Salicylic acid (SA), a hormone essential for defense against biotrophic pathogens, triggers increased susceptibility of plants against necrotrophic attackers by suppressing the jasmonic acid-ethylene ...(ET) defense response. Here, we show that this diseasepromoting SA effect is abolished in plants lacking the three related TGACG sequence-specific binding proteins TGA2, TGA5, and TGA6 (class II TGAs). After treatment of plants with the ET precursor 1-aminocyclopropane-1-carboxylic acid (ACC), activation of all those genes that are suppressed by SA depended on class II TGAs. Rather than TGA binding sites, GCC-box motifs were significantly enriched in the corresponding promoters. GCC-box motifs are recognized by members of the superfamily of APETALA2/ETHYLENE RESPONSE FACTORS (ERFs). Of 11 activating ACC-induced APETALA2/ERFs, only ORA59 (for OCTADECANOID-RESPONSIVE ARABIDOPSIS APETALA2/ETHYLENE RESPONSE FACTOR domain protein59) and ERF96 were strongly suppressed by SA. ORA59 is the master regulator of the jasmonic acid-ET-induced defense program. ORA59 transcript levels do not reach maximal levels in the tga2 tga5 tga6 triple mutant, and this residual activity cannot be suppressed by SA. The ORA59 promoter contains an essential TGA binding site and is a direct target of class II TGAs as revealed by chromatin immunoprecipitation experiments. We suggest that class II TGAs at the ORA59 promoter constitute an important regulatory hub for the activation and SA suppression of ACC-induced genes.
The first exposure to light marks a crucial transition in plant development. This transition relies on the transcription factor HY5 controlling a complex downstream growth program. Despite its ...importance, its function in transcription remains unclear. Previous studies have generated lists of thousands of potential target genes and competing models of HY5 transcription regulation. In this work, we carry out detailed phenotypic and molecular analysis of constitutive activator and repressor HY5 fusion proteins. Using this strategy, we were able to filter out large numbers of genes that are unlikely to be direct targets, allowing us to eliminate several proposed models of HY5's mechanism of action. We demonstrate that the primary activity of HY5 is promoting transcription and that this function relies on other, likely light-regulated, factors. In addition, this approach reveals a molecular feedback loop via the COP1/SPA E3 ubiquitin ligase complex, suggesting a mechanism that maintains low HY5 in the dark, primed for rapid accumulation to reprogram growth upon light exposure. Our strategy is broadly adaptable to the study of transcription factor activity. Lastly, we show that modulating this feedback loop can generate significant phenotypic diversity in both Arabidopsis (
) and tomato (
).
Ankrd11 is a potential chromatin regulator implicated in neural development and autism spectrum disorder (ASD) with no known function in the brain. Here, we show that knockdown of Ankrd11 in ...developing murine or human cortical neural precursors caused decreased proliferation, reduced neurogenesis, and aberrant neuronal positioning. Similar cellular phenotypes and aberrant ASD-like behaviors were observed in Yoda mice carrying a point mutation in the Ankrd11 HDAC-binding domain. Consistent with a role for Ankrd11 in histone acetylation, Ankrd11 was associated with chromatin and colocalized with HDAC3, and expression and histone acetylation of Ankrd11 target genes were altered in Yoda neural precursors. Moreover, the Ankrd11 knockdown-mediated decrease in precursor proliferation was rescued by inhibiting histone acetyltransferase activity or expressing HDAC3. Thus, Ankrd11 is a crucial chromatin regulator that controls histone acetylation and gene expression during neural development, thereby providing a likely explanation for its association with cognitive dysfunction and ASD.
•Ankrd11 regulates proliferation and neurogenesis in the embryonic brain•Ankrd11 mutation causes cortical abnormalities and ASD-like behavior•Ankrd11 is associated with chromatin and colocalizes with HDAC3•Ankrd11 mutation alters histone acetylation and gene expression in neural precursors
Gallagher et al. identify the ASD-associated protein Ankrd11 as a transcriptional coregulator essential for embryonic cortical development. Ankrd11 regulates histone acetylation and gene expression and thus controls precursor proliferation and neuronal genesis and positioning. Ankrd11 mutant mice display abnormal cortical development and ASD-like behaviors, thereby modeling the human situation.
Mammalian brain development requires coordination between neural precursor proliferation, differentiation and cellular organization to create the intricate neuronal networks of the adult brain. Here, ...we examined the role of the atypical cadherins Fat1 and Fat4 in this process. We show that mutation of Fat1 in mouse embryos causes defects in cranial neural tube closure, accompanied by an increase in the proliferation of cortical precursors and altered apical junctions, with perturbations in apical constriction and actin accumulation. Similarly, knockdown of Fat1 in cortical precursors by in utero electroporation leads to overproliferation of radial glial precursors. Fat1 interacts genetically with the related cadherin Fat4 to regulate these processes. Proteomic analysis reveals that Fat1 and Fat4 bind different sets of actin-regulating and junctional proteins. In vitro data suggest that Fat1 and Fat4 form cis-heterodimers, providing a mechanism for bringing together their diverse interactors. We propose a model in which Fat1 and Fat4 binding coordinates distinct pathways at apical junctions to regulate neural progenitor proliferation, neural tube closure and apical constriction.
