MERISTEM-DEFECTIVE regulates the balance between stemness and differentiation in the root meristem through RNA splicing control.

Plants respond to environmental stresses through controlled stem cell maintenance and meristem activity. One level of gene regulation is RNA alternative splicing. However, the mechanistic link between stress, meristem function and RNA splicing is poorly understood. The MERISTEM-DEFECTIVE (MDF) Arabidopsis gene encodes an SR-related family protein, required for meristem function and leaf vascularization, and is the likely orthologue of the human SART1 and yeast Snu66 splicing factors. MDF is required for the correct splicing and expression of key transcripts associated with root meristem function. We identified RSZ33 and ACC1, both known to regulate cell patterning, as splicing targets required for MDF function in the meristem. MDF expression is modulated by osmotic and cold stress, associated with differential splicing and specific isoform accumulation and shuttling between nucleus and cytosol, and acts in part via a splicing target SR34. We propose a model in which MDF controls splicing in the root meristem to promote stemness and to repress stress response, cell differentiation and cell death pathways.

GhTCE1-GhTCEE1 dimers regulate transcriptional reprogramming during wound-induced callus formation in cotton.

Wounded plant cells can form callus to seal the wound site. Alternatively, wounding can cause adventitious organogenesis or somatic embryogenesis. These distinct developmental pathways require specific cell fate decisions. Here, we identify GhTCE1, a basic helix-loop-helix family transcription factor, and its interacting partners as a central regulatory module of early cell fate transition during in vitro dedifferentiation of cotton (Gossypium hirsutum). RNAi- or CRISPR/Cas9-mediated loss of GhTCE1 function resulted in excessive accumulation of reactive oxygen species (ROS), arrested callus cell elongation, and increased adventitious organogenesis. In contrast, GhTCE1-overexpressing tissues underwent callus cell growth, but organogenesis was repressed. Transcriptome analysis revealed that several pathways depend on proper regulation of GhTCE1 expression, including lipid transfer pathway components, ROS homeostasis, and cell expansion. GhTCE1 bound to the promoters of the target genes GhLTP2 and GhLTP3, activating their expression synergistically, and the heterodimer TCE1-TCEE1 enhances this activity. GhLTP2- and GhLTP3-deficient tissues accumulated ROS and had arrested callus cell elongation, which was restored by ROS scavengers. These results reveal a unique regulatory network involving ROS and lipid transfer proteins, which act as potential ROS scavengers. This network acts as a switch between unorganized callus growth and organized development during in vitro dedifferentiation of cotton cells.

Drought response revealed by chromatin organization variation and transcriptional regulation in cotton.

BACKGROUND: Cotton is a major world cash crop and an important source of natural fiber, oil, and protein. Drought stress is becoming a restrictive factor affecting cotton production. To facilitate the development of drought-tolerant cotton varieties, it is necessary to study the molecular mechanism of drought stress response by exploring key drought-resistant genes and related regulatory factors. RESULTS: In this study, two cotton varieties, ZY007 (drought-sensitive) and ZY168 (drought-tolerant), showing obvious phenotypic differences under drought stress, were selected. A total of 25,898 drought-induced genes were identified, exhibiting significant enrichment in pathways related to plant stress responses. Under drought induction, At subgenome expression bias was observed at the whole-genome level, which may be due to stronger inhibition of Dt subgenome expression. A gene co-expression module that was significantly associated with drought resistance was identified. About 90% of topologically associating domain (TAD) boundaries were stable, and 6613 TAD variation events were identified between the two varieties under drought. We identified 92 genes in ZY007 and 98 in ZY168 related to chromatin 3D structural variation and induced by drought stress. These genes are closely linked to the cotton response to drought stress through canonical hormone-responsive pathways, modulation of kinase and phosphatase activities, facilitation of calcium ion transport, and other related molecular mechanisms. CONCLUSIONS: These results lay a foundation for elucidating the molecular mechanism of the cotton drought response and provide important regulatory locus and gene resources for the future molecular breeding of drought-resistant cotton varieties.

