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Stomata in leaves regulate gas (carbon dioxide and water vapor) exchange and water transpiration between plants and the atmosphere. SLow Anion Channel 1 (SLAC1) mediates anion efflux from guard cells and plays a crucial role in controlling stomatal aperture. It serves as a central hub for multiple signaling pathways in response to environmental stimuli, with its activity regulated through phosphorylation via various plant protein kinases. However, the molecular mechanism underlying SLAC1 phosphoactivation has remained elusive. Through a combination of protein sequence analyses, AlphaFold-based modeling and electrophysiological studies, we unveiled that the highly conserved motifs on the N- and C-terminal segments of SLAC1 form a cytosolic regulatory domain (CRD) that interacts with the transmembrane domain(TMD), thereby maintaining the channel in an autoinhibited state. Mutations in these conserved motifs destabilize the CRD, releasing autoinhibition in SLAC1 and enabling its transition into an activated state. Our further studies demonstrated that SLAC1 activation undergoes an autoinhibition-release process and subsequent structural changes in the pore helices. These findings provide mechanistic insights into the activation mechanism of SLAC1 and shed light on understanding how SLAC1 controls stomatal closure in response to environmental stimuli., Competing Interests: Competing interests statement:The authors declare no competing interest.

Bacterial pathogens deliver effectors into host cells to suppress immunity. How host cells target these effectors is critical in pathogen-host interactions. SUMOylation, an important type of posttranslational modification in eukaryotic cells, plays a critical role in immunity, but its effect on bacterial effectors remains unclear in plant cells. In this study, using bioinformatic and biochemical approaches, we found that at least 16 effectors from the bacterial pathogen Pseudomonas syringae pv. tomato DC3000 are SUMOylated by the enzyme cascade from Arabidopsis thaliana. Mutation of SUMOylation sites on the effector HopB1 enhances its function in the induction of plant cell death via stability attenuation of a plant receptor kinase BRASSINOSTEROID INSENSITIVE 1 (BRI1)-ASSOCIATED RECEPTOR KINASE 1. By contrast, SUMOylation is essential for the function of another effector, HopG1, in the inhibition of mitochondria activity and jasmonic acid signaling. SUMOylation of both HopB1 and HopG1 is increased by heat treatment, and this modification modulates the functions of these 2 effectors in different ways in the regulation of plant survival rates, gene expression, and bacterial infection under high temperatures. Therefore, the current work on the SUMOylation of effectors in plant cells improves our understanding of the function of dynamic protein modifications in plant-pathogen interactions in response to environmental conditions., Competing Interests: Conflict of interest statement. None declared., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society of Plant Biologists. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

For plant growth under salt stress, sensing and transducing salt signals are central to cellular Na + homoeostasis. The calcineurin B-like protein (CBL)-CBL-interacting protein kinase (CIPK) complexes play critical roles in transducing salt signals in plants. Here, we show that CBL5, an ortholog of CBL4 and CBL10 in Arabidopsis, interacts with and recruits CIPK8/CIPK24 to the plasma membrane. Yeast cells coexpressing CBL5, CIPK8/CIPK24 and SOS1 demonstrated lesser Na + accumulation and a better growth phenotype than the untransformed or SOS1 transgenic yeast cells under salinity. Overexpression of CBL5 improved the growth of the cipk8 or cipk24 single mutant but not the cipk8 cipk24 double mutant under salt stress, suggesting that CIPK8 and CIPK24 were the downstream targets of CBL5. Interestingly, seed germination in cbl5 was severely inhibited by NaCl, which was recovered by the overexpression of CBL5. Furthermore, CBL5 was mainly expressed in the cotyledons and hypocotyls, which are essential to seed germination. Na + efflux activity in the hypocotyls of cbl5 was reduced relative to the wild-type under salt stress, enhancing Na + accumulation. These findings indicate that CBL5 functions in seed germination and protects seeds and germinating seedlings from salt stress through the CBL5-CIPK8/CIPK24-SOS1 pathways., (© 2024 John Wiley & Sons Ltd.)

Circadian clocks are endogenous timekeeping mechanisms that coordinate internal physiological responses with the external environment. EARLY FLOWERING3 (ELF3), PSEUDO RESPONSE REGULATOR (PRR9), and PRR7 are essential components of the plant circadian clock and facilitate entrainment of the clock to internal and external stimuli. Previous studies have highlighted a critical role for ELF3 in repressing the expression of PRR9 and PRR7. However, the functional significance of activity in regulating circadian clock dynamics and plant development is unknown. To explore this regulatory dynamic further, we first employed mathematical modeling to simulate the effect of the prr9/prr7 mutation on the elf3 circadian phenotype. These simulations suggested that simultaneous mutations in prr9/prr7 could rescue the elf3 circadian arrhythmia. Following these simulations, we generated all Arabidopsis elf3/prr9/prr7 mutant combinations and investigated their circadian and developmental phenotypes. Although these assays could not replicate the results from the mathematical modeling, our results have revealed a complex epistatic relationship between ELF3 and PRR9/7 in regulating different aspects of plant development. ELF3 was essential for hypocotyl development under ambient and warm temperatures, while PRR9 was critical for root thermomorphogenesis. Finally, mutations in prr9 and prr7 rescued the photoperiod-insensitive flowering phenotype of the elf3 mutant. Together, our results highlight the importance of investigating the genetic relationship among plant circadian genes., Competing Interests: Conflicts of interest The author(s) declare no conflicts of interest., (© The Author(s) 2023. Published by Oxford University Press on behalf of The Genetics Society of America.)

ALKBH proteins, the homologs of Escherichia coli AlkB dioxygenase, constitute a single-protein repair system that safeguards cellular DNA and RNA against the harmful effects of alkylating agents. ALKBH10B, the first discovered N 6 A) demethylase in Arabidopsis ( 6 ), has been shown to regulate plant growth, development, and stress responses. However, until now, the functional role of the plant ALKBH10B has solely been reported in arabidopsis, cotton, and poplar, leaving its functional implications in other plant species shrouded in mystery. In this study, we identified the AlkB homolog SlALKBH10B in tomato ( Arabidopsis thaliana ), has been shown to regulate plant growth, development, and stress responses. However, until now, the functional role of the plant ALKBH10B has solely been reported in arabidopsis, cotton, and poplar, leaving its functional implications in other plant species shrouded in mystery. In this study, we identified the AlkB homolog SlALKBH10B in tomato ( Solanum lycopersicum ) through phylogenetic and gene expression analyses. SlALKBH10B exhibited a wide range of expression patterns and was induced by exogenous abscisic acid (ABA) and abiotic stresses. By employing CRISPR/Cas9 gene editing techniques to knock out SlALKBH10B , we observed an increased sensitivity of mutants to ABA treatment and upregulation of gene expression related to ABA synthesis and response. Furthermore, the Slalkbh10b mutants displayed an enhanced tolerance to drought and salt stress, characterized by higher water retention, accumulation of photosynthetic products, proline accumulation, and lower levels of reactive oxygen species and cellular damage. Collectively, these findings provide insights into the negative impact of SlALKBH10B on drought and salt tolerance in tomato plant, expanding our understanding of the biological functionality of SlALKBH10B .

The orchid is one of the most distinctive and highly valued flowering plants. Nevertheless, the CONSTANS-like ( COL ) gene family plays significant roles in the control of flowering, and its functions in Orchidaceae have been minimally explored. This research identified 68 potential COL genes within seven orchids' complete genome, divided into three groups (groups I, II, and III) via a phylogenetic tree. The modeled three-dimensional structure and the conserved domains exhibited a high degree of similarity among the orchid COL proteins. The selection pressure analysis showed that all orchid COLs suffered a strong purifying selection. Furthermore, the orchid COL genes exhibited functional and structural heterogeneity in terms of collinearity, gene structure, cis-acting elements within their promoters, and expression patterns. Moreover, we identified 50 genes in orchids with a homology to those involved in the COL transcriptional regulatory network in Arabidopsis. Additionally, the first overexpression of CsiCOL05 and CsiCOL09 in Cymbidium sinense protoplasts suggests that they may antagonize the regulation of flowering time and gynostemium development. Our study will undoubtedly provide new resources, ideas, and values for the modern breeding of orchids and other plants.

The efficient infection of plants by the bacteria Xanthomonas campestris pv. campestris (Xcc) depends on its type III effectors (T3Es). Although the functions of AvrE family T3Es have been reported in some bacteria, the member XopAM in Xcc has not been studied. As XopAM has low sequence similarity to reported AvrE-T3Es and different reports have shown that these T3Es have different targets in hosts, we investigated the functions of XopAM in the Xcc-plant interaction. Deletion of xopAM from Xcc reduced its virulence in cruciferous crops but increased virulence in Arabidopsis (Arabidopsis thaliana) Col-0, indicating that XopAM may perform opposite functions depending on the host species. We further found that XopAM is a lipase that may target the cytomembrane and that this activity might be enhanced by its membrane-targeted protein XOPAM-ACTIVATED RESISTANCE 1 (AMAR1) in Arabidopsis Col-0. The binding of XopAM to AMAR1 induced an intense hypersensitive response that restricted Xcc proliferation. Our results showed that the roles of XopAM in Xcc infection are not the same as those of other AvrE-T3Es, indicating that the functions of this type of T3E have differentiated during long-term bacterium‒host interactions., Competing Interests: Conflict of interest statement. The authors declare no competing interests., (© American Society of Plant Biologists 2023. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Salinity is one of the most severe abiotic stresses that adversely affect plant growth and agricultural productivity. The plant Na + /H + antiporter Salt Overly Sensitive 1 (SOS1) located in the plasma membrane extrudes excess Na + out of cells in response to salt stress and confers salt tolerance. However, the molecular mechanism underlying SOS1 activation remains largely elusive. Here we elucidate two cryo-electron microscopy structures of rice (Oryza sativa) SOS1, a full-length protein in an auto-inhibited state and a truncated version in an active state. The SOS1 forms a dimeric architecture, with an NhaA-folded transmembrane domain portion in the membrane and an elongated cytosolic portion of multiple regulatory domains in the cytoplasm. The structural comparison shows that SOS1 adopts an elevator transport mechanism accompanied by a conformational transition of the highly conserved Pro 148 in the unwound transmembrane helix 5 (TM 5 ), switching from an occluded conformation in the auto-inhibited state to a conducting conformation in the active state. These findings allow us to propose an inhibition-release mechanism for SOS1 activation and elucidate how SOS1 controls Na + homeostasis in response to salt stress., (© 2023. The Author(s), under exclusive licence to Springer Nature Limited.)

Salt-overly-sensitive 1 (SOS1) is a unique electroneutral Na + /H + antiporter at the plasma membrane of higher plants and plays a central role in resisting salt stress. SOS1 is kept in a resting state with basal activity and activated upon phosphorylation. Here, we report the structures of SOS1. SOS1 forms a homodimer, with each monomer composed of transmembrane and intracellular domains. We find that SOS1 is locked in an occluded state by shifting of the lateral-gate TM5b toward the dimerization domain, thus shielding the Na + /H + binding site. We speculate that the dimerization of the intracellular domain is crucial to stabilize the transporter in this specific conformation. Moreover, two discrete fragments and a residue W1013 are important to prevent the transition of SOS1 to an alternative conformational state, as validated by functional complementation assays. Our study enriches understanding of the alternate access model of eukaryotic Na + /H + exchangers., (© 2023. The Author(s).)

Brassinosteroid (BR) represents a group of steroid hormones that regulate plant growth and development as well as environmental adaptation. The fluctuation of external nutrient elements is a situation that plants frequently face in the natural environment, in which nitrogen (N) and phosphorus (P) are two of the most critical nutrients restraint of the early growth of plants. As the macronutrients, N and P are highly required by plants, but their availability or solubility in the soil is relatively low. Since iron (Fe) and P always modulate each other's content and function in plants mutually antagonistically, the regulatory mechanisms of Fe and P are inextricably linked. Recently, BR has emerged as a critical regulator in nutrient acquisition and phenotypic plasticity in response to the variable nutrient levels in Arabidopsis and rice. Here, we review the current understanding of the crosstalk between BR and the three major nutrients (N, P, and Fe), highlighting how nutrient signaling regulates BR synthesis and signaling to accommodate plant growth and development in Arabidopsis and rice., (© 2023. Science China Press.)

Abiotic stress has great impacts on plant germination, growth and development and crop yield. Therefore, it is important to understand the molecular mechanism of plants response to abiotic stress. In this study, we identified a plant specific protein AtSIEK (stress-induced protein with EXD1-like domain and KH domain) response to salt stress. AtSIEK encodes a hnRNP K homology (KH) protein localized in nucleus. Amino acid sequences analysis found that SIEK protein is specific in plants, containing two domains with EXD1-like domain and KH domain, while SIEK homolog in animals only had EXD1-like domain without KH domain. Physiology experiments revealed that AtSIEK was significantly induced under salt stress and the siek mutant shows sensitive to salt stress, indicating that AtSIEK was a positive regulator in stress response. Further, molecular, biochemical, and genetic assays suggested that AtSIEK interacts with FRY2/CPL1, a known regulator in response to abiotic stress, and they function synergistically in response to salt stress. Taken together, these results shed new light on the regulation of plant adaption to abiotic stress, which deepen our understanding of the molecular mechanisms of abiotic stress regulation in plants., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Despite extensive investigations in mammals and yeasts, the importance and specificity of COMPASS-like complex, which catalyzes histone 3 lysine 4 methylation (H3K4me), are not fully understood in plants. Here, we report that JMJ28, a Jumonji C domain-containing protein in Arabidopsis, recognizes specific DNA motifs through a plant-specific WRC domain and acts as an interacting factor to guide the chromatin targeting of ATX1/2-containing COMPASS-like complex. JMJ28 associates with COMPASS-like complex in vivo via direct interaction with RBL. The DNA-binding activity of JMJ28 is essential for both the targeting specificity of ATX1/2-COMPASS and the deposition of H3K4me at specific loci but exhibit functional redundancy with alternative COMPASS-like complexes at other loci. Finally, we demonstrate that JMJ28 is a negative regulator of plant immunity. In summary, our findings reveal a plant-specific recruitment mechanism of COMPASS-like complex. These findings help to gain deeper insights into the regulatory mechanism of COMPASS-like complex in plants., Competing Interests: Declaration of interests The authors declare that they have no conflict of interest., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

In Arabidopsis, the receptor-like kinase (RLK) FERONIA (FER) senses peptide ligands in the plasma membrane (PM), modulates plant growth and development, and integrates biotic and abiotic stress signaling for downstream adaptive responses. However, the molecular interplay of these diverse processes is largely unknown. Here, we show that FER, the receptor of Rapid Alkalinization Factor 1 (RALF1), physically interacts with C2 domain ABA-related (CAR) proteins to control the nano-organization of the PM. During this process, the RALF1-FER pathway upregulates CAR protein translation, and then more CAR proteins are recruited to the PM. This acts as a rapid feedforward loop that stabilizes the PM liquid-ordered phase. FER interacts with and phosphorylates CARs, thereby reducing their lipid-binding ability and breaking the feedback regulation at later time points. The formation of the flg22-induced FLS2-BAK1 immune complex, which depends on the integrity of FER-containing nanodomains, is impaired in fer and pentuple car14569 mutant. Together, we propose that the FER-CAR module controls the formation of PM nano-organization during RALF signaling through a self-contained amplifying loop including both positive and negative feedback., (© 2022 The Authors. New Phytologist © 2022 New Phytologist Foundation.)

Orchids are among the most precious flowers in the world. Regulation of flowering time is one of the most important targets to enhance their ornamental value. The beauty of Arundina graminifolia is its year-round flowering, although the molecular mechanism of this flowering ability remains masked. Therefore, we performed a comprehensive assessment to integrate transcriptome and miRNA sequencing to disentangle the genetic regulation of flowering in this valuable species. Clustering analyses provided a set of molecular regulators of floral transition and floral morphogenesis. We mined candidate floral homeotic genes, including FCA , FPA , GI , FT , FLC , AP2 , SOC1 , SVP , GI , TCP , and CO , which were targeted by a variety of miRNAs. MiR11091 targeted the highest number of genes, including candidate regulators of phase transition and hormonal control. The conserved miR156-miR172 pathway of floral time regulation was evident in our data, and we found important targets of these miRNAs in the transcriptome. Moreover, endogenous hormone levels were determined to decipher the hormonal control of floral buds in A. graminifolia . The qRT-PCR analysis of floral and hormonal integrators validated the transcriptome expression. Therefore, miRNA-mediated mining of candidate genes with hormonal regulation forms the basis for comprehending the complex regulatory network of perpetual flowering in precious orchids. The findings of this study can do a great deal to broaden the breeding programs for flowering time manipulation of orchids.

Protein-targeting technologies represent essential approaches in biological research. Protein knockdown tools developed recently in mammalian cells by exploiting natural degradation mechanisms allow for precise determination of protein function and discovery of degrader-type drugs. However, no method to directly target endogenous proteins for degradation is currently available in plants. Here, we describe a novel method for targeted protein clearance by engineering an autophagy receptor with a binder to provide target specificity and an ATG8-binding motif (AIM) to link the targets to nascent autophagosomes, thus harnessing the autophagy machinery for degradation. We demonstrate its specificity and broad potentials by degrading various fluorescence-tagged proteins, including cytosolic mCherry, the nucleus-localized bZIP transcription factor TGA5, and the plasma membrane-anchored brassinosteroid receptor BRI1, as well as fluorescence-coated peroxisomes, using a tobacco-based transient expression system. Stable expression of AIM-based autophagy receptors in Arabidopsis further confirms the feasibility of this approach in selective autophagy of endogenous proteins. With its wide substrate scope and its specificity, our concept of engineered AIM-based selective autophagy could provide a convenient and robust research tool for manipulating endogenous proteins in plants and may open an avenue toward degradation of cytoplasmic components other than proteins in plant research., (© 2022 The Authors New Phytologist © 2022 New Phytologist Foundation.)

AKT1 is an inward-rectifying K + channel that was originally thought to function only within a low-affinity K + concentration range. However, the growth of an akt1 mutant of Arabidopsis was shown to be severely inhibited within a high-affinity range. This suggested that AKT1 may also be a high-affinity K + transporter, but it remains unclear how the two modes of AKT1 coordinate to uptake K + . One gene (MeAKT2) encodes for a putatively inward-rectifying K + channel and was isolated from cassava. Relative to other tissues, the MeAKT2 gene was expressed mainly in roots, and its transcriptional level was observed to be significantly increased under low-K + conditions. Functional analyses were performed using a yeast expression system. When MeAKT2 was expressed alone in yeast cells, transgenic yeast could grow only in nutrient media supplied with >0.5 mM potassium. A yeast two-hybrid assay showed that both MeCIPK10 and MeCIPK12 clearly interacted with MeAKT2. Additionally, 0.05 mM K + was sufficient for the growth of yeast cells co-expressing MeAKT2 with MeCIPK10, but also their co-expression significantly enhanced the growth capacity of yeast cells in the low range of K + concentrations. Change in K + uptake rate in co-transgenic yeast cells grown across a wide range of K + concentrations showed that MeAKT2-mediated K + uptake displayed a biphasic pattern, but also the switching from low-to high-affinity K + uptake was regulated by CIPK10. This indicated that MeAKT2 functioned as a dual-affinity transporter to uptake K + under both low- and high-affinity K + conditions., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier GmbH. All rights reserved.)

APETALA2/Ethylene Responsive Factor ( AP2/ERF ) family plays important roles in reproductive development, stress responses and hormone responses in plants. However, AP2/ERF family has not been systematically studied in Dendrobium catenatum . In this study, 120 AP2/ERF family members were identified for the first time in D. catenatum , which were divided into four groups ( AP2 , RAV , ERF and DREB subfamily) according to phylogenetic analysis. Gene structures and conserved motif analysis showed that each DcAP2/ERF family gene contained at least one AP2 domain, and the distribution of motifs varied among subfamilies. Cis-element analysis indicated that DcAP2/ERF genes contained abundant cis-elements related to hormone signaling and stress response. To further identify potential genes involved in drought stress, 12 genes were selected to detect their expression under drought treatment through qRT-PCR analysis and DcAP2/ERF#96 , a nuclear localized ethylene-responsive transcription factor, showed a strong response to PEG treatment. Overexpression of DcAP2/ERF#96 in Arabidopsis showed sensitivity to ABA. Molecular, biochemical and genetic assays indicated that DcAP2ERF#96 interacts with DREB2A and directly inhibits the expression of P5CS1 in response to the ABA signal. Taken together, our study provided a molecular basis for the intensive study of DcAP2/ERF genes and revealed the biological function of DcAP2ERF#96 involved in the ABA signal.

The RPW8s (Resistance to Powdery Mildew 8) are atypical broad-spectrum resistance genes that provide resistance to the powdery mildew fungi. Powdery mildew of rubber tree is one of the serious fungal diseases that affect tree growth and latex production. However, the RPW8 homologs in rubber tree and their role of resistance to powdery mildew remain unclear. In this study, four RPW8 genes, HbRPW8-a, b, c, d, were identified in rubber tree, and phylogenetic analysis showed that HbRPW8-a was clustered with AtRPW8.1 and AtRPW8.2 of Arabidopsis. The HbRPW8-a protein was localized on the plasma membrane and its expression in rubber tree was significantly induced upon powdery mildew infection. Transient expression of HbRPW8-a in tobacco leaves induced plant immune responses, including the accumulation of reactive oxygen species and the deposition of callose in plant cells, which was similar to that induced by AtRPW8.2 . Consistently, overexpression of HbRPW8-a in Arabidopsis thaliana enhanced plant resistance to Erysiphe cichoracearum UCSC1 and Pseudomonas syringae pv. tomato DC30000 ( Pst DC3000). Moreover, such HbRPW8-a mediated resistance to powdery mildew was in a salicylic acid (SA) dependent manner. Taken together, we demonstrated a new RPW8 member in rubber tree, HbRPW8-a, which could potentially contribute the resistance to powdery mildew.

Endoreduplication is prevalent during plant growth and development, and is often correlated with large cell and organ size. Despite its prevalence, the transcriptional regulatory mechanisms underlying the transition from mitotic cell division to endoreduplication remain elusive. Here, we characterize ETHYLENE-RESPONSIVE ELEMENT BINDING FACTOR 4 (ERF4) as a positive regulator of endoreduplication through its function as a transcriptional repressor. ERF4 was specifically expressed in mature tissues in which the cells were undergoing expansion, but was rarely expressed in young organs. Plants overexpressing ERF4 exhibited much larger cells and organs, while plants that lacked functional ERF4 displayed smaller organs than the wild-type. ERF4 was further shown to regulate cell size by controlling the endopolyploidy level in the nuclei. Moreover, ERF4 physically associates with the class I TEOSINTE BRANCHED 1/CYCLOIDEA/PCF (TCP) protein TCP15, a transcription factor that inhibits endoreduplication by activating the expression of a key cell-cycle gene, CYCLIN A2;3 (CYCA2;3). A molecular and genetic analysis revealed that ERF4 promotes endoreduplication by directly suppressing the expression of CYCA2;3. Together, this study demonstrates that ERF4 and TCP15 function as a module to antagonistically regulate each other's activity in regulating downstream genes, thereby controlling the switch from the mitotic cell cycle to endoreduplication during leaf development. These findings expand our understanding of how the control of the cell cycle is fine-tuned by an ERF4-TCP15 transcriptional complex., (© 2022 Institute of Botany, Chinese Academy of Sciences.)

During evolutionary adaptation, the mechanisms for self-regulation are established between the normal growth and development of plants and environmental stress. The phytohormone jasmonate (JA) is a key tie of plant defence and development, and JASMONATE-ZIM DOMAIN (JAZ) repressor proteins are key components in JA signalling pathways. Here, we show that JAZ expression was affected by leaf senescence from the transcriptomic data. Further investigation revealed that SlJAZ10 and SlJAZ11 positively regulate leaf senescence and that SlJAZ11 can also promote plant regeneration. Moreover, we reveal that the SlJAV1-SlWRKY51 (JW) complex could suppress JA biosynthesis under normal growth conditions. Immediately after injury, SlJAZ10 and SlJAZ11 can regulate the activity of the JW complex through the effects of electrical signals and Ca2+ waves, which in turn affect JA biosynthesis, causing a difference in the regeneration phenotype between SlJAZ10-OE and SlJAZ11-OE transgenic plants. In addition, SlRbcs-3B could maintain the protein stability of SlJAZ11 to protect it from degradation. Together, SlJAZ10 and SlJAZ11 not only act as repressors of JA signalling to leaf senescence, but also regulate plant regeneration through coordinated electrical signals, Ca2+ waves, hormones and transcriptional regulation. Our study provides critical insights into the mechanisms by which SlJAZ11 can induce regeneration., Competing Interests: The authors have declared that no competing interests exist.

Chloroplasts are hypersensitive to heat stress (HS). SUMOylation, a critical post-translational modification, is conservatively involved in HS responses. However, the functional connection between SUMOylation and chloroplasts under HS remains to be studied. The bioinformatics, biochemistry, and cell biology analyses were used to detect the SUMOylation statuses of Arabidopsis nuclear-encoded chloroplast proteins and the effect of SUMOylation on subcellular localization of these proteins under HS. PSBR, a subunit of photosystem II, was used as an example for a detailed investigation of functional mechanisms. After a global SUMOylation site prediction of nuclear-encoded chloroplast proteins, biochemical data showed that most of the selected candidates are modified by SUMO3 in the cytoplasm. The chloroplast localization of these SUMOylation targets under long-term HS is partially maintained by the SUMO ligase AtSIZ1. The HS-induced SUMOylation on PSBR contributes to the maintenance of its chloroplast localization, which is dependent on its chloroplast importation efficiency correlated to phosphorylation. The complementation analysis provided evidence that SUMOylation is essential for the physiological function of PSBR under HS. Our study illustrated a general regulatory mechanism of SUMOylation in maintaining the chloroplast protein importation during HS and provided hints for further investigation on protein modifications associated with plant organelles under stress conditions., (© 2022 The Authors. New Phytologist © 2022 New Phytologist Foundation.)

Understanding the complex functions of plant leaves requires a thorough characterization of discrete cell features. Although single-cell gene expression profiling technologies have been developed, their application in characterizing cell subtypes has not been achieved yet. Here, we present scStereo-seq (single-cell spatial enhanced resolution omics sequencing) that enabled us to show the bona fide single-cell spatial transcriptome profiles of Arabidopsis leaves. Subtle but significant transcriptomic differences between upper and lower epidermal cells have been successfully distinguished. Furthermore, we discovered cell-type-specific gene expression gradients from the main vein to the leaf edge, which led to the finding of distinct spatial developmental trajectories of vascular cells and guard cells. Our study showcases the importance of physical locations of individual cells for exerting complex biological functions in plants and demonstrates that scStereo-seq is a powerful tool to integrate single-cell location and transcriptome information for plant biology study., Competing Interests: Declaration of interests The authors declare the following competing interests: the chip, procedure, and applications of Stereo-seq are covered in pending patents. Ye Yin is an employee and shareholder of BGI Genomics., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

Ubiquitination-mediated protein degradation in both the 26 S proteasome and vacuole is an important process in abscisic acid (ABA) signaling. However, the role of deubiquitination in this process remains elusive. Here, we demonstrate that two deubiquitinating enzymes (DUBs), ubiquitin-specific protease 12 (UBP12) and UBP13, modulate ABA signaling and drought tolerance by deubiquitinating and stabilizing the endosomal sorting complex required for transport-I (ESCRT-I) component vacuolar protein sorting 23A (VPS23A) and thereby affect the stability of ABA receptors in Arabidopsis thaliana . Genetic analysis showed that VPS23A overexpression could rescue the ABA hypersensitive and drought tolerance phenotypes of ubp12-2w or ubp13-1 . In addition to the direct regulation of VPS23A, we found that UBP12 and UBP13 also stabilized the E3 ligase XB3 ortholog 5 in A. thaliana (XBAT35.2) in response to ABA treatment. Hence, we demonstrated that UBP12 and UBP13 are previously unidentified rheostatic regulators of ABA signaling and revealed a mechanism by which deubiquitination precisely monitors the XBAT35/VPS23A ubiquitination module in the ABA response.

The microbiome plays an important role in shaping plant growth and immunity, but few plant genes and pathways impacting plant microbiome composition have been reported. In Arabidopsis thaliana, the phosphate starvation response (PSR) was recently found to modulate the root microbiome upon phosphate (Pi) starvation through the transcriptional regulator PHR1. Here, we report that A. thaliana PHR1 directly binds to the promoters of rapid alkalinization factor (RALF) genes, and activates their expression under phosphate-starvation conditions. RALFs in turn suppress complex formation of pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) receptor through FERONIA, a previously-identified PTI modulator that increases resistance to certain detrimental microorganisms. Suppression of immunity via the PHR1-RALF-FERONIA axis allows colonization by specialized root microbiota that help to alleviate phosphate starvation by upregulating the expression of PSR genes. These findings provide a new paradigm for coordination of host-microbe homeostasis through modulating plant innate immunity after environmental perturbations., (© 2022 The Authors.)

The transcription factors of the MYB family are involved in plant growth and development and responses to biotic and abiotic stresses. Here, we isolated the R2R3-MYB transcription factor gene SlMYB55 and found that it is responsive to abscisic acid (ABA), drought, and salt stress. Notably, the expression levels of multiple stress-related and inflorescence and flowering time-related genes were changed in SlMYB55-RNAi plants compared to wild-type plants. Transient tobacco expression experiments indicated that SlMYB55 directly targets the WUS and 4CL genes to regulate the development of inflorescence and flavonoid biosynthesis. Yeast two-hybrid experiments showed that the SlMYB55 protein interacts with the MADS-box family protein MBP21. Based on these results, we concluded that SlMYB55 affects the biosynthesis of ABA, regulates drought and salt responses through ABA-mediated signal transduction pathways, and directly or indirectly affects the expression of genes related to drought and salt response, flowering time, sepal size and inflorescence, thereby regulating stress tolerance and flower development. In summary, this study identified essential roles for SlMYB55 in regulating drought and salt tolerance and flower development., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Split firefly luciferase complementation assay (FLCA) is one of the most widely used sensitive and reliable methods for the analysis of constitutive and dynamic protein-protein interactions (PPIs). Here, we report a method for long-term in vivo detects plant protein-protein interactions in Arabidopsis F1 hybrids via Topcount™ Microplate Scintillation Counter or Deep-Cooled CCD camera. Following these protocols, we successfully detected time-dependent PPIs of EARLY FLOWERING 3 (ELF3) and EARLY FLOWERING 4 (ELF4); both of them with LUX ARRHYTHMO (LUX) belong to an evening complex which has been found to play a key role in circadian rhythms, flowering, and growth., (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Protein S-acylation is an important post-translational modification in eukaryotes, regulating the subcellular localization, trafficking, stability, and activity of substrate proteins. The dynamic regulation of this reversible modification is mediated inversely by protein S-acyltransferases and de-S-acylation enzymes, but the de-S-acylation mechanism remains unclear in plant cells. Here, we characterized a group of putative protein de-S-acylation enzymes in Arabidopsis thaliana, including 11 members of Alpha/Beta Hydrolase Domain-containing Protein 17-like acyl protein thioesterases (ABAPTs). A robust system was then established for the screening of de-S-acylation enzymes of protein substrates in plant cells, based on the effects of substrate localization and confirmed via the protein S-acylation levels. Using this system, the ABAPTs, which specifically reduced the S-acylation levels and disrupted the plasma membrane localization of five immunity-related proteins, were identified respectively in Arabidopsis. Further results indicated that the de-S-acylation of RPM1-Interacting Protein 4, which was mediated by ABAPT8, resulted in an increase of cell death in Arabidopsis and Nicotiana benthamiana, supporting the physiological role of the ABAPTs in plants. Collectively, our current work provides a powerful and reliable system to identify the pairs of plant protein substrates and de-S-acylation enzymes for further studies on the dynamic regulation of plant protein S-acylation., (� American Society of Plant Biologists 2021. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Endoplasmic reticulum-associated degradation (ERAD) is known to regulate plant responses to diverse stresses, yet its underlying molecular mechanisms and links to various stress signaling pathways are poorly understood. Here, we show that the ERAD component ubiquitin-conjugating enzyme UBC32 positively regulates drought tolerance in Arabidopsis thaliana by targeting the aquaporins PIP2;1 and PIP2;2 for degradation. Furthermore, we demonstrate that the RING-type ligase Rma1 acts together with UBC32 and that the E2 activity of UBC32 is essential for the ubiquitination of Rma1. This complex ubiquitinates a phosphorylated form of PIP2;1 at Lys276 to promote its degradation, thereby enhancing plant drought tolerance. Extending these molecular insights into crops, we show that overexpression of Arabidopsis UBC32 also improves drought tolerance in rice (Oryza sativa). Thus, beyond uncovering the molecular basis of an ERAD-regulated stress response, our study suggests multiple potential strategies for engineering crops with improved drought tolerance., (© American Society of Plant Biologists 2021. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

The core plant circadian oscillator is composed of multiple interlocked transcriptional-translational feedback loops, which synchronize endogenous diel physiological rhythms to the cyclic changes of environmental cues. PSEUDO-RESPONSE REGULATORS (PRRs) have been identified as negative components in the circadian clock, though their underlying molecular mechanisms remain largely unknown. Here, we found that a subfamily of zinc finger transcription factors, B-box (BBX)-containing proteins, have a critical role in fine-tuning circadian rhythm. We demonstrated that overexpressing Arabidopsis thaliana BBX19 and BBX18 significantly lengthened the circadian period, while the null mutation of BBX19 accelerated the circadian speed. Moreover, BBX19 and BBX18, which are expressed during the day, physically interacted with PRR9, PRR7, and PRR5 in the nucleus in precise temporal ordering from dawn to dusk, consistent with the respective protein accumulation pattern of PRRs. Our transcriptomic and genetic analysis indicated that BBX19 and PRR9, PRR7, and PRR5 cooperatively inhibited the expression of morning-phased clock genes. PRR proteins affected BBX19 recruitment to the CCA1, LHY, and RVE8 promoters. Collectively, our findings show that BBX19 interacts with PRRs to orchestrate circadian rhythms, and suggest the indispensable role of transcriptional regulators in fine-tuning the circadian clock., (© The Author(s) 2021. Published by Oxford University Press on behalf of American Society of Plant Biologists.)

The transcriptional regulators of arsenic-induced gene expression remain largely unknown. Sulfur assimilation is tightly linked with arsenic detoxification. Here, we report that mutant alleles in the SLIM1 transcription factor are substantially more sensitive to arsenic than cadmium. Arsenic treatment caused high levels of oxidative stress in the slim1 mutants, and slim1 alleles were impaired in both thiol accumulation and sulfate accumulation. We further found enhanced arsenic accumulation in roots of slim1 mutants. Transcriptome analyses indicate an important role for SLIM1 in arsenic-induced tolerance mechanisms. The present study identifies the SLIM1 transcription factor as an essential component in arsenic tolerance and arsenic-induced gene expression. Our results suggest that the severe arsenic sensitivity of the slim1 mutants is caused by altered redox status., (© 2021 The Authors. FEBS Letters published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

Stomata in leaves regulate gas exchange between the plant and its atmosphere. Various environmental stimuli elicit abscisic acid (ABA); ABA leads to phosphoactivation of slow anion channel 1 (SLAC1); SLAC1 activity reduces turgor pressure in aperture-defining guard cells; and stomatal closure ensues. We used electrophysiology for functional characterizations of Arabidopsis thaliana SLAC1 ( At SLAC1) and cryoelectron microscopy (cryo-EM) for structural analysis of Brachypodium distachyon SLAC1 ( Bd SLAC1), at 2.97-Å resolution. We identified 14 phosphorylation sites in At SLAC1 and showed nearly 330-fold channel-activity enhancement with 4 to 6 of these phosphorylated. Seven SLAC1-conserved arginines are poised in Bd SLAC1 for regulatory interaction with the N-terminal extension. This Bd SLAC1 structure has its pores closed, in a basal state, spring loaded by phenylalanyl residues in high-energy conformations. SLAC1 phosphorylation fine-tunes an equilibrium between basal and activated SLAC1 trimers, thereby controlling the degree of stomatal opening., Competing Interests: The authors declare no competing interest.

Arsenic stress causes rapid transcriptional responses in plants. However, transcriptional regulators of arsenic-induced gene expression in plants remain less well known. To date, forward genetic screens have proven limited for dissecting arsenic response mechanisms. We hypothesized that this may be due to the extensive genetic redundancy present in plant genomes. To overcome this limitation, we pursued a forward genetic screen for arsenite tolerance using a randomized library of plants expressing >2,000 artificial microRNAs (amiRNAs). This library was designed to knock-down diverse combinations of homologous gene family members within sub-clades of transcription factor and transporter gene families. We identified six transformant lines showing an altered response to arsenite in root growth assays. Further characterization of an amiRNA line targeting closely homologous CBF and ERF transcription factors show that the CBF1,2 and 3 transcription factors negatively regulate arsenite sensitivity. Furthermore, the ERF34 and ERF35 transcription factors are required for cadmium resistance. Generation of CRISPR lines, higher-order T-DNA mutants and gene expression analyses, further support our findings. These ERF transcription factors differentially regulate arsenite sensitivity and cadmium tolerance., (© 2021 John Wiley & Sons Ltd.)

The gaseous phytohormone ethylene mediates numerous aspects of plant growth and development as well as stress responses. The F-box proteins EIN3-binding F-box protein 1 (EBF1) and EBF2 are key components that ubiquitinate and degrade the master transcription factors ethylene insensitive 3 (EIN3) and EIN3-like 1 (EIL1) in the ethylene response pathway. Notably, EBF1 and EBF2 themselves undergo the 26S proteasome-mediated proteolysis induced by ethylene and other stress signals. However, despite their importance, little is known about the mechanisms regulating the degradation of these proteins. Here, we show that a really interesting new gene (RING)-type E3 ligase, salt- and drought-induced ring finger 1 (SDIR1), positively regulates the ethylene response and promotes the accumulation of EIN3. Further analyses indicate that SDIR1 directly interacts with EBF1/EBF2 and targets them for ubiquitination and proteasome-dependent degradation. We show that SDIR1 is required for the fine tuning of the ethylene response to ambient temperature changes by mediating temperature-induced EBF1/EBF2 degradation and EIN3 accumulation. Thus, our work demonstrates that SDIR1 functions as an important modulator of ethylene signaling in response to ambient temperature changes, thereby enabling plant adaptation under fluctuating environmental conditions., Competing Interests: The authors declare no competing interest.

Key Message: CCA1α and CCA1β protein variants respond to environmental light and temperature cues, and higher temperature promotes CCA1β protein production and causes its retention detectable in the cytoplasm. CIRCADIAN CLOCK ASSOCIATED1 (CCA1), as the core transcription factor of circadian clock, is involved in the regulation of endogenous circadian rhythm in Arabidopsis. Previous studies have shown that CCA1 consists of two abundant splice variants, fully spliced CCA1α and intron-retaining CCA1β. CCA1β is believed to form a nonfunctional heterodimer with CCA1α and its closed-related homolog LHY. Many studies have established that CCA1β is a transcription product, while how CCA1β protein is produced and how two CCA1 isoforms respond to environmental cues have not been elucidated. In this study, we identified CCA1α and CCA1β protein variants under different photoperiods with warm or cold temperature cycles, respectively. Our results showed that CCA1 protein production is regulated by prolonged light exposure and warm temperature. The protein levels of CCA1α and CCA1β peak in the morning, but the detection of CCA1β is dependent on immunoprecipitation enrichment at 22 °C. Higher temperature of 37 °C promotes CCA1β protein production and causes its retention to be detectable in the cytoplasm. