD-exposed seedlings. Genetic studies implicated NITRATE REDUCTASE1/2 (NIA1/2) in the response to CWD. In nia1/2 seedlings CWD induced neither expression of CKX2/3 and cell cycle genes nor accumulation of jasmonic acid, salicylic acid and lignin.\nC_LIO_LIThis suggests that CWD causes increased CKX2/3 expression through a NIA1/2-mediated process. Increased CKX expression seems to cause changes in cytokinin levels, leading to reduced cell cycle gene expression.\nC_LI

plant biology↗