Biology subjects

Wrzaczek, M.

Publications and source records attributed to Wrzaczek, M..

18 recordsLinked to original sources

bioRxiv2026

The interactome of the receptor kinase THESEUS1 reveals how cell wall integrity is linked to hormone signalling in Arabidopsis

The receptor kinase THESEUS1 (THE1) activates responses to impairment of cell wall integrity (CWI). However, the interaction partners enabling THE1 to mediate the responses remain underexplored, impeding a mechanistic understanding of THE1. We produced a set of THE1 GFP fusion constructs in which THE1 is either wild type, mutated in its kinase domain, has a loss of function mutation in its ectodomain, or lacks the kinase or extracellular domain. We used these constructs to perform localization studies and immunoprecipitation liquid chromatography tandem mass spectrometry under control and stress conditions (isoxaben induced cell wall damage and sorbitol induced hyperosmotic stress) to identify the THE1 interactome. The interactome of THE1 changed in a context dependent manner, linking THE1 to known signaling components in CWI maintenance and phytohormone response (such as proton pumps, calcium pumps, other receptor kinases or cellulose synthesis regulators). Moreover, we identified novel interactors, associating THE1 with hitherto unexplored signaling pathways in response to CWI impairment. Our results also suggest that the THE1G37D protein (corresponding to the the1 1 allele) is retained in the endoplasmic reticulum, explaining its loss of function phenotype. The THE1 interactome and localization data provide the foundation to build a mechanistic understanding of THE1 mediated CWI maintenance.

plant biology↗

bioRxiv2026

The molecular architecture of the Arabidopsis callose synthase complex

ABSTRACTCallose synthase is responsible for the targeted deposition of the {beta}-1,3-glucan polymer, callose which underlines essential plant developmental processes, including cell division, pathogen defense or cell-cell communication. The architecture of the callose synthase complex (CALSC) as well as the molecular mechanisms of callose synthesis remain unknown. Here we report an integrative characterisation of the Arabidopsis CALS complex, with the most enriched subunits, CALS1, CALS2 and CALS3, forming its core. Structurally, CALSC assembles into a trimer, requiring the plant-specific Bag domain to mediate inter-subunit associations. The biological importance of CALSC assembly is highlighted by the simultaneous loss of CALS1 and CALS3, which abolishes plasmodesmal callose deposition and affects symplastic transport. Site-directed mutagenesis and molecular dynamics simulations depict the topology of the CALS1 active site in detail, including the components of the enzymatic reaction. We pinpoint the translocating tunnel through which the nascent glucan is delivered and mechanistically confirm the role of transmembrane helix 8 in regulating glucan export. Our work provides unprecedented insight into the molecular architecture of the CALSC and the distinct changes from maturation to activity at the plasma membrane, while showcasing the mechanisms involved in callose synthesis at the molecular level.

plant biology↗

bioRxiv2026

Unstructured regions differentially modulate the activation of RBOHD and RBOHH

Reactive oxygen species (ROS) produced by plant NADPH oxidases (RBOHs) must be precisely controlled in their concentration and spatial distribution to support diverse developmental and stress responses. RBOHs are activated by Ca2+ binding and phosphorylation, yet how internal regulatory domains within RBOHs have evolved to translate these inputs into precise levels of ROS production remains unclear. To address this, we performed phylogenetic analyses to define RBOH subfamilies and identify protein regions underlying functional diversification. This analysis revealed that the most variable regions across land-plant RBOHs are two unstructured regions in the N-terminus, UR1 and UR2, which flank the EF-hand Ca2+-binding domain (EFD). We dissected the roles of these regions in Arabidopsis RBOHD, which is central to plant immunity, and in RBOHH, which drives pollen tube elongation and exhibits high Ca2+-induced ROS production. Our analyses revealed that UR1 plays opposing roles in these RBOHs: in RBOHD, UR1 functions as an autoinhibitory module that restrains Ca2+-mediated activation, whereas in RBOHH, UR1 is essential for Ca2+-dependent activation and has coevolved with the EFD to maximize Ca2+-induced ROS production. We further uncovered divergent regulatory roles for UR2. In RBOHD, but not in RBOHH, phosphorylation of UR2 stabilizes an -helical conformation that promotes interaction with the catalytic domain required for enzymatic activation. Furthermore, unlike in RBOHH, the EFD of RBOHD has coevolved with UR2 to maximize phosphorylation-induced activity. Together, our results show how evolution of unstructured regulatory regions adapts a conserved enzymatic core to distinct demands of immune signaling and polarized growth.

