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Salt and drought are documented among the most detrimental and persistent abiotic stresses for crop production. Here, we investigated the impact of Pseudomonas koreensis strain S4T10 on plant performance under salt and drought stress. Arabidopsis thaliana Col-0 wild type and atnced3 mutant plants were inoculated with P. koreensis or tap water and exposed to NaCl (100 mM) for five days and drought stress by withholding water for seven days. P. koreensis significantly enhanced plant biomass and photosynthetic pigments under salt and drought stress conditions. Moreover, P. koreensis activated the antioxidant defence by modulating glutathione (GSH), superoxide dismutase (SOD), peroxidase (POD), and polyphenol oxidase (PPO) activities to scavenge the reactive oxygen species produced due to the stress. In addition, the application of P. koreensis upregulated the expression of genes associated with antioxidant responses, such as AtCAT1, AtCAT3, and AtSOD. Similarly, genes linked to salt stress, such as AtSOS1, AtSOS2, AtSOS3, AtNHX1, and AtHKT1, were also upregulated, affirming the positive role of P. koreensis S4T10 in streamlining the cellular influx and efflux transport systems during salt stress. Likewise, the PGPB inoculation was observed to regulate the expression of drought-responsive genes AtDREB2A, AtDREB2B, and ABA-responsive genes AtAO3, AtABA3 indicating that S4T10 enhanced drought tolerance via modulation of the ABA pathway. The results of this study affirm that P. koreensis S4T10 could be further developed as a biofertilizer to mitigate salt and drought stress at the same time., (© 2024 The Authors. Physiologia Plantarum published by John Wiley & Sons Ltd on behalf of Scandinavian Plant Physiology Society.)

Chloroplast biogenesis is crucial in plant development, as it is essential for the transition to autotrophic growth. This process is light-induced and relies on the orchestrated transcription of nuclear and plastid genes, enabling the effective assembly and regulation of the photosynthetic machinery. Here we reveal a new regulation level for this process by showing the involvement of chromatin remodelling in the nuclear control of plastid gene expression for proper chloroplast biogenesis and function. The two Arabidopsis homologues of yeast EPL1 protein, components of the NuA4 histone acetyltransferase complex, are essential for plastid transcription and correct chloroplast development and performance. We show that EPL1 proteins are light-regulated and necessary for concerted expression of nuclear genes encoding most components of chloroplast transcriptional machinery, directly mediating H4K5ac deposition at these loci and promoting the expression of plastid genes required for chloroplast biogenesis. These data unveil a NuA4-mediated mechanism regulating chloroplast biogenesis that links the transcription of nuclear and plastid genomes during chloroplast development., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

Disease lesion mimic (DLM) or necrotic mutants display necrotic lesions in the absence of pathogen infections. They can show improved resistance to some pathogens and their molecular dissection can contribute to revealing components of plant defense pathways. Although forward-genetics strategies to find genes causal to mutant phenotypes are available in crops, these strategies require the production of experimental cross populations, mutagenesis, or gene editing and are time- and resource-consuming or may have to deal with regulated plant materials. In this study, we described a collection of 34 DLM mutants in barley (Hordeum vulgare L.) and applied a novel method called complementation by sequencing (CBS), which enables the identification of the gene responsible for a mutant phenotype given the availability of two or more chemically mutagenized individuals showing the same phenotype. Complementation by sequencing relies on the feasibility to obtain all induced mutations present in chemical mutants and on the low probability that different individuals share the same mutated genes. By CBS, we identified a cytochrome P450 CYP71P1 gene as responsible for orange blotch DLM mutants, including the historical barley nec3 locus. By comparative phylogenetic analysis we showed that CYP71P1 gene family emerged early in angiosperm evolution but has been recurrently lost in some lineages including Arabidopsis thaliana (L.) Heynh. Complementation by sequencing is a straightforward cost-effective approach to clone genes controlling phenotypes in a chemically mutagenized collection. The TILLMore (TM) collection will be instrumental for understanding the molecular basis of DLM phenotypes and to contribute knowledge about mechanisms of host-pathogen interaction., (© 2022 The Authors. The Plant Genome published by Wiley Periodicals LLC on behalf of Crop Science Society of America.)

Questions in developmental biology are most frequently addressed by using fluorescent markers of otherwise invisible cell states. In plants, such questions can be addressed most conveniently in leaves. Indeed, from the formation of stomata and trichomes within the leaf epidermis to that of vein networks deep into the leaf inner tissue, leaf cells and tissues differentiate anew during the development of each leaf. Moreover, leaves are produced in abundance and are easily accessible to visualization and perturbation. Yet a detailed procedure for the perturbation, dissection, mounting, and imaging of developing leaves has not been described. Here we address this limitation (1) by providing robust, step-by-step protocols for the local application of the plant hormone auxin to developing leaves and for the routine dissection and mounting of leaves and leaf primordia, and (2) by offering practical guidelines for the optimization of imaging parameters for confocal microscopy. We describe the procedure for the first leaves of Arabidopsis, but the same approach can be easily applied to other leaves of Arabidopsis or to leaves of other plants. © 2022 Wiley Periodicals LLC. Support Protocol 1: Preparation of plant growth medium Support Protocol 2: Preparation of growth medium plates Basic Protocol 1: Seed sterilization, sowing, and germination, and seedling growth Support Protocol 3: Preparation of IAA-lanolin paste Basic Protocol 2: Application of IAA-lanolin paste to 3.5-DAG first leaves Basic Protocol 3: Dissection of 3- to 6-DAG first leaves and leaf primordia Basic Protocol 4: Dissection of 1- and 2-DAG first-leaf primordia Basic Protocol 5: Mounting of dissected leaves and leaf primordia Support Protocol 4: Quality check of mounted leaves and leaf primordia by fluorescence microscopy Basic Protocol 6: Imaging of mounted leaves and leaf primordia by confocal microscopy., (© 2022 Wiley Periodicals LLC.)

In an ethyl methanesulfonate oat (Avena sativa) mutant population we have found a mutant with striking differences to the wild-type (WT) cv. Belinda. We phenotyped the mutant and compared it to the WT. The mutant was crossed to the WT and mapping-by-sequencing was performed on a pool of F2 individuals sharing the mutant phenotype, and variants were called. The impacts of the variants on genes present in the reference genome annotation were estimated. The mutant allele frequency distribution was combined with expression data to identify which among the affected genes was likely to cause the observed phenotype. A brassinosteroid sensitivity assay was performed to validate one of the identified candidates. A literature search was performed to identify homologs of genes known to be involved in seed shape from other species. The mutant had short kernels, compact spikelets, altered plant architecture, and was found to be insensitive to brassinosteroids when compared to the WT. The segregation of WT and mutant phenotypes in the F2 population was indicative of a recessive mutation of a single locus. The causal mutation was found to be one of 123 single-nucleotide polymorphisms (SNPs) spanning the entire chromosome 3A, with further filtering narrowing this down to six candidate genes. In-depth analysis of these candidate genes and the brassinosteroid sensitivity assay suggest that a Pro303Leu substitution in AVESA.00010b.r2.3AG0419820.1 could be the causal mutation of the short kernel mutant phenotype. We identified 298 oat proteins belonging to orthogroups of previously published seed shape genes, with AVESA.00010b.r2.3AG0419820.1 being the only of these affected by a SNP in the mutant. The AVESA.00010b.r2.3AG0419820.1 candidate is functionally annotated as a GSK3/SHAGGY-like kinase with homologs in Arabidopsis, wheat, barley, rice, and maize, with several of these proteins having known mutants giving rise to brassinosteroid insensitivity and shorter seeds. The substitution in AVESA.00010b.r2.3AG0419820.1 affects a residue with a known gain-of function substitution in Arabidopsis BRASSINOSTEROID-INSENSITIVE2. We propose a gain-of-function mutation in AVESA.00010b.r2.3AG0419820.1 as the most likely cause of the observed phenotype, and name the gene AsGSK2.1. The findings presented here provide potential targets for oat breeders, and a step on the way towards understanding brassinosteroid signaling, seed shape and nutrition in oats. [ABSTRACT FROM AUTHOR]

