Phosphatidylinositol 5-Phosphate Links Dehydration Stress to the Activity of ARABIDOPSIS TRITHORAX-LIKE Factor ATX1

Changes in gene expression enable organisms to respond to environmental stress. Levels of cellular lipid second messengers, such as the phosphoinositide PtdIns5P, change in response to a variety of stresses and can modulate the localization, conformation and activity of a number of intracellular proteins. The plant trithorax factor (ATX1) tri-methylates the lysine 4 residue of histone H3 (H3K4me3) at gene coding sequences, which positively correlates with gene transcription. Microarray analysis has identified a target gene (WRKY70) that is regulated by both ATX1 and by the exogenous addition of PtdIns5P in Arabidopsis. Interestingly, ATX1 contains a PtdIns5P interaction domain (PHD finger) and thus, phosphoinositide signaling, may link environmental stress to changes in gene transcription.Using the plant Arabidopsis as a model system, we demonstrate a link between PtdIns5P and the activity of the chromatin modifier ATX1 in response to dehydration stress. We show for the first time that dehydration leads to an increase in cellular PtdIns5P in Arabidopsis. The Arabidopsis homolog of myotubularin (AtMTM1) is capable of generating PtdIns5P and here, we show that AtMTM1 is essential for the induced increase in PtdIns5P upon dehydration. Furthermore, we demonstrate that the ATX1-dependent gene, WRKY70, is downregulated during dehydration and that lowered transcript levels are accompanied by a drastic reduction in H3K4me3 of its nucleosomes. We follow changes in WRKY70 nucleosomal K4 methylation as a model to study ATX1 activity at chromatin during dehydration stress. We found that during dehydration stress, the physical presence of ATX1 at the WRKY70 locus was diminished and that ATX1 depletion resulted from it being retained in the cytoplasm when PtdIns5P was elevated. The PHD of ATX1 and catalytically active AtMTM1 are required for the cytoplasmic localization of ATX1.The novelty of the manuscript is in the discovery of a mechanistic link between a chromatin modifying activity (ATX1) and a lipid (PtdIns5P) synthesis in a signaling pathway that ultimately results in altered expression of ATX1 dependent genes downregulated in response to dehydration stress

Corrigendum: The \u3ci\u3eArabidopsis\u3c/i\u3e Homologs of Trithorax (ATX1) and Enhancer of Zeste (CLF) Establish ‘Bivalent Chromatin Marks’ at the Silent \u3ci\u3eAGAMOUS\u3c/i\u3e Locus

The authors wish to draw the attention to two irregularities in Figures 2a and 4. Both concern errors in duplicating images of empty lanes illustrating absence of DNA bands. We regret these omissions and apologize to readers for the inconvenience caused. The results and conclusions remain valid

The \u3ci\u3eArabidopsis\u3c/i\u3e Homologs of Trithorax (ATX1) and Enhancer of Zeste (CLF) Establish ‘Bivalent Chromatin Marks’ at the Silent \u3ci\u3eAGAMOUS\u3c/i\u3e Locus

Tightly balanced antagonism between the Polycomb group (PcG) and the Trithorax group (TrxG) complexes maintain Hox expression patterns in Drosophila and murine model systems. Factors belonging to the PcG/TrxG complexes control various processes in plants as well but whether they participate in mechanisms that antagonize, balance or maintain each other’s effects at a particular gene locus is unknown. CURLY LEAF (CLF), an Arabidopsis homolog of enhancer of zeste (EZ) and the ARABIDOPSIS HOMOLOG OF TRITHORAX (ATX1) control the expression of the flower homeotic gene AGAMOUS (AG). Disrupted ATX1 or CLF function results in misexpression of AG, recognizable phenotypes and loss of H3K4me3 or H3K27me3 histone H3-tail marks, respectively. A novel idea suggested by our results here, is that PcG and TrxG complexes function as a specific pair generating bivalent chromatin marks at the silent AG locus. Simultaneous loss of ATX1 and CLF restored AG repression and normalized leaf phenotypes. At the molecular level, disrupted ATX1 and CLF functions did not lead to erasure of the CLF and ATX1-generated epigenetic marks, as expected: instead, in the double mutants, H3K27me3 and H3K4me3 tags were partially restored. We demonstrate that ATX1 and CLF physically interact linking mechanistically the observed effects

A cytoplasm-specific activity encoded by the Trithorax-like ATX1 gene

Eukaryotes produce multiple products from a single gene locus by alternative splicing, translation or promoter usage as mechanisms expanding the complexity of their proteome. Trithorax proteins, including the Arabidopsis Trithorax-like protein ATX1, are histone modifiers regulating gene activity. Here, we report that a novel member of the Trithorax family has a role unrelated to chromatin. It is encoded from an internal promoter in the ATX1 locus as an isoform containing only the SET domain (soloSET). It is located exclusively in the cytoplasm and its substrate is the elongation factor 1A (EF1A). Loss of SET, but not of the histone modifying ATX1-SET activity, affects cytoskeletal actin bundling illustrating that the two isoforms have distinct functions in Arabidopsis cells

Divergent functions of the myotubularin (MTM) homologs AtMTM1 and AtMTM2 in \u3ci\u3eArabidopsis thaliana\u3c/i\u3e: evolution of the plant MTM family

Myotubularin and myotubularin-related proteins are evolutionarily conserved in eukaryotes. Defects in their function result in muscular dystrophy, neuronal diseases and leukemia in humans. In contrast to the animal lineage, where genes encoding both active and inactive myotubularins (phosphoinositide 3-phosphatases) have appeared and proliferated in the basal metazoan group, myotubularin genes are not found in the unicellular relatives of green plants. However, they are present in land plants encoding proteins highly similar to the active metazoan enzymes. Despite their remarkable structural conservation, plant and animal myotubularins have significantly diverged in their functions. While loss of myotubularin function causes severe disease phenotypes in humans it is not essential for the cellular homeostasis under normal conditions in Arabidopsis thaliana. Instead, myotubularin deficiency is associated with altered tolerance to dehydration stress. The two Arabidopsis genes AtMTM1 and AtMTM2 have originated from a segmental chromosomal duplication and encode catalytically active enzymes. However, only AtMTM1 is involved in elevating the cellular level of phosphatidylinositol 5-phosphate in response to dehydration stress, and the two myotubularins differentially affect the Arabidopsis dehydration stress-responding transcriptome. AtMTM1 and AtMTM2 display different localization patterns in the cell, consistent with the idea that they associate with different membranes to perform specific functions. A single amino acid mutation in AtMTM2 (L250W) results in a dramatic loss of subcellular localization. Mutations in this region are linked to disease conditions in humans

Divergent Functions of the Myotubularin (MTM) Homologs AtMTM1 and AtMTM2 in \u3ci\u3eArabidopsis thaliana\u3c/i\u3e: Evolution of the Plant MTM Family

Myotubularin and myotubularin-related proteins are evolutionarily conserved in eukaryotes. Defects in their function result in muscular dystrophy, neuronal diseases, and leukemia in humans. In contrast to the animal lineage, where genes encoding both active and inactive myotubularins (phosphoinositide 3-phosphatases) have appeared and proliferated in the basal metazoan group, myotubularin genes are not found in the unicellular relatives of green plants. However, they are present in land plants encoding proteins highly similar to the active metazoan enzymes. Despite their remarkable structural conservation, plant and animal myotubularins have significantly diverged in their functions. While loss of myotubularin function causes severe disease phenotypes in humans, it is not essential for the cellular homeostasis under normal conditions in Arabidopsis thaliana. Instead, myotubularin deficiency is associated with altered tolerance to dehydration stress. The two Arabidopsis genes AtMTM1 and AtMTM2 have originated from a segmental chromosomal duplication and encode catalytically active enzymes. However, only AtMTM1 is involved in elevating the cellular level of phosphatidylinositol 5-phosphate in response to dehydration stress, and the two myotubularins differentially affect the Arabidopsis dehydration stress-responding transcriptome. AtMTM1 and AtMTM2 display different localization patterns in the cell, consistent with the idea that they associate with different membranes to perform specific functions. A single amino acid mutation in AtMTM2 (L250W) results in a dramatic loss of subcellular localization. Mutations in this region are linked to disease conditions in humans

A cytoplasm-specific activity encoded by the Trithorax-like ATX1 gene

Eukaryotes produce multiple products from a single gene locus by alternative splicing, translation or promoter usage as mechanisms expanding the complexity of their proteome. Trithorax proteins, including the Arabidopsis Trithorax-like protein ATX1, are histone modifiers regulating gene activity. Here, we report that a novel member of the Trithorax family has a role unrelated to chromatin. It is encoded from an internal promoter in the ATX1 locus as an isoform containing only the SET domain (soloSET). It is located exclusively in the cytoplasm and its substrate is the elongation factor 1A (EF1A). Loss of SET, but not of the histone modifying ATX1-SET activity, affects cytoskeletal actin bundling illustrating that the two isoforms have distinct functions in Arabidopsis cells

Und seine Rolle in der Regulation des Jasmonsäure-Weges.