plant biology↗

bioRxiv2026

GCN5 negatively regulates class III peroxidases PRX71 and PRX33 to promote lignin deposition in Arabidopsis

Histone acetylation shapes transcriptional programs during environmental stress. The Arabidopsis histone acetyltransferase GCN5/HAG1, a catalytic subunit of the SAGA complex, has been implicated in salt stress responses and cell wall integrity. Here, we show that loss of GCN5 enhances reactive oxygen species (ROS) accumulation under NaCl stress and is accompanied by altered expression of class III peroxidase genes, with strong salt-induced upregulation of PRX71 and elevated basal PRX33 transcript abundance. Consistent with a role for these peroxidases in stress-associated cell wall remodeling, overexpression of PRX71 or PRX33 in the wild-type is sufficient to promote ectopic lignin deposition in roots. Conversely, prx71 and prx33 mutants show improved growth under salt stress and on the cellulose biosynthesis inhibitor isoxaben, and they lack the pronounced ectopic root lignification observed in gcn5 under salt stress. Chromatin immunoprecipitation followed by qPCR (ChIP-qPCR) reveals reduced H3K9 acetylation at PRX71 and PRX33 promoter regions in gcn5 compared with the wild-type, and reduced transcript abundance of candidate upstream transcription factors (TFs), including GATA21 and MYBS2, accompanied by reduced H3K9ac at their loci. Together, our results support a model in which GCN5 constrains PRX71/PRX33-mediated lignification during stress, likely through an indirect regulatory route that integrates chromatin state and transcription factor activity to limit stress-associated lignification while maintaining root growth under salt stress.

plant biology↗

bioRxiv2025

Coordinated action of CRK2 and QSK1 regulate osmotic stress response in Arabidopsis

Precise control of intercellular communication is essential for normal growth and stress responses in all multicellular organisms. In Arabidopsis, two membrane-localized receptor like kinases (RLKs), the Cysteine-rich RLK CRK2 and the Leucine-rich repeat (LRR) RLK QSK1 relocalize from the general plasma membrane (PM) to plasmodesmata (PD) in response to osmotic stress. Both these RLKs regulate callose deposition thereby modulating PD permeability. However, unchecked callose deposition can block the PD and disrupt proper intercellular communication. Here, we show that under normal growth conditions, CRK2 phosphorylates and sequesters QSK1 at the general PM, preventing unnecessary callose deposition at PD. We show that osmotic stress-induced enrichment of QSK1 at PD requires functional CRK2 and establish that phosphorylation of QSK1 in its C-terminal region is inhibitory in this process. We propose that osmotic stress triggers dephosphorylation and release of QSK1 from the CRK2-QSK1 complex, enabling its relocalization from general PM to PD, where it promotes stress-induced callose deposition. Subsequently, CRK2 relocalizes to PD where it negatively influences callose deposition. Our work reveals a tightly coordinated distribution of QSK1 and CRK2 at PM, establishing a dynamic gating mechanism that balances growth and stress responsiveness.

plant biology↗

bioRxiv2025

CYSTEINE-RICH RLK2 regulates development via callose synthase-dependent symplastic transport in Arabidopsis