Mechanical stimuli, such as wind, rain, and touch affect plant development, growth, pest resistance, and ultimately reproductive success. Using water spray to simulate rain, we demonstrate that ...jasmonic acid (JA) signaling plays a key role in early geneexpression changes, well before it leads to developmental changes in flowering and plant architecture. The JA-activated transcription factors MYC2/MYC3/MYC4 modulate transiently induced expression of 266 genes, most of which peak within 30 min, and control 52% of genes induced >100-fold. Chromatin immunoprecipitationsequencing analysis indicates that MYC2 dynamically binds >1,300 promoters and trans-activation assays show that MYC2 activates these promoters. By mining our multiomic datasets, we identified a core MYC2/MYC3/MYC4-dependent “regulon” of 82 genes containing many previously unknown MYC2 targets, including transcription factors bHLH19 and ERF109. bHLH19 can in turn directly activate the ORA47 promoter, indicating that MYC2/MYC3/MYC4 initiate a hierarchical network of downstream transcription factors. Finally, we also reveal that rapid water spray-induced accumulation of JA and JA-isoleucine is directly controlled by MYC2/MYC3/MYC4 through a positive amplification loop that regulates JA-biosynthesis genes.
Glutaredoxins are small heat-stable oxidoreductases that transfer electrons from glutathione （GSH） to oxi- dized cysteine residues, thereby contributing to protein integrity and regulation. In ...Arabidopsis thaliana, floral glutare- doxins ROXY1 and ROXY2 and pathogen-induced ROXY19/GRX480 interact with bZIP transcription factors of the TGACG （TGA） motif-binding family. ROXY1, ROXY2, and TGA factors PERIANTHIA, TGA9, and TGA10 play essential roles in floral development. In contrast, ectopically expressed ROXY19/GRX480 negatively regulates expression of jasmonic acid （JA）/ ethylene （ET）-induced defense genes through an unknown mechanism that requires clade II transcription factors TGA2, TGA5, and/or TGA6. Here, we report that at least 17 of the 21 land plant-specific glutaredoxins encoded in the Arabidopsis genome interact with TGA2 in a yeast-two-hybrid system. To investigate their capacity to interfere with the expression of JA/ET-induced genes, we developed a transient expression system. Activation of the ORA59 （OCTADECANOID-RESPONSIVE ARABIDOPSIS AP2/ERF-domain protein 59） promoter by transcription factor EIN3 （ETHYLENE INSENSITVE 3） was sup- pressed by co-expressed ROXY19/GRX480. Suppression depended on the L＊＊LL motif in the C-terminus of ROXY19/ GRX480. This putative protein interaction domain was recently described as being essential for the TGA/ROXY interaction. Ten of the 17 tested ROXY proteins suppressed ORA59 promoter activity, which correlated with the presence of the C-terminal ALWL motif, which is essential for ROXY1 function in flower development. ROXY19/GRX480-mediated repres- sion depended on the GSH binding site, suggesting that redox modification of either TGA factors or as yet unknown target proteins is important for the suppression of ORA59 promoter activity.
The bacterium Agrobacterium tumefaciens has been the workhorse in plant genome engineering. Customized replacement of native tumor-inducing (Ti) plasmid elements enabled insertion of a sequence of ...interest called Transfer-DNA (T-DNA) into any plant genome. Although these transfer mechanisms are well understood, detailed understanding of structure and epigenomic status of insertion events was limited by current technologies. Here we applied two single-molecule technologies and analyzed Arabidopsis thaliana lines from three widely used T-DNA insertion collections (SALK, SAIL and WISC). Optical maps for four randomly selected T-DNA lines revealed between one and seven insertions/rearrangements, and the length of individual insertions from 27 to 236 kilobases. De novo nanopore sequencing-based assemblies for two segregating lines partially resolved T-DNA structures and revealed multiple translocations and exchange of chromosome arm ends. For the current TAIR10 reference genome, nanopore contigs corrected 83% of non-centromeric misassemblies. The unprecedented contiguous nucleotide-level resolution enabled an in-depth study of the epigenome at T-DNA insertion sites. SALK_059379 line T-DNA insertions were enriched for 24nt small interfering RNAs (siRNA) and dense cytosine DNA methylation, resulting in transgene silencing via the RNA-directed DNA methylation pathway. In contrast, SAIL_232 line T-DNA insertions are predominantly targeted by 21/22nt siRNAs, with DNA methylation and silencing limited to a reporter, but not the resistance gene. Additionally, we profiled the H3K4me3, H3K27me3 and H2A.Z chromatin environments around T-DNA insertions using ChIP-seq in SALK_059379, SAIL_232 and five additional T-DNA lines. We discovered various effect s ranging from complete loss of chromatin marks to the de novo incorporation of H2A.Z and trimethylation of H3K4 and H3K27 around the T-DNA integration sites. This study provides new insights into the structural impact of inserting foreign fragments into plant genomes and demonstrates the utility of state-of-the-art long-range sequencing technologies to rapidly identify unanticipated genomic changes.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Airway pressure is usually measured by sensors placed in the ventilator or on the ventilator side of the endotracheal tube (ETT), at the Y-piece. These remote measurements serve as a surrogate for ...the tracheal or alveolar pressure. Tracheal pressure can only be predicted correctly by using a model that incorporates the pressure at the remote location, the flow through the ETT, and the resistance of the ETT if the latter is a predictable function of Y-piece flow. However, this is not consistently appropriate, and accuracy of prediction is hampered.