Salicylsäure (SA) ist ein Signalmolekül, welches in der Pflanze Abwehrreaktionen nach Pathogenbefall auslöst. Das cis-Element Activation sequence-1 (as-1) und die verwandten basic/leucin zipper (bZIP)-Typ Transkriptionsfaktoren der TGA-Familie regulieren die Expression in Abhängigkeit von SA und xenobiotischen Chemikalien. TGA-Faktoren interagieren mit NPR1 (NON EXPRESSOR OF PR GENES 1), einem zentralen Aktivator vieler SA-induzierter Abwehrreaktionen. Unter induzierten Bedingungen konnten Änderungen im Redox-Status von TGA1 und NPR1 nachgewiesen werden. Wenn erhöhte Mengen von SA und Jasmonsäure (JA) in der Zelle vorhanden sind, ist NPR1 außerdem an der Hemmung von (JA)-induzierbaren Genen beteiligt.Mit Hilfe eines Protein-Interaktions-Tests in Hefe mit NtTGA2.2 aus Tabak und einer cDNA-Bank aus Arabidopsis thaliana konnte ein Mitglied der Glutaredoxin Familie (GRX480; At1g28480) als mit TGA interagierendes Protein identifiziert werden. Außerdem konnte im Hefe-Dreihybridsystem ein GRX480/NtTGA2.2/NPR1 Komplex nachgewiesen werden. Glutaredoxine gelten als Kandidaten der Redox-Regulation in der Zelle, da sie in der Lage sind, Disulfid-Interaktionen zu katalysieren. In Protoplasten ist GRX480 im Zytosol sowie im Nukleus lokalisiert.In dieser Studie sollte der Einfluss der zwei katalytischen Cysteine von GRX480 auf die Interaktion mit AtTGA2 untersucht werden. Dafür wurden Hefe-Zweihybridsystem in Hefe und Tabak-Protoplasten durchgeführt. Es konnte gezeigt werden, dass das redox-defiziente GRX480 weiterhin mit AtTGA2 in Hefe interagiert, in Protoplasten jedoch nicht. Für die Interaktion ist der GRX480-spezifische N-Terminus nicht erforderlich. Des weiteren interagiert AtTGA2 nicht mit GRX370 (At5g40370).GRX480 wird verstärkt nach SA-Induktion oder Pathogenbefall exprimiert, wofür NPR1 und TGA Transkriptionsfaktoren notwendig sind. Eine Induktion durch JA konnte nicht nachgewiesen werden, obwohl JA in der Lage ist, die SA-abhängige Expression zweifach zu steigern.Arabidopsis Linien, welche GRX480 ektopisch exprimieren zeigen eine reduzierte Aktivität des verkürzten CaMV 35S Promotors, der an regulatorischen Elementen nur ein as-1 Element enthält. Dies wurde nicht beobachtet für die ektopische Expression von GRX370, was darauf hinweist, dass die Interaktion mit TGA Faktoren wichtig ist für diesen Effekt.Das JA-abhängige Abwehrgen PDF1.2 welches Bestandteil des SA/JA Antagonismus ist, wird ebenfalls durch GRX480 negativ reguliert, was ein Hinweis darauf ist, dass GRX480 ein Regulator dieses Wechselwirkung ist. GRX480 mit Cysteine-Serin-Austausch im aktiven Zentrum, zeigt diesen Effekt nicht. Epistatische Analysen zeigten, dass GRX480 unabhängig oder unterhalb von NPR1 wirkt. Das Vorhandensein des SA/JA Antagonismus in einer grx480 Verlustmutanten lässt vermuten, dass redundante Mechanismen in der Pflanze existieren, welche zur Hemmung der PDF1.2 Expression beitragen

Phosphatidylinositol 5-Phosphate Links Dehydration Stress to the Activity of \u3ci\u3eArabidopsis\u3c/i\u3e Trithorax-Like Factor ATX1

Background: Changes in gene expression enable organisms to respond to environmental stress. Levels of cellular lipid second messengers, such as the phosphoinositide PtdIns5P, change in response to a variety of stresses and can modulate the localization, conformation and activity of a number of intracellular proteins. The plant trithorax factor (ATX1) trimethylates the lysine 4 residue of histone H3 (H3K4me3) at gene coding sequences, which positively correlates with gene transcription. Microarray analysis has identified a target gene (WRKY70) that is regulated by both ATX1 and by the exogenous addition of PtdIns5P in Arabidopsis. Interestingly, ATX1 contains a PtdIns5P interaction domain (PHD finger) and thus, phosphoinositide signaling, may link environmental stress to changes in gene transcription. Principal Findings: Using the plant Arabidopsis as a model system, we demonstrate a link between PtdIns5P and the activity of the chromatin modifier ATX1 in response to dehydration stress. We show for the first time that dehydration leads to an increase in cellular PtdIns5P in Arabidopsis. The Arabidopsis homolog of myotubularin (AtMTM1) is capable of generating PtdIns5P and here, we show that AtMTM1 is essential for the induced increase in PtdIns5P upon dehydration. Furthermore, we demonstrate that the ATX1-dependent gene, WRKY70, is downregulated during dehydration and that lowered transcript levels are accompanied by a drastic reduction in H3K4me3 of its nucleosomes. We follow changes in WRKY70 nucleosomal K4 methylation as a model to study ATX1 activity at chromatin during dehydration stress. We found that during dehydration stress, the physical presence of ATX1 at the WRKY70 locus was diminished and that ATX1 depletion resulted from it being retained in the cytoplasm when PtdIns5P was elevated. The PHD of ATX1 and catalytically active AtMTM1 are required for the cytoplasmic localization of ATX1. Conclusions/Significance: The novelty of the manuscript is in the discovery of a mechanistic link between a chromatin modifying activity (ATX1) and a lipid (PtdIns5P) synthesis in a signaling pathway that ultimately results in altered expression of ATX1 dependent genes downregulated in response to dehydration stress