Stochastic modeling of the mRNA life process: A generalized master equation.

The mRNA life cycle is a complex biochemical process, involving transcription initiation, elongation, termination, splicing, and degradation. Each of these molecular events is multistep and can create a memory. The effect of this molecular memory on gene expression is not clear, although there are many related yet scattered experimental reports. To address this important issue, we develop a general theoretical framework formulated as a master equation in the sense of queue theory, which can reduce to multiple previously studied gene models in limiting cases. This framework allows us to interpret experimental observations, extract kinetic parameters from experimental data, and identify how the mRNA kinetics vary under regulatory influences. Notably, it allows us to evaluate the influences of elongation processes on mature RNA distribution; e.g., we find that the non-exponential elongation time can induce the bimodal mRNA expression and there is an optimal elongation noise intensity such that the mature RNA noise achieves the lowest level. In a word, our framework can not only provide insight into complex mRNA life processes but also bridge a dialogue between theoretical studies and experimental data.

N6-methyladenosine RNA modification regulates cotton drought response in a Ca2+ and ABA-dependent manner.

N6 -methyladenosine (m6 A) is the most prevalent internal modification present in mRNAs, and is considered to participate in a range of developmental and biological processes. Drought response is highly regulated at the genomic, transcriptional and post-transcriptional levels. However, the biological function and regulatory mechanism of m6 A modification in the drought stress response is still poorly understood. We generated a transcriptome-wide m6 A map using drought-resistant and drought-sensitive varieties of cotton under different water deficient conditions to uncover patterns of m6 A methylation in cotton response to drought stress. The results reveal that m6 A represents a common modification and exhibit dramatic changes in distribution during drought stress. More 5'UTR m6 A was deposited in the drought-resistant variety and was associated with a positive effect on drought resistance by regulating mRNA abundance. Interestingly, we observed that increased m6 A abundance was associated with increased mRNA abundance under drought, contributing to drought resistance, and vice versa. The demethylase GhALKBH10B was found to decrease m6 A levels, facilitating the mRNA decay of ABA signal-related genes (GhZEP, GhNCED4 and GhPP2CA) and Ca2+ signal-related genes (GhECA1, GhCNGC4, GhANN1 and GhCML13), and mutation of GhALKBH10B enhanced drought resistance at seedling stage in cotton. Virus-induced gene silencing (VIGS) of two Ca2+ -related genes, GhECA1 and GhCNGC4, reduced drought resistance with the decreased m6 A enrichment on silenced genes in cotton. Collectively, we reveal a novel mechanism of post-transcriptional modification involved in affecting drought response in cotton, by mediating m6 A methylation on targeted transcripts in the ABA and Ca2+ signalling transduction pathways.

A PXY-Mediated Transcriptional Network Integrates Signaling Mechanisms to Control Vascular Development in Arabidopsis.

The cambium and procambium generate the majority of biomass in vascular plants. These meristems constitute a bifacial stem cell population from which xylem and phloem are specified on opposing sides by positional signals. The PHLOEM INTERCALATED WITH XYLEM (PXY) receptor kinase promotes vascular cell division and organization. However, how these functions are specified and integrated is unknown. Here, we mapped a putative PXY-mediated transcriptional regulatory network comprising 690 transcription factor-promoter interactions in Arabidopsis (Arabidopsis thaliana). Among these interactions was a feedforward loop containing transcription factors WUSCHEL HOMEOBOX RELATED14 (WOX14) and TARGET OF MONOPTEROS6 (TMO6), each of which regulates the expression of the gene encoding a third transcription factor, LATERAL ORGAN BOUNDARIES DOMAIN4 (LBD4). PXY signaling in turn regulates the WOX14, TMO6, and LBD4 feedforward loop to control vascular proliferation. Genetic interaction between LBD4 and PXY suggests that LBD4 marks the phloem-procambium boundary, thus defining the shape of the vascular bundle. These data collectively support a mechanism that influences the recruitment of cells into the phloem lineage, and they define the role of PXY signaling in this context in determining the arrangement of vascular tissue.