During leaf development, the timing of transition from cell proliferation to expansion is an important factor in determining the final organ size. However, the regulatory system involved in this transition remains less understood. To get an insight into this system, we investigated the compensation phenomenon, in which the cell number decreases while the cell size increases in organs with determinate growth. Compensation is observed in several plant species suggesting coordination between cell proliferation and expansion. In this study, we examined an Arabidopsis mutant of ANGUSTIFOLIA 3 (AN3)/GRFINTERACTING FACTOR 1, a positive regulator of cell proliferation, which exhibits the compensation. Though the AN3 role has been extensively investigated, the mechanism underlying excess cell expansion in the an3 mutant remains unknown. Focusing on the early stage of leaf development, we performed kinematic, cytological, biochemical, and transcriptome analyses, and found that the cell size had already increased during the proliferation phase, with active cell proliferation in the an3 mutant. Moreover, at this stage, chloroplasts, vacuoles, and xylem cells developed earlier than in the wild-type cells. Transcriptome data showed that photosynthetic activity and secondary cell wall biosynthesis were activated in an3 proliferating cells. These results indicated that precocious cell differentiation occurs in an3 cells. Therefore, we suggest a novel AN3 role in the suppression of cell expansion/differentiation during the cell proliferation phase. [ABSTRACT FROM AUTHOR]

The Elongator complex in eukaryotes has conserved tRNA modification functions and contributes to various physiological processes such as transcriptional control, DNA replication and repair, and chromatin accessibility. ARABIDOPSIS ELONGATOR PROTEIN 4 (AtELP4) is one of the six subunits (AtELP1-AtELP6) in Arabidopsis Elongator. In addition, there is an Elongator-associated protein, DEFORMED ROOTS AND LEAVES 1 (DRL1), whose homolog in yeast (Kti12) binds tRNAs. In this study, we explored the functions of AtELP4 in plant-specific aspects such as leaf morphogenesis and evolutionarily conserved ones between yeast and Arabidopsis. ELP4 comparison between yeast and Arabidopsis revealed that plant ELP4 possesses not only a highly conserved P-loop ATPase domain but also unknown plant-specific motifs. ELP4 function is partially conserved between Arabidopsis and yeast in the growth sensitivity toward caffeine and elevated cultivation temperature. Either single Atelp4 or drl1-102 mutants and double Atelp4 drl1-102 mutants exhibited a reduction in cell proliferation and changed the adaxial-abaxial polarity of leaves. In addition, the single Atelp4 and double Atelp4 drl1-102 mutants showed remarkable downward curling at the whole part of leaf blades in contrast to wild-type leaf blades. Furthermore, our genetic study revealed that AtELP4 might epistatically act on DRL1 in the regulation of cell proliferation and dorsoventral polarity in leaves. Taken together, we suggest that AtELP4 as part of the plant Elongator complex may act upstream of a regulatory pathway for adaxial-abaxial polarity and cell proliferation during leaf development. [ABSTRACT FROM AUTHOR]

ATP-Binding Cassette E (ABCE) proteins dissociate cytoplasmic ribosomes after translation terminates, and contribute to ribosome recycling, thus linking translation termination to initiation. This function has been demonstrated to be essential in animals, fungi, and archaea, but remains unexplored in plants. In most species, ABCE is encoded by a single-copy gene; by contrast, Arabidopsis thaliana has two ABCE paralogs, of which ABCE2 seems to conserve the ancestral function. We isolated apiculata7-1 (api7-1), the first viable, hypomorphic allele of ABCE2, which has a pleiotropic morphological phenotype reminiscent of mutations affecting ribosome biogenesis factors and ribosomal proteins. We also studied api7-2, a null, recessive lethal allele of ABCE2. Co-immunoprecipitation experiments showed that ABCE2 physically interacts with components of the translation machinery. An RNA-seq study of the api7-1 mutant showed increased responses to iron and sulfur starvation. We also found increased transcript levels of genes related to auxin signaling and metabolism. Our results support for the first time a conserved role for ABCE proteins in translation in plants, as previously shown for the animal, fungal, and archaeal lineages. In Arabidopsis, the ABCE2 protein seems important for general growth and vascular development, likely due to an indirect effect through auxin metabolism. [ABSTRACT FROM AUTHOR]

Overexpression of Arabidopsis dehydration response element binding 1a (DREB1a) is a well‐known approach for developing salinity, cold and/or drought stress tolerance. However, understanding of the genetic mechanisms associated with DREB1a expression in rice is generally limited. In this study, DREB1a‐associated early responses were investigated in a transgenic rice line harboring cold‐inducible DREB1a at a gene stacked locus. Although the function of other genes in the stacked locus was not relevant to stress tolerance, this study demonstrates DREB1a can be co‐localized with other genes for multigenic trait enhancement. As expected, the transgenic lines displayed improved tolerance to salinity stress and water withholding as compared with non‐transgenic controls. RNA sequencing and transcriptome analysis showed upregulation of complex transcriptional networks and metabolic reprogramming as DREB1a expression led to the upregulation of multiple transcription factor gene families, suppression of photosynthesis, and induction of secondary metabolism. In addition to the detection of previously described mechanisms such as production of protective molecules, potentially novel pathways were also revealed. These include jasmonate, auxin, and ethylene signaling, induction of JAZ and WRKY regulons, trehalose synthesis, and polyamine catabolism. These genes regulate various stress responses and ensure timely attenuation of the stress signal. Furthermore, genes associated with heat stress response were downregulated in DREB1a expressing lines, suggesting antagonism between heat and dehydration stress response pathways. In summary, through a complex transcriptional network, multiple stress signaling pathways are induced by DREB1a that presumably lead to early perception and prompt response toward stress tolerance as well as attenuation of the stress signal to prevent deleterious effects of the runoff response. [ABSTRACT FROM AUTHOR]

ARGONAUTE1 (AGO1) encodes a key component of the complexes mediating microRNA (miRNA) function in Arabidopsis. To study the regulation, action and interactions of AGO1, we conducted a genetic screen to identify second-site mutations modifying the morphological phenotype of ago1-52, a partial loss-of-function allele of AGO1. Unlike null ago1 mutations, the hypomorphic ago1-52 allele does not cause lethality or sterility; however, ago1-52 does produce a morphological phenotype clearly distinct from wild type. In our screen for modifiers of ago1-52, we identified suppressor mutations that partially restore wild-type morphology in the ago1-52 background and we termed these mas (morphology of argonaute1-52 suppressed). We focused on 23 of these putative suppressors. Linkage analysis of the mas mutations together with sequencing of the AGO1 gene in genomic DNA and cDNA from ago1-52 mas plants indicated that 22 of the mas lines contain extragenic suppressors, and one contains an intragenic suppressor that affects splicing of ago1-52. In the presence of the wild-type allele of AGO1, most of the mas mutations cause a mild or no mutant phenotype on their own, indicating that the ago1-52 mutant may provide a sensitized background for examining the interactions of AGO1. [ABSTRACT FROM AUTHOR]

The dynamic assembly of SKP1•CUL1•F-box protein (SCF) ubiquitin ligases is important for protein ubiquitination and degradation. This process is enabled by CAND1, which exchanges F-box proteins associated with the common CUL1 scaffold, and thereby, recycles the limited CUL1 core and allows diverse F-box proteins to assemble active SCFs. Previous human cell biological and computational studies have led to the adaptive exchange hypothesis, which suggests that the CAND1-mediated exchange confers plasticity on the SCF system, allowing cells to tolerate large variations in F-box protein expression. Here, we tested this hypothesis using Arabidopsis thaliana, a multicellular organism expressing hundreds of F-box protein genes at variable levels in different tissues. The cand1 null mutant in Arabidopsis is viable but produce almost no seeds. Bioinformatic, cell biological, and developmental analyses revealed that the low fertility in the cand1 mutant is associated with cell death in pollen, where the net expression of F-box protein genes is significantly higher than any other Arabidopsis tissue. In addition, we show that the transmission efficiency of the cand1 null allele was reduced through the male but not the female gametophyte. Our results suggest that CAND1 activity is essential in cells or tissues expressing high levels of F-box proteins. This finding is consistent with the proposed adaptive exchange hypothesis, demonstrating the necessity of the evolutionarily conserved CAND1-mediated exchange system in the development of a multicellular organism. [ABSTRACT FROM AUTHOR]