This in vitro study systematically examined the ventilator pressure in dependence of compliance of the respiratory system (C
), inspiratory time, and expiratory time during pressure-controlled ventilation by using a small intratracheal pressure sensor and a mechanical lung simulator. Pressures were measured simultaneously at the ventilator outlet, at the Y-piece, and in the trachea during pressure-controlled ventilation with a peak inspiratory pressure of 20 cm H
O and a PEEP of 5 cm H
O while changing C
(10, 30, 60, 90, and 100 mL/cm H
O) and varying inspiratory time and expiratory time.
Tracheal pressures were always lower (maximum 8 cm H
O during inspiration) or higher (maximum 4 cm H
O during expiration) than the pressures measured proximal to the ETT if zero-flow conditions were not achieved at the end of the breathing cycles.
Dependent on C
and the breathing cycle, tracheal pressures deviated from those measured proximal to the ETT under non-zero-flow conditions. Intratracheal pressure and pressure curve dynamics can differ greatly from the ventilator pressure, depending on the ventilator setting and the C
. The small pressure sensor may be used as a measurement method of tracheal pressure via integration onto an ETT.
Understanding the systems-level actions of transcriptional responses to hormones provides insight into how the genome is reprogrammed in response to environmental stimuli. Here, we investigated the ...signalling pathway of the hormone jasmonic acid (JA), which controls a plethora of critically important processes in plants and is orchestrated by the transcription factor MYC2 and its closest relatives in Arabidopsis thaliana. We generated an integrated framework of the response to JA, which spans from the activity of master and secondary regulatory transcription factors, through gene expression outputs and alternative splicing, to protein abundance changes, protein phosphorylation and chromatin remodelling. We integrated time-series transcriptome analysis with (phospho)proteomic data to reconstruct gene regulatory network models. These enabled us to predict previously unknown points of crosstalk of JA to other signalling pathways and to identify new components of the JA regulatory mechanism, which we validated through targeted mutant analysis. These results provide a comprehensive understanding of how a plant hormone remodels cellular functions and plant behaviour, the general principles of which provide a framework for analyses of cross-regulation between other hormone and stress signalling pathways.
Summary
The three closely related Arabidopsis basic leucine zipper (bZIP) transcription factors TGA2, TGA5 and TGA6 are required for the establishment of the salicylic acid (SA)‐dependent plant ...defense response systemic acquired resistance, which is effective against biotrophic pathogens. Here we show that the same transcription factors are essential for the activation of jasmonic acid (JA)‐ and ethylene (ET)‐dependent defense mechanisms that counteract necrotrophic pathogens: the tga256 triple mutant is impaired in JA/ET‐induced PDF1.2 and b‐CHI expression, which correlates with a higher susceptibility against the necrotroph Botrytis cinerea. JA/ET induction of the trans‐activators ERF1 and ORA59, which act upstream of PDF1.2, was slightly increased (ERF1) or unaffected (ORA59). PDF1.2 expression can be restored in the tga256 mutant by increased expression of ORA59, as observed in the tga256 jin1 quadruple mutant, which lacks the transcription factor JIN1/AtMYC2 that functions as a negative regulator of the JA/ET‐dependent anti‐fungal defense program. Whereas JA/ET‐induced PDF1.2 expression is strongly suppressed by SA in wild‐type plants, no negative effect of SA on PDF1.2 expression was observed in the tga256 jin1 quadruple mutant. These results imply that the antagonistic effects of TGA factors and JIN1/AtMYC2 on the JA/ET pathway are necessary to evoke the SA‐mediated suppression of JA/ET‐induced defense responses.
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