CYSTEINE-RICH RECEPTOR-LIKE PROTEIN KINASEs (CRKs) play an important role in plant development and stress responses. One of the best described members of the Arabidopsis CRK family is CRK2, which was proposed as a crucial regulator of intercellular transport facilitated by plasmodesmata (PD). As intercellular channels allowing symplastic communication, PD-mediated transport is predominantly regulated by callose synthase (CALS)-mediated callose deposition. This process can impact not just the distribution of molecules between adjacent cells, but also the symplastic loading of vascular tissue, thereby influencing plant stress responses and developmental processes. Here we described the overlapping expression pattern of genes encoding phylogenetically closely related CALS1 and CALS3. Both CALSs were phosphorylated in vitro by CRK2, and the genetic interaction between genes encoding CRK2 and CALS1 or CALS3 revealed their impact on callose deposition, rosette growth, primary root length, and development, represented as a decreased number of true leaves. Importantly, we observed significant accumulation of starch in crk2 mutant plants, especially in developmentally older leaves, which was reverted by the independent introduction of cals1.5 and cals3.1 into the crk2 mutant background. The observed starch accumulation was accompanied by photosynthesis inhibition. We propose that the growth and developmental alterations of crk2 are caused by decreased phloem loading, which resulted in starch accumulation in source organs, and subsequent sink tissue starvation. Our results propose CRK2 as negative regulator of CALS1 and CALS3 regulating source to sink transport, which impacts plant growth and development.

plant biology↗

bioRxiv2025

BBX transcription factor evolution in the green plant lineage

BBX transcription factors play versatile roles in integrating environment cues and regulating plant development. The model plant Arabidopsis continues to be a primary tool for the molecular investigation of protein function, but comprehensive evolutionary studies across different plant species can support translational research. Here, we constructed robust maximum likelihood phylogenies of full-length BBX proteins, the B-box domain, and the CCT domains by using diverse plant species ranging from algae to eudicots and monocots. Our analysis resolved five major structural groups (I-V) with BBXs from algae at the root of major clades. Our results suggest that Group II emerged after the split between Chlorophytes and Streptophytes. Our results also highlight subgroupings in the different BBXs clades. The evolution of the B-box domains showed a dynamic domain losses and acquisitions overall correlating with the BBX phylogeny. While the CCT domain was overall highly conserved across all groups, we identified secondary loss of the CCT domain in select lineages. We also detected significant expansions and contractions among angiosperm BBX orthogroups. Overall, the BBX family demonstrates ancient origins, structural plasticity via domain gain, loss and duplication, and lineage-specific innovations that underpin their diverse roles in light signaling, development and stress responses.

plant biology↗

bioRxiv2025

CRK2 modulates flowering in Arabidopsis together with GLYCINE-RICH RNA BINDING PROTEIN 7 (GRP7)

Timely flowering is critical for the reproductive success of plants. Plants adjust flowering time through integration of multiple internal and environmental cues. A complex signalling system lies behind this integration, including multiple proteins like the GLYCINE-RICH RNA-BINDING PROTEIN 7 (GRP7), linked to the modulation of stress and flowering in Arabidopsis. Available data show the interaction between GRP7 and the receptor-like kinase CYSTEINE-RICH RECEPTOR-LIKE KINASE 2 (CRK2). Like GRP7, CRK2 plays multifaceted roles in the coordination of stress responses and development, but its role in development remains unclear. Phenotypic analyses of the crk2 and grp7-1 single mutants as well as the crk2/grp7-1 double mutant supported the interaction between CRK2 and GRP7. Moreover, exogenous gibberellic acid did not recover the rosette phenotype of crk2 mutants but as in sensitive plants, inhibited GA-biosynthesis transcripts. The pseudo insensitivity of crk2 mutant was recovered in the crk2/grp7-1 line pointing to a common role of CRK2 and GRP7 in the modulation of gibberellin signalling. Together, CRK2 and GRP7 integrate hormonal and spatial signals required for floral transition. These insights highlight a previously unrecognized layer of regulation in GA signalling and open new avenues to understand how membrane-localized kinases and RNA-binding proteins intersect to control developmental timing.

plant biology↗

bioRxiv2025

Pattern-triggered immunity in blue and white seed cultivars of Papaver somniferum