The submergence-induced hypoxic condition negatively affects the plant growth and development, and causes early onset of senescence. Hypoxia alters the expression of a number of microRNAs (miRNAs). However, the molecular function of submergence stress-induced miRNAs in physiological or developmental changes and recovery remains poorly understood. Here, we show that miR775 is an Arabidopsis thaliana-specific young and unique miRNA that possibly evolved non-canonically. miR775 post-transcriptionally regulates GALACTOSYLTRANSFERASE 9 (GALT9) and their expression is inversely affected at 24 h of complete submergence stress. The overexpression ofmiR775 (miR775-Oe) confers enhanced recovery from submergence stress and reduced accumulation of RBOHD and ROS, in contrast to wild-type and MIM775 Arabidopsis shoot. A similar recovery phenotype in the galt9 mutant indicates the role of the miR775-GALT9 module in post-submergence recovery.We predicted that Golgi-localized GALT9 is potentially involved in protein glycosylation. The altered expression of senescence-associated genes (SAG12, SAG29 and ORE1), ethylene signalling (EIN2 and EIN3) and abscisic acid (ABA) biosynthesis (NCED3) pathway genes occurs in miR775-Oe, galt9 and MIM775 plants. Thus, our results indicate the role for the miR775-GALT9 module in post-submergence recovery through a crosstalk between the ethylene signalling and ABA biosynthesis pathways. [ABSTRACT FROM AUTHOR]

The N 6-methyladenosine (m6A) pathway has been widely described as a viral regulatory mechanism in animals. We previously reported that the capsid protein (CP) of alfalfa mosaic virus (AMV) interacts with the Arabidopsis m6A demethylase ALKBH9B regulating m6A abundance on viral RNAs (vRNAs) and systemic invasion of floral stems. Here, we analyze the involvement of other ALKBH9 proteins in AMV infection and we carry out a detailed evaluation of the infection restraint observed in alkbh9b mutant plants. Thus, via viral titer quantification experiments and in situ hybridization assays, we define the viral cycle steps that are altered by the absence of the m6A demethylase ALKBH9B in Arabidopsis. We found that ALKBH9A and ALKBH9C do not regulate the AMV cycle, so ALKBH9B activity seems to be highly specific. We also define that not only systemic movement is affected by the absence of the demethylase, but also early stages of viral infection. Moreover, our findings suggest that viral upload into the phloem could be blocked in alkbh9b plants. Overall, our results point to ALKBH9B as a possible new component of phloem transport, at least for AMV, and as a potential target to obtain virus resistance crops. [ABSTRACT FROM AUTHOR]

SUMMARY: The NDC80 complex is a conserved eukaryotic complex composed of four subunits (NUF2, SPC25, NDC80, and SPC24). In yeast and animal cells, the complex is located at the outer layer of the kinetochore, connecting the inner layer of the kinetochore and spindle microtubules (MTs) during cell division. In higher plants, the relationship of the NDC80 complex with MTs is still unclear. In this study, we characterized the biological function of AtNUF2, a subunit of the Arabidopsis NDC80 complex. We found that AtNUF2 is widely expressed in various organs, especially in different stages of embryonic development. It was verified that AtNUF2 co‐localized with α‐tubulin on MTs during mitosis by immunohistochemical assays. Mutation of AtNUF2 led to severe mitotic defects, not only in the embryo and endosperm, but also in seedlings, resulting in seed abortion and stagnating seedling growth. Furthermore, the biological function of AtNUF2 was studied using partially complemented nuf2‐3/‐DD45;ABI3pro::AtNUF2 (nuf2‐3/‐DA) seedlings. The chromosome bridge and lagging chromatids occurred in nuf2‐3/‐DA root apical meristem cells, along with aberration of spindle MTs, resulting in blocked root growth. Meanwhile, the direct binding of AtNUF2 and AtSPC25 to MTs was determined by an MT co‐sedimentation assay in vitro. This study revealed the function of AtNUF2 in mitosis and the underlying mechanisms, modulating spindle MT organization and ensuring chromosome segregation during embryo, endosperm, and root development, laying the foundation for subsequent research of the NDC80 complex. Significance Statement: AtNUF2, a subunit of the Arabidopsis NDC80 complex, binds directly to AtSPC25, modulating spindle microtubule organization and chromosome segregation, thereby influencing mitosis. [ABSTRACT FROM AUTHOR]

Plant oxylipins are signaling molecules produced from fatty acids by oxidative pathways, mainly initiated by 9- and 13-lipoxygenases (9-LOX and 13-LOX), alpha-dioxygenases or non-enzymatic oxidation. Oxylipins from the 9-LOX pathway induce oxidative stress and control root development and plant defense. These activities have been associated with mitochondrial processes, but precise cellular targets and pathways remain unknown. In order to study oxylipin signaling, we previously generated a collection of Arabidopsis thaliana mutants that were insensitive to the 9-LOX products 9(S)-hydroxy-10,12, 15-octadecatrienoic acid (9-HOT) and its ketone derivative 9-KOT (noxy mutants). Here, we describe noxy1, noxy3, noxy5, noxy23 , and noxy54 mutants, all affected in nucleus-encoded mitochondrial proteins, and use them to study the role of mitochondria in oxylipin signaling. Functional and phenotypic analyses showed that noxy plants displayed mitochondrial aggregation, reduced respiration rates and resistance to the complex III inhibitor Antimycin A (AA), thus indicating a close similarity of the oxylipin signaling and mitochondrial stress. Application of 9-HOT and 9-KOT protected plants against subsequent mitochondrial stress, whereas they boosted root growth reduction when applied in combination with complex III inhibitors but did not with inhibitors of other respiratory complexes. A similar effect was caused by linear-chain oxylipins from 13-LOX or non-enzymatic pathways having α,β-unsaturated hydroxyl or keto groups in their structure. Studies to investigate 9-HOT and 9-KOT activity indicated that they do not reduce respiration rates, but their action is primarily associated with enhanced ROS responses. This was supported by the results showing that 9-HOT or 9-KOT combined with AA amplified the expression of oxylipin- and ROS-responding genes but not of the AA marker AOX1a , thus implying the activation of a specific mitochondria retrograde signaling pathway. Our results implicate mitochondrial complex III as a hub in the signaling activity of multiple oxylipin pathways and point at downstream ROS responses as components of oxylipin function. [ABSTRACT FROM AUTHOR]

The shoot apical meristem (SAM) is a reservoir of stem cells that gives rise to all post-embryonic above-ground plant organs. The size of the SAM remains stable over time owing to a precise balance of stem cell replenishment versus cell incorporation into organ primordia. The WUSCHEL (WUS)/CLAVATA (CLV) negative feedback loop is central to SAM size regulation. Its correct function depends on accurate spatial expression of WUS and CLV3. A signaling pathway, consisting of ERECTA family (ERf) receptors and EPIDERMAL PATTERNING FACTOR LIKE (EPFL) ligands, restricts SAM width and promotes leaf initiation. Although ERf receptors are expressed throughout the SAM, EPFL ligands are expressed in its periphery. Our genetic analysis of Arabidopsis demonstrated that ERfs and CLV3 synergistically regulate the size of the SAM, and wus is epistatic to ERf genes. Furthermore, activation of ERf signaling with exogenous EPFLs resulted in a rapid decrease of CLV3 andWUS expression. ERf-EPFL signaling inhibits expression of WUS and CLV3 in the periphery of the SAM, confining them to the center. These findings establish the molecular mechanism for stem cell positioning along the radial axis. [ABSTRACT FROM AUTHOR]

Abscisic acid (ABA) is an important phytohormone regulating plant growth, development and stress responses. A multitude of key factors implicated in ABA signaling have been identified; however, the regulation network of these factors needs for further information. AtS40.4, a plant-specific DUF584 domain-containing protein, was identified previously as a senescence regulator in Arabidopsis. In this study, our finding showed that AtS40.4 was negatively involved in ABA signaling during seed germination and early seedling growth. AtS40.4 was highly expressed in seeds and seedlings, and the expression level was promoted by ABA. AtS40.4 was localized both in the nucleus and the cytoplasm. Moreover, the subcellular localization pattern of AtS40.4 was affected by ABA. The knockdown mutants of AtS40.4 exhibited an increased sensitivity to ABA, whereas the overexpression of AtS40.4 decreased the ABA response during seed germination and seedling growth of Arabidopsis. Furthermore, AtS40.4 was involved in ABRE-dependent ABA signaling and influenced the expression levels of ABA INSENTIVE (ABI) 1-5 and SnRK2.6. Further genetic evidence demonstrated that AtS40.4 functioned upstream of ABI4. These findings support the notion that AtS40.4 is a novel negative regulator of the ABA response network during seed germination and early seedling growth. [ABSTRACT FROM AUTHOR]

Leaf colour is an important agronomic trait for studying molecular mechanisms in chlorophyll biosynthesis and chloroplast development. Here, a novel mutant oil yellow leaf 2 (Oy2) with a typical yellow leaf phenotype at the seedling stage was identified from the mutant population derived from the maize inbred line RP125. Compared with wild type, Oy2 mutant displays decreased chlorophyll content, reduced photosynthetic capacity and impaired chloroplast structure, which is likely controlled by a single recessive gene. The Oy2 locus was then delimited into a 117 kb region on chromosome 5 harbouring four genes, amongst which the gene Zm00001d013013, encoding a magnesium chelatase subunit D, was identified as the only candidate gene associated with Oy2 mutant phenotype. Moreover, the expression levels of candidate gene Oy2 and genes associated with chlorophyll biosynthesis and photosynthesis were tested by RNA‐seq and qRT‐PCR, implying that the causal gene Oy2 playing a critical role in chlorophyll synthesis. Taken together, we propose that the causal gene Oy2 highly associated with the yellow leaf phenotype may be helpful in elucidating photosynthetic pigments biosynthesis and chloroplast development in maize. [ABSTRACT FROM AUTHOR]

Mitochondrial transcription termination factors (mTERFs) are highly conserved proteins in metazoans. Plants have many more mTERF proteins than animals. The functions and the underlying mechanisms of plants' mTERFs remain largely unknown. In plants, mTERF family proteins are present in both mitochondria and plastids and are involved in gene expression in these organelles through different mechanisms. In this study, we screened Arabidopsis mutants with pigment-defective phenotypes and isolated a T-DNA insertion mutant exhibiting seedling-lethal and albino phenotypes [ seedling lethal 1 (sl1)]. The SL1 gene encodes an mTERF protein localized in the chloroplast stroma. The sl1 mutant showed severe defects in chloroplast development, photosystem assembly, and the accumulation of photosynthetic proteins. Furthermore, the transcript levels of some plastid-encoded proteins were significantly reduced in the mutant, suggesting that SL1/mTERF3 may function in the chloroplast gene expression. Indeed, SL1/mTERF3 interacted with PAP12/PTAC7, PAP5/PTAC12, and PAP7/PTAC14 in the subgroup of DNA/RNA metabolism in the plastid-encoded RNA polymerase (PEP) complex. Taken together, the characterization of the plant chloroplast mTERF protein, SL1/mTERF3, that associated with PEP complex proteins provided new insights into RNA transcription in the chloroplast. [ABSTRACT FROM AUTHOR]

Drought and salinity reduce seed germination, seedling emergence, and early seedling establishment, affect plant metabolism, and hence, reduce crop yield. Development of technologies that can increase plant tolerance of these challenging growth conditions is a major current interest among plant scientists and breeders. Seed priming has become established as one of the practical approaches that can alleviate the negative impact of many environmental stresses and improve the germination and overall performance of crops. Hormopriming using different plant growth regulators has been widely demonstrated as effective, but information about using cytokinins (CKs) as priming agents is limited to only a few studies using kinetin or 6-benzylaminopurine (BAP). Moreover, the mode of action of these compounds in improving seed and plant fitness through priming has not yet been studied. For many years, BAP has been one of the CKs most commonly applied exogenously to plants to delay senescence and reduce the impact of stress. However, rapid endogenous N 9-glucosylation of BAP can result in negative effects. This can be suppressed by hydroxylation of the benzyl ring or by appropriate N 9 purine substitution. Replacement of the 2′ or 3′ hydroxyl groups of a nucleoside with a fluorine atom has shown promising results in drug research and biochemistry as a means of enhancing biological activity and increasing chemical or metabolic stability. Here, we show that the application of this chemical modification in four new N 9-substituted CK derivatives with a fluorinated carbohydrate moiety improved the antisenescence properties of CKs. Besides, detailed phenotypical analysis of the growth and development of Arabidopsis plants primed with the new CK analogs over a broad concentration range and under various environmental conditions revealed that they improve growth regulation and antistress activity. Seed priming with, for example, 6-(3-hydroxybenzylamino)-2′-deoxy-2′-fluoro-9-(β)-D-arabinofuranosylpurine promoted plant growth under control conditions and alleviated the negative effects of the salt and osmotic stress. The mode of action of this hormopriming and its effect on plant metabolism were further analyzed through quantification of the endogenous levels of phytohormones such as CKs, auxins and abscisic acid, and the results are discussed. [ABSTRACT FROM AUTHOR]

Plant ribosomal proteins play universal roles in translation, although they are also involved in developmental processes and hormone signaling pathways. Among Arabidopsis RPL10 family members, RPL10A exhibits the highest expression during germination and early development, suggesting that RPL10A is the main contributor to these processes. In this work, we first analyzed RPL10A expression pattern in Arabidopsis thaliana using transgenic RPL10Apro:GUS plants. The gene exhibits a ubiquitous expression pattern throughout the plant, but it is most strongly expressed in undifferentiated tissues. Interestingly, gene expression was also detected in stomatal cells. We then examined protein function during seedling establishment and abscisic acid (ABA) response. Heterozygous rpl10A mutant plants show decreased ABA-sensitivity during seed germination, are impaired in early seedling and root development, and exhibit reduced ABA-inhibition of stomatal aperture under light conditions. Overexpression of RPL10A does not affect the germination and seedling growth, but RPL10A -overexpressing lines are more sensitive to ABA during early plant development and exhibit higher stomatal closure under light condition both with and without ABA treatment than wild type plants. Interestingly, RPL10A expression is induced by ABA. Together, we conclude that RPL10A could act as a positive regulator for ABA-dependent responses in Arabidopsis plants. [ABSTRACT FROM AUTHOR]

Drought stress adversely affects plant growth and development and significantly reduces crop productivity and yields. The phytohormone abscisic acid (ABA) rapidly accumulates in response to drought stress and mediates the expression of stress-responsive genes that help the plant to survive dehydration. The protein Powerdress (PWR), which interacts with Histone Deacetylase 9 (HDA9), has been identified as a critical component regulating plant growth and development, flowering time, floral determinacy, and leaf senescence. However, the role and function of PWR and HDA9 in abiotic stress response had remained elusive. Here we report that a complex of PWR and HDA9 interacts with ABI4 and epigenetically regulates drought signaling in plants. T-DNA insertion mutants of PWR and HDA9 are insensitive to ABA and hypersensitive to dehydration. Furthermore, the expression of ABA-responsive genes (RD29A , RD29B , and COR15A) is also downregulated in pwr and hda9 mutants. Yeast two-hybrid assays showed that PWR and HDA9 interact with ABI4. Transcript levels of genes that are normally repressed by ABI4, such as CYP707A1 , AOX1a and ACS4 , are increased in pwr. More importantly, during dehydration stress, PWR and HDA9 regulate the acetylation status of the CYP707A1 , which encodes a major enzyme of ABA catabolism. Taken together, our results indicate that PWR, in association with HDA9 and ABI4, regulates the chromatin modification of genes responsible for regulation of both the ABA-signaling and ABA-catabolism pathways in response to ABA and drought stress. [ABSTRACT FROM AUTHOR]

Patterning of the root xylem into protoxylem (PX) and metaxylem is regulated by auxin-cytokinin signaling and microRNA miR165a/166bmediated suppression of genes encoding Class III HOMEODOMAIN LEU-ZIPPER (HD-ZIPIII) proteins. We found that, in Arabidopsis, osmotic stress via core abscisic acid (ABA) signaling in meristematic endodermal cells induces differentiation of PX in radial and longitudinal axes in association with increased VND7 expression. Similarly, in tomato, ABA enhanced PX differentiation longitudinally and radially, indicating an evolutionarily conserved mechanism. ABA increased expression of miR165a/166b and reduced expression of the miR165a/166b repressor ZWILLE (ZLL) (also known as ARGONAUTE10), resulting in reduced levels of all five HD-ZIPIII RNAs. ABA treatments failed to induce additional PX files in a miR165a/166b-resistant PHB mutant, phb1-d, and in scr and shr mutants, in which miR165a/166b expression is strongly reduced. Thus, ABA regulates xylem patterning and maturation via miR165a/166b-regulated expression of HD-ZIPIII mRNAs and associated VND7 levels. In lateral root initials, ABA induced an increase in miR165a levels in endodermal precursors and inhibited their reduction in the future quiescent center specifically at preemergence stage. Hence, ABA-induced inhibition of lateral root is associated with reduced HD-ZIPIII levels. [ABSTRACT FROM AUTHOR]

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Summary: Underground roots normally reside in darkness. However, they are often exposed to ambient light that penetrates through cracks in the soil layers which can occur due to wind, heavy rain or temperature extremes. In response to light exposure, roots produce reactive oxygen species (ROS) which promote root growth. It is known that ROS‐induced growth promotion facilitates rapid escape of the roots from non‐natural light. Meanwhile, long‐term exposure of the roots to light elicits a ROS burst, which causes oxidative damage to cellular components, necessitating that cellular levels of ROS should be tightly regulated in the roots. Here we demonstrate that the red/far‐red light photoreceptor phytochrome B (phyB) stimulates the biosynthesis of abscisic acid (ABA) in the shoots, and notably the shoot‐derived ABA signals induce a peroxidase‐mediated ROS detoxification reaction in the roots. Accordingly, while ROS accumulate in the roots of the phyb mutant that exhibits reduced primary root growth in the light, such an accumulation of ROS did not occur in the dark‐grown phyb roots that exhibited normal growth. These observations indicate that mobile shoot‐to‐root ABA signaling links shoot phyB‐mediated light perception with root ROS homeostasis to help roots adapt to unfavorable light exposure. We propose that ABA‐mediated shoot‐to‐root phyB signaling contributes to the synchronization of shoot and root growth for optimal propagation and performance in plants. [ABSTRACT FROM AUTHOR]

Organ size regulation is dependent on the precise spatial and temporal regulation of cell proliferation and cell expansion. A number of transcription factors have been identified that play a key role in the determination of aerial lateral organ size, but their functional relationship to various chromatin modifiers has not been well understood. To understand how leaf size is regulated, we previously isolated the oligocellula1 (oli1) mutant of Arabidopsis thaliana that develops smaller first leaves than the wild type (WT) mainly due to a reduction in the cell number. In this study, we further characterized oli1 leaf phenotypes and identified the OLI1 gene as well as interaction partners of OLI1. Detailed characterizations of leaf development suggested that the cell proliferation rate in oli1 leaf primordia is lower than that in the WT. In addition, oli1 was associated with a slight delay of the progression from the juvenile to adult phases of leaf traits. A classical map-based approach demonstrated that OLI1 is identical to HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENES15 (HOS15). HOS15/OLI1 encodes a homolog of human transducin β-like protein1 (TBL1). TBL1 forms a transcriptional repression complex with the histone deacetylase (HDAC) HDAC3 and either nuclear receptor corepressor (N-CoR) or silencing mediator for retinoic acid and thyroid receptor (SMRT). We found that mutations in HISTONE DEACETYLASE9 (HDA9) and a switchingdefective protein 3, adaptor 2, N-CoR, and transcription factor IIIB-domain protein gene, POWERDRESS (PWR), showed a small-leaf phenotype similar to oli1. In addition, hda9 and pwr did not further enhance the oli1 small-leaf phenotype, suggesting that these three genes act in the same pathway. Yeast two-hybrid assays suggested physical interactions, wherein PWR probably bridges HOS15/OLI1 and HDA9. Earlier studies suggested the roles of HOS15, HDA9, and PWR in transcriptional repression. Consistently, transcriptome analyses showed several genes commonly upregulated in the three mutants. From these findings, we propose a possibility that HOS15/OLI1, PWR, and HDA9 form an evolutionary conserved transcription repression complex that plays a positive role in the regulation of final leaf size. [ABSTRACT FROM AUTHOR]

Leaf abaxial-adaxial patterning is dependent on the mutual repression of leaf polarity genes expressed either adaxially or abaxially. In Arabidopsis thaliana, this process is strongly affected by mutations in ribosomal protein genes and in ribosome biogenesis genes in a sensitized genetic background, such as asymmetric leaves2 (as2). Most ribosome-related mutants by themselves do not show leaf abaxialization, and one of their typical phenotypes is the formation of pointed rather than rounded leaves. In this study, we characterized two ribosome-related mutants to understand how ribosome biogenesis is linked to several aspects of leaf development. Previously, we isolated oligocellula2 (oli2) which exhibits the pointed-leaf phenotype and has a cell proliferation defect. OLI2 encodes a homolog of Nop2 in Saccharomyces cerevisiae, a ribosome biogenesis factor involved in pre-60S subunit maturation. In this study, we found another pointed-leaf mutant that carries a mutation in a gene encoding an uncharacterized protein with a G-patch domain. Similar to oli2, this mutant, named g-patch domain protein1 (gdp1), has a reduced number of leaf cells. In addition, gdp1 oli2 double mutants showed a strong genetic interaction such that they synergistically impaired cell proliferation in leaves and produced markedly larger cells. On the other hand, they showed additive phenotypes when combined with several known ribosomal protein mutants. Furthermore, these mutants have a defect in pre-rRNA processing. GDP1 and OLI2 are strongly expressed in tissues with high cell proliferation activity, and GDP1-GFP and GFP-OLI2 are localized in the nucleolus. These results suggest that OLI2 and GDP1 are involved in ribosome biogenesis. We then examined the effects of gdp1 and oli2 on adaxial-abaxial patterning by crossing them with as2. Interestingly, neither gdp1 nor oli2 strongly enhanced the leaf polarity defect of as2. Similar results were obtained with as2 gdp1 oli2 triple mutants although they showed severe growth defects. These results suggest that the leaf abaxialization phenotype induced by ribosome-related mutations is not merely the result of a general growth defect and that there may be a sensitive process in the ribosome biogenesis pathway that affects adaxial-abaxial patterning when compromised by a mutation. [ABSTRACT FROM AUTHOR]

Mitochondria are the powerhouse of eukaryotic cells because they are responsible for energy production through the aerobic respiration required for growth and development. These organelles harbour their own genomes and translational apparatus: mitochondrial ribosomes or mitoribosomes. Deficient mitochondrial translation would impair the activity of this organelle, and is expected to severely perturb different biological processes of eukaryotic organisms. In plants, mitoribosomes consist of three rRNA molecules, encoded by the mitochondrial genome, and an undefined set of ribosomal proteins (mitoRPs), encoded by nuclear and organelle genomes. A detailed functional and structural characterisation of the mitochondrial translation apparatus in plants is currently lacking. In some plant species, presence of small gene families of mitoRPs whose members have functionally diverged has led to the proposal of the heterogeneity of the mitoribosomes. This hypothesis supports a dynamic composition of the mitoribosomes. Information on the effects of the impaired function of mitoRPs on plant development is extremely scarce. Nonetheless, several works have recently reported the phenotypic and molecular characterisation of plant mutants affected in mitoRPs that exhibit alterations in specific development aspects, such as embryogenesis, leaf morphogenesis or the formation of reproductive tissues. Some of these results would be in line with the ribosomal filter hypothesis, which proposes that ribosomes, besides being the machinery responsible for performing translation, are also able to regulate gene expression. This review describes the phenotypic effects on plant development displayed by the mutants characterised to date that are defective in genes which encode mitoRPs. The elucidation of plant mitoRPs functions will provide a better understanding of the mechanisms that control organelle gene expression and their contribution to plant growth and morphogenesis. [ABSTRACT FROM AUTHOR]

To ensure successful plant reproduction and crop production, the spatial and temporal control of the termination of the floral meristem must be coordinated. In Arabidopsis, the timing of this termination is determined by AGAMOUS (AG). Following its termination, the floral meristem underdoes gynoecium formation. A direct target of AG, CRABS CLAW (CRC), is involved in both floral meristem determinacy and gynoecium development. However, how floral meristem termination is coordinated with gynoecium formation is not understood. Here, we identify a mechanistic link between floral meristem termination and gynoecium development through fine-tuning of auxin homeostasis by CRC. CRC controls auxin homeostasis in the medial region of the developing gynoecium to generate proper auxin maxima. This regulation partially occurs via direct transcriptional repression of TORNADO2 (TRN2) by CRC. Plasma membrane-localized TRN2 modulates auxin homeostasis. We propose a model describing how regulation of auxin homeostasis mediates the transition from floral meristem termination to gynoecium development. [ABSTRACT FROM AUTHOR]

AtLSG1-2 is a circularly permuted GTPase required for ribosome biogenesis and recently shown to be involved in early leaf development, although it was unclear how AtLSG1-2 affects leaf growth. Here, we found that atlsg1-2 mutants had reduced leaf size as a result of decreased cell size and cell number. Leaf kinematic analysis and CYCB1;1::GUS expression pattern in atlsg1-2 mutant indicated that loss of function of AtLSG1-2 delays the transition from cell division to cell expansion. Decreases in ploidy levels and trichome branch number suggest that AtLSG1-2 deficiency suppresses endoreduplication. Real-time PCR analysis showed that genes specifically expressed in the proliferation stage were highly expressed and those involved in endoreduplication were differentially regulated. LSG1 is known to mediate the recruitment of nucleocytoplasmic shuttling protein NMD3 back to the nucleus in yeast, yet their relationship was unclear in plants. Our genetic analysis revealed that the atlsg1 atnmd3 double mutant displayed enhanced phenotypes as compared with the respective single mutant and that AtLSG1-2 and AtNMD3 synergistically affect the cell proliferation process. [ABSTRACT FROM AUTHOR]

Polycomb group proteins form multiple protein complexes such as Polycomb Repressive Complex (PRC) 1 and PRC2, which repress the expression of thousands of genes. PRC1 and PRC2 are essential for normal development in Arabidopsis. Recently, significant progress has been made in understanding the functions and regulatory mechanisms of PRC1. In this review, we focus on the discovery of the composition of PRC1, functions of its components, the recruitment of PRC1 to target genes and the control of PRC1 function in Arabidopsis. Perspectives on dissecting the roles of PRC1 in plant gene expression and development are also given. [ABSTRACT FROM AUTHOR]

A major challenge in morphometrics is to analyse complex biological shapes formed by structures at different scales. Leaves exemplify this challenge as they combine differences in their overall shape with smaller shape variations at their margin, leading to lobes or teeth. Current methods based on contour or on landmark analysis are successful in quantifying either overall leaf shape or leaf margin dissection, but fail in combining the two. Here, we present a comprehensive strategy and its associated freely available platform for the quantitative, multiscale analysis of the morphology of leaves with different architectures. For this, biologically relevant landmarks are automatically extracted and hierarchised, and used to guide the reconstruction of accurate average contours that properly represent both global and local features. Using this method, we establish a quantitative framework of the developmental trajectory of Arabidopsis leaves of different ranks and retrace the origin of leaf heteroblasty. When applied to different mutant forms, our method can contribute to a better understanding of gene function, as we show here for the role of CUC2 during Arabidopsis leaf serration. Finally, we illustrate the wider applicability of our tool by analysing hand morphometrics. [ABSTRACT FROM AUTHOR]

Abscisic acid (ABA) is a key phytohormone that regulates plant growth and developmental processes, including seed germination and stomatal closing. Here, we report the identification and functional characterization of a novel type 2C protein phosphatase, CaADIP1 ( Capsicum annuum ABA and Drought- Induced Protein phosphatase 1). The expression of CaADIP1 was induced in pepper leaves by ABA, drought and NaCl treatments. Arabidopsis plants overexpressing CaADIP1 ( CaADIP1-OX) exhibited an ABA-hyposensitive and drought-susceptible phenotype. We used a yeast two-hybrid screening assay to identify CaRLP1 ( Capsicum annuum RCAR- Like Protein 1), which interacts with CaADIP1 in the cytoplasm and nucleus. In contrast to CaADIP1-OX plants, CaRLP1-OX plants displayed an ABA-hypersensitive and drought-tolerant phenotype, which was characterized by low levels of transpirational water loss and increased expression of stress-responsive genes relative to those of wild-type plants. In CaADIP1-OX/ CaRLP1-OX double transgenic plants, ectopic expression of the CaRLP1 gene led to strong suppression of CaADIP1-induced ABA hyposensitivity during the germinative and post-germinative stages, indicating that CaADIP1 and CaRLP1 act in the same signalling pathway and CaADIP1 functions downstream of CaRLP1. Our results indicate that CaADIP1 and its interacting partner CaRLP1 antagonistically regulate the ABA-dependent defense signalling response to drought stress. [ABSTRACT FROM AUTHOR]

When plants grow in close proximity basic resources such as light can become limiting. Under such conditions plants respond to anticipate and/or adapt to the light shortage, a process known as the shade avoidance syndrome (SAS). Following genetic screening using a shade-responsive luciferase reporter line (PHYB:LUC), we identified DRACULA2 (DRA2), which encodes an Arabidopsis homolog of mammalian nucleoporin 98, a component of the nuclear pore complex (NPC). DRA2, together with other nucleoporins, participates positively in the control of the hypocotyl elongation response to plant proximity, a role that can be considered dependent on the nucleocytoplasmic transport of macromolecules (i.e. is transport dependent). In addition, our results reveal a specific role for DRA2 in controlling shade-induced gene expression. We suggest that this novel regulatory role of DRA2 is transport independent and that it might rely on its dynamic localization within and outside of the NPC. These results provide mechanistic insights in to how SAS responses are rapidly established by light conditions. They also indicate that nucleoporins have an active role in plant signaling. [ABSTRACT FROM AUTHOR]

Efficient use of seed nutrient reserves is crucial for germination and establishment of plant seedlings. Mobilizing seed oil reserves in Arabidopsis involves b-oxidation, the glyoxylate cycle, and gluconeogenesis, which provide essential energy and the carbon skeletons needed to sustain seedling growth until photoautotrophy is acquired. We demonstrated that HC-PPase activity is required for gluconeogenesis. Lack of HC- PPase in fugu5 mutants increases cytosolic pyrophosphate (PPi) levels, which partially reduces sucrose synthesis de novo and inhibits cell division. In contrast, post-mitotic cell expansion in cotyledons was unusually enhanced, a phenotype called compensation. Therefore, it appears that PPi inhibits several cellular functions, including cell cycling, to trigger compensated cell enlargement (CCE). Here, we mutagenized fugu5-1 seeds with 12C6+ heavy-ion irradiation and screened mutations that restrain CCE to gain insight into the genetic pathway(s) involved in CCE. We isolated A#3-1, in which cell size was severely reduced, but cell number remained similar to that of original fugu5-1. Moreover, cell number decreased in A#3-1 single mutant (A#3-1sm), similar to that of fugu5-1, but cell size was almost equal to that of the wild type. Surprisingly, A#3-1 mutation did not affect CCE in other compensation exhibiting mutant backgrounds, such as an3-4 and fugu2-1/fas1-6. Subsequent map-based cloning combined with genome sequencing and HRM curve analysis identified enoyl-CoA hydratase 2 (ECH2) as the causal gene of A#3-1. The above phenotypes were consistently observed in the ech2-1 allele and supplying sucrose restored the morphological and cellular phenotypes in fugu5-1, ech2- 1, A#3-1sm, fugu5-1 ech2-1, and A#3-1; fugu5-1. Taken together, these results suggest that defects in either HC-PPase or ECH2 compromise cell proliferation due to defects in mobilizing seed storage lipids. In contrast, ECH2 alone likely promotes CCE during the post-mitotic cell expansion stage of cotyledon development, probably by converting indolebutyric acid to indole acetic acid. [ABSTRACT FROM AUTHOR]

DEAD-box RNA helicases belong to an RNA helicase family that plays specific roles in various RNA metabolism processes, including ribosome biogenesis, mRNA splicing, RNA export, mRNA translation and RNA decay. This study investigated a DEAD-box RNA helicase, AtRH7/PRH75, in Arabidopsis. Expression of AtRH7/PRH75 was ubiquitous; however, the levels of mRNA accumulation were increased in cell division regions and were induced by cold stress. The phenotypes of two allelic AtRH7/PRH75-knockout mutants, atrh7-2 and atrh7-3, resembled auxin-related developmental defects that were exhibited in several ribosomal protein mutants, and were more severe under cold stress. Northern blot and circular reverse transcription-PCR (RT-PCR) analyses indicated that unprocessed 18S pre-rRNAs accumulated in the atrh7 mutants. The atrh7 mutants were hyposensitive to the antibiotic streptomycin, which targets ribosomal small subunits, suggesting that AtRH7 was also involved in ribosome assembly. In addition, the atrh7-2 and atrh7-3 mutants displayed cold hypersensitivity and decreased expression of CBF1, CBF2 and CBF3, which might be responsible for the cold intolerance. The present study indicated that AtRH7 participates in rRNA biogenesis and is also involved in plant development and cold tolerance in Arabidopsis. [ABSTRACT FROM AUTHOR]

Epigenetic regulatory states can persist through mitosis and meiosis, but the connection between chromatin structure and DNA replication remains unclear. Arabidopsis INCURVATA2 (ICU2) encodes the catalytic subunit of DNA polymerase α, and null alleles of ICU2 have an embryo-lethal phenotype. Analysis of icu2-1, a hypomorphic allele of ICU2, demonstrated that ICU2 functions in chromatin-mediated cellular memory; icu2-1 strongly impairs ICU2 function in the maintenance of repressive epigenetic marks but does not seem to affect ICU2 polymerase activity. To better understand the global function of ICU2 in epigenetic regulation, here we performed a microarray analysis of icu2-1 mutant plants. We found that the genes up-regulated in the icu2-1 mutant included genes encoding transcription factors and targets of the Polycomb Repressive Complexes. The downregulated genes included many known players in salicylic acid (SA) biosynthesis and accumulation, ABA signaling and ABA-mediated responses. In addition, we found that icu2-1 plants had reduced SA levels in normal conditions; infection by Fusarium oxysporum induced SA accumulation in the En-2 wild type but not in the icu2-1 mutant. The icu2- 1 plants were also hypersensitive to salt stress and exogenous ABA in seedling establishment, post-germination growth and stomatal closure, and accumulated more ABA than the wild type in response to salt stress. The icu2-1 mutant also showed high tolerance to the oxidative stress produced by 3-amino-1,2,4-triazole (3-AT). Our results uncover a role for ICU2 in the regulation of genes involved in ABA signaling as well as in SA biosynthesis and accumulation. [ABSTRACT FROM AUTHOR]

The evolutionarily conserved Elongator complex functions in diverse biological processes including salicylic acid-mediated immune response. However, how Elongator functions in jasmonic acid ( JA)/ethylene ( ET)-mediated defense is unknown. Here, we show that Elongator is required for full induction of the JA/ ET defense pathway marker gene PLANT DEFENSIN1.2 ( PDF1.2) and for resistance to the necrotrophic fungal pathogens Botrytis cinerea and Alternaria brassicicola. A loss-of-function mutation in the Arabidopsis Elongator subunit 2 ( ELP2) alters B. cinerea-induced transcriptome reprogramming. Interestingly, in elp2, expression of WRKY33, OCTADECANOID- RESPONSIVE ARABIDOPSIS AP2/ ERF59 ( ORA59), and PDF1.2 is inhibited, whereas transcription of MYC2 and its target genes is enhanced. However, overexpression of WRKY33 or ORA59 and mutation of MYC2 fail to restore PDF1.2 expression and B. cinerea resistance in elp2, suggesting that ELP2 is required for induction of not only WRKY33 and ORA59 but also PDF1.2. Moreover, elp2 is as susceptible as coronatine-insensitive1 ( coi1) and ethylene-insensitive2 ( ein2) to B. cinerea, indicating that ELP2 is an important player in B. cinerea resistance. Further analysis of the lesion sizes on the double mutants elp2 coi1 and elp2 ein2 and the corresponding single mutants revealed that the function of ELP2 overlaps with COI1 and is additive to EIN2 for B. cinerea resistance. Finally, basal histone acetylation levels in the coding regions of WRKY33, ORA59, and PDF1.2 are reduced in elp2 and a functional ELP2- GFP fusion protein binds to the chromatin of these genes, suggesting that constitutive ELP2-mediated histone acetylation may be required for full activation of the WRKY33/ ORA59/ PDF1.2 transcriptional cascade. [ABSTRACT FROM AUTHOR]

Leaf shape in Arabidopsis is modulated by patterning events in the margin that utilize a PIN-based auxin exporter/CUC2 transcription factor system to define regions of promotion and retardation of growth, leading to morphogenesis. In addition to auxin exporters, leaves also express auxin importers, notably members of the AUX1/ LAX family. In contrast to their established roles in embryogenesis, lateral root and leaf initiation, the function of these transporters in leaf development is poorly understood. We report that three of these genes ( AUX1, LAX1 and LAX2) show specific and dynamic patterns of expression during early leaf development in Arabidopsis, and that loss of expression of all three genes is required for observation of a phenotype in which morphogenesis (serration) is decreased. We used these expression patterns and mutant phenotypes to develop a margin-patterning model that incorporates an AUX1/ LAX1/ LAX2 auxin import module that influences the extent of leaf serration. Testing of this model by margin-localized expression of axr3-1 ( AXR17) provides further insight into the role of auxin in leaf morphogenesis. [ABSTRACT FROM AUTHOR]

Secondary cell-wall thickening takes place in sclerenchyma cells, but not in surrounding parenchyma cells. The molecular mechanism of switching on and off secondary wall synthesis in various cell types is still elusive. Here, we report the identification of a dominant mutant stp-2d showing secondary wall thickening in pith cells ( STP). Immunohistochemistry assays confirmed accumulation of secondary cell walls in the pith cells of the stp-2d mutant. Activation of micro RNA 165b ( miR165b) expression is responsible for the STP phenotype, as demonstrated by transgenic over-expression experiments. The expression of three class III HD- ZIP transcription factor genes, including At HB15, was repressed in the stp-2d mutant. Transgenic over-expression of a mutant form of At HB15 that is resistant to miR165-mediated cleavage reversed the stp-2d mutant phenotype to wild-type, indicating that At HB15 represses secondary wall development in pith. Characterization of two athb15 mutant alleles further confirmed that functional At HB15 is necessary for retaining primary walls in parenchyma pith cells. Expression analyses of cell-wall synthetic genes and wall-related transcription factors indicated that a transcriptional pathway is involved in At HB15 function. These results provide insight into the molecular mechanism of secondary cell-wall development. [ABSTRACT FROM AUTHOR]

Recent advances in the isolation of single cell-types in plants provides an opportunity to conduct detailed analyses of their molecular characteristics at high resolution. This kind of cell-type specific molecular phenotyping is likely to enhance forward genetics studies to dissect the effect of mutations and thereby aid gene function assignment. Recent experimental results support this view, demonstrating that different cell-types exhibit substantial variation in transcript, protein, and metabolite accumulation and these molecular phenotypes are often sensitive to genetic and environmental alterations. The use of single cell-type molecular phenotyping approach to define plant gene function is most amenable to cell-types with well-characterized molecular tools and isolation protocols. [ABSTRACT FROM AUTHOR]

Plant shoot organs such as stems, leaves and flowers are derived from specialized groups of stem cells organized at the shoot apical meristem (SAM). Organogenesis involves two major processes, namely cell proliferation and differentiation, whereby the former contributes to increasing the cell number and the latter involves substantial increases in cell volume through cell expansion. Co-ordination between the above processes in time and space is essential for proper organogenesis. To identify regulatory factors involved in proper organogenesis, heavy-ion beam-irradiated de-etiolated (det) 3-1 seeds have been used to identify striking phenotypes in the A#26-2; det3-1 mutant. In addition to the stunted plant stature mimicking det3-1, the A#26-2; det3-1 mutant exhibited stem thickening, increased floral organ number and a fruit shape reminiscent of clavata (clv) mutants. DNA sequencing analysis demonstrated that A#26-2; det3-1 harbors a mutation in the CLV3 gene. Importantly, A#26-2; det3-1 displayed cracks that randomly occurred on the main stem with a frequency of approximately 50%. Furthermore, the double mutants clv3-8 det3-1, clv1-4 det3-1 and clv2-1 det3-1 consistently showed stem cracks with frequencies of approximately 97, 38 and 35%, respectively. Cross-sections of stems further revealed an increase in vascular bundle number, cell number and size in the pith of clv3-8 det3-1 compared with det3-1. These findings suggest that the stem inner volume increase due to clv mutations exerts an outward mechanical stress; that in a det3-1 background (defective in cell expansion) resulted in cracking of the outermost layer of epidermal cells. [ABSTRACT FROM AUTHOR]

Plant responses to insects and wounding involve substantial transcriptional reprogramming that integrates hormonal, metabolic, and physiological events. The ability to respond differentially to various stresses, including wounding, generally involves hormone signaling and trans-acting regulatory factors. Evidence of the importance of transcription factors (TFs) in responses to insects is also accumulating. However, the relationships among hormone signaling, TF activity, and ability to respond specifically to different insects are uncertain. We examined transcriptional and hormonal changes in Arabidopsis thaliana after herbivory by larvae of two lepidopteran species, Spodoptera exigua (Hübner) and Pieris rapae L. over a 24-hour time course. Transcriptional responses to the two insects differed and were frequently weaker or absent in response to the specialist P. rapae. Using microarray analysis and qRT-PCR, we found 141 transcription factors, including many AP2/ERFs (Ethylene Response Factors) and selected defense-related genes, to be differentially regulated in response to the two insect species or wounding. Jasmonic Acid (JA), JA-isoleucine, and ethylene production by Arabidopsis plants increased after attack by both insect species. However, the amounts and timing of ethylene production differed between the two herbivory treatments. Our results support the hypothesis that the different responses to these two insects involve modifications of JA-signaling events and activation of different subsets of ERF transcription factors, resulting in different degrees of divergence from responses to wounding alone. [ABSTRACT FROM AUTHOR]

Stomata are two-celled valves that control epidermal pores whose opening and spacing optimizes shoot-atmosphere gas exchange. Arabidopsis stomatal formation involves at least one asymmetric division and one symmetric division. Stomatal formation and patterning are regulated by the frequency and placement of asymmetric divisions. This model system has already led to significant advances in developmental biology, such as the regulation of cell fate, division, differentiation, and patterning. Over the last 30 years, stomatal development has been found to be controlled by numerous intrinsic genetic and environmental factors. This mini review focuses on the signaling involved in stomatal initiation and in divisions in the cell lineage. [ABSTRACT FROM AUTHOR]

Seed size in higher plants is an important agronomic trait, and is also crucial for evolutionary fitness. In flowering plants, the seed comprises three major anatomical components, the embryo, the en dosperm and the seed coat, each with different genetic compositions. Therefore, seed size is coordinately determined by the growth of the embryo, endosperm and maternal tissue. Recent studies have revealed multiple pathways that influence seed size in plants. Several factors involved in ubiquitin-related activities have been recently known to determine seed size in Arabidopsis and rice. In this review, we summarize current knowledge of ubiquitin-mediated control of seed size and discuss the role of the ubiquitin pathway in seed size control. [ABSTRACT FROM AUTHOR]

Functional redundancy arises between gene paralogs as well as non-homologous genes that play a common role at a shared node. The bHLH transcription factor FAMA, along with the paralogous MYB genes, FOUR LIPS ( FLP) and MYB88 all ensure that Arabidopsis stomata contain just two guard cells (GCs) by enforcing a single symmetric precursor cell division before stomatal maturity. Consistent with this function, FLP and FAMA exhibit the same expression pattern in which both translational GFP fusions emit fluorescence just before and after symmetric division; however, FAMA but not FLP is required to confer GC fate. Strikingly, swapping the genes and promoters of the FLP and FAMA genes results in the reciprocal complementation of respective loss-of-function mutants. Thus, an FLP transgene can restore GC fate to a fama mutant background. FAMA, FLP and the FLP paralog MYB88 were previously shown to influence higher order functions in stomatal development, including maintaining and stabilizing stomatal fate. Here we show that these overlapping functions are likely to also involve interactions between FLP and FAMA with the RETINOBLASTOMA-RELATED (RBR) protein. [ABSTRACT FROM AUTHOR]

Trichomes are specialized epidermal cells located on aerial parts of plants and are associated with a wide array of biological processes. Trichomes protect plants from adverse conditions including UV light and herbivore attack and are also an important source of a number of phytochemicals. The simple unicellular trichomes of Arabidopsis serve as an excellent model to study molecular mechanism of cell differentiation and pattern formation in plants. The emerging picture suggests that the developmental process is controlled by a transcriptional network involving three major groups of transcription factors: the R2R3 MYB, basic helix-loop-helix (bHLH) and WD40 repeat (WDR) protein. These regulatory proteins form a trimeric activator complex that positively regulates trichome development. The single repeat R3 MYBs act as negative regulators of trichome development. They compete with the R2R3 MYBs to bind the bHLH factor and form a repressor complex. In addition to activator-repressor mechanism, a depletion mechanism may operate in parallel during trichome development. In this mechanism, the bHLH factor traps the WDR protein which results in depletion of WDR protein in neighboring cells. Consequently, the cells with high levels of bHLH and WDR proteins are developed into trichomes. A group of C2H2 zinc finger TFs has also been implicated in trichome development. Phytohormones, including gibberellins and jasmonic acid, play significant roles in this developmental process. Recently, microRNAs have been shown to be involved in trichome development. Furthermore, it has been demonstrated that the activities of the key regulatory proteins involved in trichome development are controlled by the 26S/ubiquitin proteasome system (UPS), highlighting the complexity of the regulatory network controlling this developmental process. To complement several excellent recent relevant reviews, this review focuses on the transcriptional network and hormonal interplay controlling trichome development in Arabidopsis. [ABSTRACT FROM AUTHOR]

Maintenance of organ separation is one of the essential phenomena for normal plant development. We have identified and analyzed ONION3 (ONI3), which is required for avoiding organ fusions in rice. Loss-of-function mutations of ONI3, which were identified as mutants with ectopic expression of KNOX genes in leaves and morphologically resembling KNOX overexpressors, showed abnormal organ fusions in developing shoots. The mutant seedlings showed fusions between neighboring organs and also within an organ; they stopped growing soon after germination and subsequently died. ONI3 was shown to encode an enzyme that is most similar to Arabidopsis HOTHEAD and is involved in biosynthesis of long-chain fatty acids. Expression analyses showed that ONI3 was specifically expressed in the outermost cell layer in the shoot apex throughout life cycle, and the oni3 mutants had an aberrant outermost cell layer. Our results together with previous studies suggest that long-chain fatty acids are required for avoiding organ fusions and promoting normal shoot development in rice. [ABSTRACT FROM PUBLISHER]