Single-cell RNA-seq reveals fate determination control of an individual fibre cell initiation in cotton (Gossypium hirsutum).

Cotton fibre is a unicellular seed trichome, and lint fibre initials per seed as a factor determines fibre yield. However, the mechanisms controlling fibre initiation from ovule epidermis are not understood well enough. Here, with single-cell RNA sequencing (scRNA-seq), a total of 14 535 cells were identified from cotton ovule outer integument of Xu142_LF line at four developmental stages (1.5, 1, 0.5 days before anthesis and the day of anthesis). Three major cell types, fibre, non-fibre epidermis and outer pigment layer were identified and then verified by RNA in situ hybridization. A comparative analysis on scRNA-seq data between Xu142 and its fibreless mutant Xu142 fl further confirmed fibre cluster definition. The developmental trajectory of fibre cell was reconstructed, and fibre cell was identified differentiated at 1 day before anthesis. Gene regulatory networks at four stages revealed the spatiotemporal pattern of core transcription factors, and MYB25-like and HOX3 were demonstrated played key roles as commanders in fibre differentiation and tip-biased diffuse growth respectively. A model for early development of a single fibre cell was proposed here, which sheds light on further deciphering mechanism of plant trichome and the improvement of cotton fibre yield.

Transcriptome and metabolome profiling of interspecific CSSLs reveals general and specific mechanisms of drought resistance in cotton.

In order to understand the molecular mechanism of cotton's response to drought during the flowering and boll stage, transcriptomics and metabolomics were carried out for two introgression lines (drought-tolerant line: T307; drought-sensitive line: S48) which were screened from Gossypium hirsutum cv. 'Emian22' with some gene fragments imported from Gossypium barbadense acc. 3-79, under drought stress by withdrawing water at flowering and boll stage. Results showed that the basic drought response in cotton included a series of broad-spectrum responses, such as amino acid synthesis, hormone (abscisic acid, ABA) signal transduction, and mitogen-activated protein kinases signal transduction pathway, which activated in both drought-tolerant and drought-sensitive lines. However, the difference of their imported fragments and diminished sequences triggers endoplasmic reticulum (ER) protein processing, photosynthetic-related pathways (in leaves), and membrane solute transport (in roots) in drought-tolerant line T307, while these are missed or not activated in drought-sensitive line S48, reflecting the different drought tolerance of the two genotypes. Virus-induced gene silencing assay of drought-tolerant differentially expressed heat shock protein (HSP) genes (mainly in leaf) and ATP-binding cassette (ABC) transporter genes (mainly in roots) indicated that those genes play important role in cotton drought tolerant. Combined analysis of transcriptomics and metabolomics highlighted the important roles of ER-stress-related HSP genes and root-specific ABC transporter genes in plants drought tolerance. These results provide new insights into the molecular mechanisms underlying the drought stress adaptation in cotton.

Silent transcription intervals and translational bursting lead to diverse phenotypic switching.

Gene-expression bimodality, as a potential mechanism generating phenotypic cell diversity, can enhance the survival of cells in a fluctuating environment. Previous studies have shown that intrinsic or extrinsic regulations could induce bimodal gene expressions, but it is unclear whether this bimodality can occur without regulation. Here we develop an interpretable and tractable model, namely a generalized telegraph model (GTM), which considers silent transcription intervals and translational bursting, each being characterized by a general distribution. Using the queuing theory, we derive the analytical expressions of protein distributions, and show that non-exponential inactive times and translational bursting can lead to two peaks of the protein distribution away from the origin, which are different from those occurring in classical telegraph models. We also find that both silent-interval noise and translational burst-size noise can amplify gene-expression noise and induce diverse dynamic expression patterns. Our results not only provide an alternative mechanism of phenotypic switching but also could be used in explaining the bimodal phenomenon in experimental observations.

Identification and Functional Analysis of bZIP Genes in Cotton Response to Drought Stress.

The basic leucine zipper (bZIP) transcription factors, which harbor a conserved bZIP domain composed of two regions, a DNA-binding basic region and a Leu Zipper region, operate as important switches of transcription networks in eukaryotes. However, this gene family has not been systematically characterized in cotton (Gossypium hirsutum). Here, we identified 197 bZIP family members in cotton. The chromosome distribution pattern indicates that the GhbZIP genes have undergone 53 genome-wide segmental and 7 tandem duplication events which contribute to the expansion of the cotton bZIP family. Phylogenetic analysis showed that cotton GhbZIP proteins cluster into 13 subfamilies, and homologous protein pairs showed similar characteristics. Inspection of the DNA-binding basic region and leucine repeat heptads within the bZIP domains indicated different DNA-binding site specificities as well as dimerization properties among different groups. Comprehensive expression analysis indicated the most highly and differentially expressed genes in root and leaf that might play significant roles in cotton response to drought stress. GhABF3D was identified as a highly and differentially expressed bZIP family gene in cotton leaf and root under drought stress treatments that likely controls drought stress responses in cotton. These data provide useful information for further functional analysis of the GhbZIP gene family and its potential application in crop improvement.

The Calcium Sensor CBL2 and Its Interacting Kinase CIPK6 Are Involved in Plant Sugar Homeostasis via Interacting with Tonoplast Sugar Transporter TST2.

Calcineurin B-like protein (CBL) and CBL-interacting protein kinase (CIPK)-mediated calcium signaling has been widely reported to function in plant development and various stress responses, particularly in ion homeostasis. Sugars are the most important primary metabolites, and thus sugar homeostasis requires precise regulation. Here, we describe a CBL2-CIPK6-Tonoplast-Localized Sugar Transporter2 (TST2) molecular module in cotton (Gossypium hirsutum) that regulates plant sugar homeostasis, in particular Glc homeostasis. GhCIPK6 is recruited to the tonoplast by GhCBL2 and interacts with the tonoplast-localized sugar transporter GhTST2. Overexpression of either GhCBL2, GhCIPK6, or GhTST2 was sufficient to promote sugar accumulation in transgenic cotton, whereas RNAi-mediated knockdown of GhCIPK6 expression or CRISPR-Cas9-mediated knockout of GhTST2 resulted in significantly decreased Glc content. Moreover, mutation of GhCBL2 or GhTST2 in GhCIPK6-overexpressing cotton reinstated sugar contents comparable to wild-type plants. Heterologous expression of GhCIPK6 in Arabidopsis (Arabidopsis thaliana) also promoted Glc accumulation, whereas mutation of AtTST1/2 in GhCIPK6-overexpressing Arabidopsis similarly reinstated wild-type sugar contents, thus indicating conservation of CBL2-CIPK6-TST2-mediated sugar homeostasis among different plant species. Our characterization of the molecular players behind plant sugar homeostasis may be exploited to improve sugar contents and abiotic stress resistance in plants.

Genome-wide analysis of CBL and CIPK family genes in cotton: conserved structures with divergent interactions and expression.

Calcineurin B-like proteins (CBLs) interact with CBL-interacting protein kinases (CIPKs) to form complex molecular modules in response to diverse abiotic stresses. Although previous studies demonstrated that the CBL-CIPK networks play a crucial role in plants response to abiotic stresses, however, little is known about their functions in cotton. In the present study, a total of 22 GhCBL and 79 GhCIPK gene family members were identified in upland cotton (Gossypium hirsutum Linn). Synteny analysis revealed that most genes of GhCBL and GhCIPK exist in pairs between At sub-genome and Dt sub-genome. Interaction analysis between GhCBL and GhCIPK proteins by yeast two-hybrid (Y2H) suggested that the GhCBL-GhCIPK networks were complex, and exhibited functional redundancy in cotton. Quantitative expression analysis by public transcriptome datasets revealed that some GhCBL and GhCIPK genes are differentially expressed under abiotic stress treatments, and especially under drought stress. Our results not only contribute to understanding the structural features of GhCBL and GhCIPK genes but also provide the basis for in-depth functional studies of GhCBL-GhCIPK networks in stress response for plants. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at (doi:10.1007/s12298-021-00943-1).

Genome-wide identification of MAPK cascade genes reveals the GhMAP3K14-GhMKK11-GhMPK31 pathway is involved in the drought response in cotton.

The mitogen-activated protein kinase (MAPK) cascade pathway, which has three components, MAP3Ks, MKKs and MPKs, is involved in diverse biological processes in plants. In the current study, MAPK cascade genes were identified in three cotton species, based on gene homology with Arabidopsis. Selection pressure analysis of MAPK cascade genes revealed that purifying selection occurred among the cotton species. Expression pattern analysis showed that some MAPK cascade genes differentially expressed under abiotic stresses and phytohormones treatments, and especially under drought stress. Yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) experiments showed extensive interactions between different MAPK cascade proteins. Virus-induced gene silencing (VIGS) assays showed that some MAPK cascade modules play important roles in the drought stress response, and the GhMAP3K14-GhMKK11-GhMPK31 signal pathway was demonstrated to regulate drought stress tolerance in cotton. This study provides new information on the function of MAPK cascade genes in the drought response, and will help direct molecular breeding for improved drought stress tolerance in cotton.

Phenomics-based GWAS analysis reveals the genetic architecture for drought resistance in cotton.

Drought resistance (DR) is a complex trait that is regulated by a variety of genes. Without comprehensive profiling of DR-related traits, the knowledge of the genetic architecture for DR in cotton remains limited. Thus, there is a need to bridge the gap between genomics and phenomics. In this study, an automatic phenotyping platform (APP) was systematically applied to examine 119 image-based digital traits (i-traits) during drought stress at the seedling stage, across a natural population of 200 representative upland cotton accessions. Some novel i-traits, as well as some traditional i-traits, were used to evaluate the DR in cotton. The phenomics data allowed us to identify 390 genetic loci by genome-wide association study (GWAS) using 56 morphological and 63 texture i-traits. DR-related genes, including GhRD2, GhNAC4, GhHAT22 and GhDREB2, were identified as candidate genes by some digital traits. Further analysis of candidate genes showed that Gh_A04G0377 and Gh_A04G0378 functioned as negative regulators for cotton drought response. Based on the combined digital phenotyping, GWAS analysis and transcriptome data, we conclude that the phenomics dataset provides an excellent resource to characterize key genetic loci with an unprecedented resolution which can inform future genome-based breeding for improved DR in cotton.

The ß-ketoacyl-CoA synthase KCS13 regulates the cold response in cotton by modulating lipid and oxylipin biosynthesis.

Cold stress is a key environmental factor that affects plant development and productivity. In this study, RNA-seq in cotton following cold-stress treatment resulted in the identification of 5239 differentially expressed genes (DEGs) between two cultivars with differing sensitivity to low temperatures, among which GhKCS13 was found to be involved in the response. Transgenic plants overexpressing GhKCS13 showed increased sensitivity to cold stress. KEGG analysis of 418 DEGs in both GhKCS13-overexpressing and RNAi lines after treatment at 4 °C indicated that lipid biosynthesis and linoleic acid metabolism were related to cold stress. ESI-MS/MS analysis showed that overexpression of GhKCS13 led to modifications in the composition of sphingolipids and glycerolipids in the leaves, which might alter the fluidity of the cell membrane under cold conditions. In particular, differences in levels of jasmonic acid (JA) in GhKCS13 transgenic lines suggested that, together with lysophospholipids, it might mediate the cold-stress response. Our results suggest that overexpression of GhKCS13 probably causes remodeling of lipids in the endoplasmic reticulum and biosynthesis of lipid-derived JA in chloroplasts, which might account for the increased sensitivity to cold stress in the transgenic plants. Complex interactions between lipid components, lipid signaling molecules, and JA appear to determine the response to cold stress in cotton.

Myofibroblast-Derived Exosomes Contribute to Development of a Susceptible Substrate for Atrial Fibrillation.

OBJECTIVE: Atrial fibrosis plays a critical role in atrial fibrillation (AF). A key event in the pathogenesis of fibrosis is the activation of fibroblasts (FBs) into myofibroblasts (MFBs). Paracrine factors released from MFBs lead to ion channel expression changes in cardiomyocytes (CMs). Downregulation of L-type calcium channel Cav1.2 expression is a hallmark of AF-associated ionic remodeling. However, whether exosome (Exo)-mediated crosstalk between MFBs and CMs regulates Cav1.2 expression remains unknown. METHODS: Atrial FBs and CMs were isolated and cultured from neonatal rats by enzymatic digestion. The activation of FBs into MFBs was induced by angiotensin II. Co-culture assay and in vitro Exo treatment were used to determine the effect of MFB-derived Exos on Cav1.2 expression. Confocal Ca2+ imaging was performed to examine the adrenergic stimulation-elicited Ca2+ influx signals. The levels of potential Cav1.2-inhibitory microRNAs (miRNAs) were measured by qRT-PCR. RESULTS: Untreated FBs expressed limited amounts of alpha smooth muscle actin (α-SMA), while angiotensin II induced a significant upregulation of α-SMA-expressing MFBs. Co-cultures of MFBs and CMs resulted in downregulation of Cav1.2 expression in CMs, which was largely abolished by pretreatment of MFBs with exosomal inhibitor GW4869. More importantly, treatment with MFB-derived Exos caused repression of Cav1.2 expression in CMs. Additionally, the adrenergic receptor agonist-elicited Ca2+ influx signals in CMs were remarkably attenuated by pretreatment with MFB-derived Exos, corresponding to the paralleled change in Cav1.2 expression. Finally, miR-21-3p, a potential Cav1.2-inhibitory miRNA, was enriched in MFB-derived Exos and upregulated in CMs in response to MFB-derived Exos. CONCLUSION: We uncover an Exo-mediated crosstalk between MFBs and CMs, contributing to increased vulnerability to AF by reducing the expression of Cav1.2 in CMs.

Characterization of a novel cotton MYB gene, GhMYB108-like responsive to abiotic stresses.

Transcriptional factors are the major regulators of plant signaling pathways in response to environmental stresses i.e., drought, salinity and cold. Hereby, the GhMYB108-like was characterized to determine whether it regulate these stresses. The GhMYB108-like cDNA consisted of 1107 base pairs (bp) with 807 open reading frame encoded a protein of 268 amino acids. Its isoelectric point and molecular weight are 5.51 and 30.3 kDa respectively. Phylogenetic analysis and online databases revealed that GhMYB108-like proteins are closely related with the Arabidopsis thaliana MYB2. Important cis-elements were detected in the promotor region of GhMYB108-like responding to stresses and phytohormones. The 3D structure of GhMYB108-like protein has been predicted. In addition, various physico-chemical properties of GhMYB108-like have been determined. Subcellular localization confirmed that GhMYB108-like are nuclear localized protein. Quantitative expression analysis showed that polyethylene glycol and salt treatments significantly induced the expression of GhMYB108-like. Overall, our findings suggest that GhMYB108-like is an important gene that would plays important regulatory role in response to drought and salt stresses.

GhL1L1 affects cell fate specification by regulating GhPIN1-mediated auxin distribution.

Auxin is as an efficient initiator and regulator of cell fate during somatic embryogenesis (SE), but the molecular mechanisms and regulating networks of this process are not well understood. In this report, we analysed SE process induced by Leafy cotyledon1-like 1 (GhL1L1), a NF-YB subfamily gene specifically expressed in embryonic tissues in cotton. We also identified the target gene of GhL1L1, and its role in auxin distribution and cell fate specification during embryonic development was analysed. Overexpression of GhL1L1 accelerated embryonic cell formation, associated with an increased concentration of IAA in embryogenic calluses (ECs) and in the shoot apical meristem, corresponding to altered expression of the auxin transport gene GhPIN1. By contrast, GhL1L1-deficient explants showed retarded embryonic cell formation, and the concentration of IAA was decreased in GhL1L1-deficient ECs. Disruption of auxin distribution accelerated the specification of embryonic cell fate together with regulation of GhPIN1. Furthermore, we showed that PHOSPHATASE 2AA2 (GhPP2AA2) was activated by GhL1L1 through targeting the G-box of its promoter, hence regulating the activity of GhPIN1 protein. Our results indicate that GhL1L1 functions as a key regulator in auxin distribution to regulate cell fate specification in cotton and contribute to the understanding of the complex process of SE in plant species.

Laccase GhLac1 Modulates Broad-Spectrum Biotic Stress Tolerance via Manipulating Phenylpropanoid Pathway and Jasmonic Acid Synthesis.

Plants are constantly challenged by a multitude of pathogens and pests, which causes massive yield and quality losses annually. A promising approach to reduce such losses is to enhance the immune system of plants through genetic engineering. Previous work has shown that laccases (p-diphenol:dioxygen oxidoreductase, EC 1.10.3.2) function as lignin polymerization enzymes. Here we demonstrate that transgenic manipulation of the expression of the laccase gene GhLac1 in cotton (Gossypium hirsutum) can confer an enhanced defense response to both pathogens and pests. Overexpression of GhLac1 leads to increased lignification, associated with increased tolerance to the fungal pathogen Verticillium dahliae and to the insect pests cotton bollworm (Helicoverpa armigera) and cotton aphid (Aphis gosypii). Suppression of GhLac1 expression leads to a redirection of metabolic flux in the phenylpropanoid pathway, causing the accumulation of JA and secondary metabolites that confer resistance to V. dahliae and cotton bollworm; it also leads to increased susceptibility to cotton aphid. Plant laccases therefore provide a new molecular tool to engineer pest and pathogen resistance in crops.

Golgi-localized cation/proton exchangers regulate ionic homeostasis and skotomorphogenesis in Arabidopsis.

Multiple transporters and channels mediate cation transport across the plasma membrane and tonoplast to regulate ionic homeostasis in plant cells. However, much less is known about the molecular function of transporters that facilitate cation transport in other organelles such as Golgi. We report here that Arabidopsis KEA4, KEA5, and KEA6, members of cation/proton antiporters-2 (CPA2) superfamily were colocalized with the known Golgi marker, SYP32-mCherry. Although single kea4,5,6 mutants showed similar phenotype as the wild type under various conditions, kea4/5/6 triple mutants showed hypersensitivity to low pH, high K+ , and high Na+ and displayed growth defects in darkness, suggesting that these three KEA-type transporters function redundantly in controlling etiolated seedling growth and ion homeostasis. Detailed analysis indicated that the kea4/5/6 triple mutant exhibited cell wall biosynthesis defect during the rapid etiolated seedling growth and under high K+ /Na+ condition. The cell wall-derived pectin homogalacturonan (GalA)3 partially suppressed the growth defects and ionic toxicity in the kea4/5/6 triple mutants when grown in the dark but not in the light conditions. Together, these data support the hypothesis that the Golgi-localized KEAs play key roles in the maintenance of ionic and pH homeostasis, thereby facilitating Golgi function in cell wall biosynthesis during rapid etiolated seedling growth and in coping with high K+ /Na+ stress.