Papaver somniferum (poppy) is a traditional component of Central and Eastern European cuisine and an important oilseed crop in the region. The crucial thread for poppy stable yield is pathogen infection. Thus, we need to understand poppy defence mechanisms in detail. The first robust layer of plant immunity, which plays a crucial role in combat against pathogens, is pattern-triggered immunity (PTI). Here, we provide the first insights into PTI in poppy. We selected four poppy varieties used in the food industry. We investigated poppy response to various peptide elicitors acting as microbe-associated molecular patterns (MAMPs) and damage-associated molecular patterns (DAMPs). Flg22 induced the most robust reactive oxygen species (ROS) burst among all tested peptides. Flg22 also triggered putative mitogen-activated protein kinase (MAPK) phosphorylation and seedling growth inhibition in all tested cultivars. We identified PsWRKY22 and PsPR2 as candidate marker genes suitable for monitoring poppy PTI responses. The tested poppy cultivars have low levels of salicylic acid. Callose accumulation was triggered by wounding but not by flg22. For studying PTI in poppy, wounding is a challenge that needs to be considered as it can obscure potential PTI responses. Our findings highlight conserved aspects of poppy immunity and challenges in studying poppy PTI. The established pipeline facilitates improving our understanding of poppy immunity and has the potential for widespread application in poppy breeding and improving selection for broad-spectrum disease resistance provided by enhanced PTI. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=147 SRC="FIGDIR/small/639761v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@1680d52org.highwire.dtl.DTLVardef@42895eorg.highwire.dtl.DTLVardef@488ffeorg.highwire.dtl.DTLVardef@1e5a95d_HPS_FORMAT_FIGEXP M_FIG Graphical abstract The establishment of the methods for studying pattern-triggered immunity (PTI) in Papaver somniferum L. (poppy) was inspired by the knowledge of the model plant Arabidopsis thaliana. The study showed a similarity between Arabidopsis and poppy in response to flg22 but also pointed out the obstacles for PTI analysis in poppy and the differences compared to the model plant. Created with BioRender.com. C_FIG

plant biology↗

bioRxiv2025

Calcium-triggered apoplastic ROS bursts balance gravity and mechanical signals to navigate soil

Reactive oxygen species (ROS) have been implicated repeatedly in multiple signaling processes in plants but the underlying mechanisms and roles remain enigmatic. Here, we developed live imaging of apoplastic ROS at the root surface. Different signals, including auxin, extracellular ATP and RALF1 peptide, all induce cytosolic calcium transients and apoplastic ROS bursts. Genetic and optogenetic manipulations identified calcium transients as necessary and sufficient for ROS bursts via activation of NADPH oxidases RBOHC and RBOHF. Apoplastic ROS bursts are not required but rather limit the gravity-induced root bending. Root bending is sensed by stretch-activated calcium channel MCA1 leading to NADPH oxidase activation at the stretched side. The resulting ROS production stiffens cell wall for better soil penetration. Apoplastic ROS thus provides a means to balance tissue flexibility and stiffness to efficiently navigate soil.

plant biology↗

bioRxiv2023

Cell-specialized chloroplast signaling orchestrates photosynthetic and extracellular reactive oxygen species for stress responses

Cellular responses to abiotic stress involve multiple signals including secondary messengers such as reactive oxygen species (ROS) and Ca2+, phytohormones such as abscisic acid (ABA) and chloroplast-to-nucleus retrograde signals such as 3-phosphoadenosine 5-phosphate (PAP). Mechanism(s) by which these messengers, produced in different subcellular compartments, intersect for cell regulation remain enigmatic. Previously we showed that the chloroplast retrograde signal PAP, similar to ABA, induces an increase in ROS levels in guard cells (Pornsiriwong et al, 2017). Here we demonstrate a mechanistic link enabling ABA and PAP to coordinate both chloroplast and plasma membrane ROS production. In whole leaves, PAP alters various ROS-related processes including plasmodesmal permeability as well as responses to ozone and the bacterial elicitor flg22, but mainly initiates processes that quench ROS during oxidative stress. Conversely, we show in guard cells, both PAP and ABA induce an increase in ROS levels in both chloroplasts via photosynthetic electron transport, and the apoplast via the RESPIRATORY BURST OXIDASE HOMOLOG (RBOH). Both subcellular ROS sources were necessary for ABA- and PAP-mediated stomatal closure. However, PAP signaling diverges from ABA by activating RBOHD, instead of RBOHF, for apoplastic ROS mediated stomatal closure. We identified three calcium-dependent protein kinases (CPKs) as the post-translational activators of RBOHD-mediated ROS production. CPK13, CPK32, and CPK34 were transcriptionally induced by PAP and concurrently activate RBOHD and the slow anion channel SLAC1 by phosphorylating two Serine (S) residues, including S120 which is also targeted by the core ABA signaling kinase OPEN STOMATA 1 (OST1). Consequently, overexpression of the PAP-induced CPKs rescues stomatal closure in ost1. Our data identify stomatal chloroplasts, to be nodes in the multifaceted cellular stress response networks as they are both sources and mediators of ROS and retrograde signals such PAP. Thus, chloroplasts are not just mediators of photosynthesis in response to, for example, excess light, but can serve as critical nodes in the multifaceted cellular stress response networks in specialized cells via retrograde signals, providing support to the concept of sensory plastids. Significance StatementThe chloroplast is an environmental sensor for stresses such as excess light and drought via the activation of photosynthetic-mediated retrograde signals. However, how does it function in specialized cells for which carbon fixation is secondary? Here we show the chloroplast is an important node to coordinate multiple plant signaling pathways in response to stresses such as drought. The chloroplast retrograde signal 3-phosphoadenosine 5-phosphate (PAP) plays multiple roles in reactive oxygen species (ROS) signaling and homeostasis. While PAP suppresses ROS in photosynthetic tissue, PAP instead induces guard cell ROS in chloroplasts and extracellular space to induce stomatal closure. We decipher how PAP-induced proteins activate both extracellular ROS production and anion channels for stomatal closure, thus providing a mechanism by which chloroplasts provide a strategic complement to canonical hormonal pathways in regulating plant physiological responses in specialized cells.

plant biology↗

bioRxiv2020

Arabidopsis Poly(ADP-ribose)-binding protein RCD1 interacts with Photoregulatory Protein Kinases in nuclear bodies

Continuous reprogramming of gene expression in response to environmental signals is required for plant survival in changing environment. One mechanism responsible for this is signaling through hub proteins that integrate external stimuli and transcriptional responses. RADICAL-INDUCED CELL DEATH1 (RCD1) functions as a nuclear hub protein, that interacts with a variety of transcription factors through its C-terminal RST domain and acts as a co-regulator of numerous stress responses in plants. Here, a previously unknown function for RCD1 as a novel plant poly(ADP-ribose) (PAR) reader protein is described. RCD1 localizes to specific locations inside the nucleus, in a PAR-dependent manner; its N-terminal WWE domain o binds PAR and together with the PARP-like domain determines its localization to nuclear bodies (NBs), which is prevented by inhibition of PAR synthesis. RCD1 also interacts with Photoregulatory Protein Kinases (PPKs) that co-localize with RCD1 in the NBs. The PPKs, that have been associated with circadian clock, abscisic acid, and light signaling pathways, phosphorylate RCD1 at multiple sites in the intrinsically disordered region between the WWE and PARP-like domains. This affects its stability and functions in the nucleus and1 provides a mechanism where the turnover of a PAR-binding transcriptional co-regulator is controlled by nuclear protein kinases.

plant biology↗

bioRxiv2019

Golgi anti-apoptotic proteins are evolutionarily conserved ion channels that regulate cell death in plants

Programmed cell death regulates developmental and stress responses in eukaryotes. Golgi anti-apoptotic proteins (GAAPs) are evolutionarily conserved cell death regulators. Human and viral GAAPs inhibit apoptosis and modulate intracellular Ca2+ fluxes, and viral GAAPs form cation-selective channels. Although most mammalian cell death regulators are not conserved at the sequence level in plants, the GAAP gene family shows expansion, with five paralogues (AtGAAP1-5) in the Arabidopsis genome. We pursued molecular and physiological characterization of AtGAAPs making use of the advanced knowledge of their human and viral counterparts. Structural modeling of AtGAAPs predicted the presence of a channel-like pore, and electrophysiological recordings from purified AtGAAP3 reconstituted into lipid bilayers confirmed that plant GAAPs can function as ion channels. AtGAAP1 and AtGAAP4 localized exclusively to the Golgi within the plant cell, while AtGAAP2, AtGAAP3 and AtGAAP5 also showed tonoplastic localization. Gene expression analysis revealed differential spatial expression and abundance of transcript for AtGAAP paralogues in Arabidopsis tissues. We demonstrate that AtGAAP1-5 inhibit Bax-induced cell death in yeast. However, overexpression of AtGAAP1 induces cell death in Nicotiana benthamiana leaves and lesion mimic phenotype in Arabidopsis. We propose that AtGAAPs function as Golgi-localized ion channels that regulate cell death by affecting ionic homeostasis within the cell. HighlightArabidopsis Golgi anti-apoptotic proteins (GAAPs) share functional conservation with their human and viral counterparts in cell death regulation and ion channel activity AbbreviationsAtGAAP, Arabidopsis thaliana GAAP; BI-1, Bax inhibitor-1; CFP, cyan fluorescent protein; CMLV, camelpox virus; ER, Endoplasmic reticulum; GAAP, Golgi anti-apoptotic protein; GFP, green fluorescent protein; hGAAP, human GAAP; LFG, Lifeguard; LMM, lesion mimic mutant; PCD, programmed cell death; TMBIM, transmembrane Bax inhibitor-1 motif-containing; TMDs, transmembrane domains; vGAAP, viral GAAP; YFP, yellow fluorescent protein

plant biology↗

bioRxiv2019

CRK2-mediated control of ROS production by phosphorylation of the RBOHD C-terminus in Arabidopsis

Reactive oxygen species (ROS) are important messengers in eukaryotic organisms and their production is tightly controlled. Active extracellular ROS production by NADPH oxidases in plants is triggered by receptor-like protein kinase (RLK)-dependent signaling networks. Here we show that the cysteine-rich RLK CRK2 kinase activity is required for plant growth and CRK2 exists in a preformed complex with the NADPH oxidase RBOHD in Arabidopsis. Functional CRK2 is required for the full elicitor-induced ROS burst and consequently the crk2 mutant is impaired in defense against the bacterial pathogen Pseudomonas syringae pv. tomato DC3000. Our work demonstrates that CRK2 regulates plant innate immunity. We identified in vitro CRK2-dependent phosphorylation sites in the C-terminal region of RBOHD. Phosphorylation of S703 RBOHD is enhanced upon flg22 treatment and substitution of S703 with alanine reduced ROS production in Arabidopsis. Phylogenetic analysis suggests that phospho-sites in C-terminal region of RBOHD are conserved throughout the plant lineage and between animals and plants. We propose that regulation of NADPH oxidase activity by phosphorylation of the C-terminal region might be an ancient mechanism and that CRK2 is an important element in regulating MAMP-triggered ROS production. One-sentence summaryCRK2 associates with and activates RBOHD to trigger MAMP-induced ROS production and reveals a novel regulatory mechanism for plant NADPH oxidases through phosphorylation of the C-terminus.

plant biology↗

bioRxiv2018

Mechanistic insights into the evolution of DUF26-containing proteins in land plants

Large protein families are a prominent feature of plant genomes and their size variation is a key element for adaptation in plants. Here we infer the evolutionary history of a representative protein family, the DOMAIN OF UNKNOWN FUNCTION (DUF) 26-containing proteins. The DUF26 first appeared in secreted proteins. Domain duplications and rearrangements led to the emergence of CYSTEINE-RICH RECEPTOR-LIKE PROTEIN KINASES (CRKs) and PLASMODESMATA-LOCALIZED PROTEINS (PDLPs). While the DUF26 itself is specific to land plants, structural analyses of Arabidopsis PDLP5 and PDLP8 ectodomains revealed strong similarity to fungal lectins. Therefore, we propose that DUF26-containing proteins constitute a novel group of plant carbohydrate-binding proteins. Following their appearance, CRKs expanded both through tandem duplications and preferential retention of duplicates in whole genome duplication events, whereas PDLPs evolved according to the dosage balance hypothesis. Based on our findings, we suggest that the main mechanism of expansion in new gene families is small-scale duplication, whereas genome fractionation and genetic drift after whole genome multiplications drive families towards dosage balance.

plant biology↗

bioRxiv2018

CRK2 enhances salt tolerance in Arabidopsis thaliana by regulating endocytosis and callose deposition in connection with PLDα1

High salinity has become an increasingly prevalent source of stress to which plants need to adapt. The receptor-like protein kinases (RLKs), including the cysteine-rich receptor-like kinase (CRK) subfamily, are a highly expanded family of transmembrane proteins in plants and are largely responsible for communication between cells and the extracellular environment. Various CRKs have been implicated in biotic and abiotic stress responses, however their functions on a cellular level remain largely uncharacterized. Here we have shown that CRK2 enhances salt tolerance at the germination stage in Arabidopsis thaliana. We identified CRK2 as a negative regulator of endocytosis, under both normal growth conditions and salt stress. We also established that functional CRK2 is required for salt-induced callose deposition. In doing so, we revealed a novel role for callose deposition, in response to increased salinity, and demonstrated its importance for salt tolerance during germination. Using fluorescently tagged proteins we observed specific changes in CRK2s subcellular localization in response to various stress treatments. Many of CRK2s cellular functions were dependent on phospholipase D (PLD) activity, as were the subcellular localization changes. Thus we propose that CRK2 acts downstream of PLD during salt stress to regulate endocytosis and promote callose deposition, and that CRK2 adopts specific stress-dependent subcellular localization patterns in order to carry out its functions.\n\nOne sentence summaryThe receptor-like kinase CRK2 acts in connection with PLD1 to regulate endocytosis and callose deposition at plasmodesmata, enhancing salt tolerance in Arabidopsis thaliana.

plant biology↗

bioRxiv2018

RCD1 Coordinates Chloroplastic and Mitochondrial Electron Transfer through Interaction with ANAC Transcription Factors

Signaling from chloroplasts and mitochondria, both dependent on reactive oxygen species (ROS), merge at the nuclear protein RADICAL-INDUCED CELL DEATH1 (RCD1). ROS produced in the chloroplasts affect the abundance, thiol redox state and oligomerization of RCD1. RCD1 directly interacts in vivo with ANAC013 and ANAC017 transcription factors, which are the mediators of the ROS-related mitochondrial complex III retrograde signa and suppresses activity of ANAC013 and ANAC017. Inactivation of RCD1 leads to increased expression of ANAC013 and ANAC017-regulated genes belonging to the mitochondrial dysfunction stimulon (MDS), including genes for mitochondrial alternative oxidases (AOXs). Accumulating AOXs and other MDS gene products alter electron transfer pathways in the chloroplasts, leading to diminished production of chloroplastic ROS and increased protection of photosynthetic apparatus from ROS damage. RCD1-dependent regulation affects chloroplastic and mitochondrial retrograde signaling including chloroplast signaling by 3-phosphoadenosine 5-phosphate (PAP). Sensitivity of RCD1 to organellar ROS provides feedback control of nuclear gene expression.

plant biology↗

bioRxiv2017

An Oomycete Effector Protein Induces Shade Avoidance In Arabidopsis And Attenuates Salicylate Signaling By Binding To Host Proteins Of The RADICAL-INDUCED CELL DEATH1 Family

The oomycete pathogen Hyaloperonospora arabidopsidis (Hpa) causes downy mildew disease on Arabidopsis. During infection, Hpa like other biotrophic pathogens, suppresses activation of plant innate immunity by translocating effector proteins into host cells. Some of these effectors localize to the host cell nucleus where they may manipulate transcriptional reprogramming of plant defense genes. Here we report that the nuclear-localized Hpa effector HaRxL106, when expressed in Arabidopsis, induces shade avoidance and attenuates the transcriptional response to the defense signaling molecule salicylic acid. HaRxL106 interacts with RADICAL-INDUCED CELL DEATH1 (RCD1) and loss of RCD1 function renders Arabidopsis resilient against HaRxL106-mediated suppression of immunity. To further characterize the molecular functions of RCD1 we solved a crystal structure of RCD1s Poly-(ADP-ribose)-Polymerase (PARP) domain and, based on non-conservation of amino acids constituting the active site of canonical PARPs, conclude that RCD1 has no PARP activity. We report that RCD1-type proteins are phosphorylated and identified histone-modifying Mut9-like kinases (MLKs) as RCD1-interacting proteins. A mlk1,3,4 triple mutant exhibits stronger SA-induced defense marker gene expression compared to wild-type plants. Our data suggest that HaRxL106 suppresses Arabidopsis innate immunity by manipulating the function(s) of RCD1 in the host cell nucleus and point towards a role of RCD1 as a transcriptional co-regulator that integrates signals from light and pathogen sensors.

plant biology↗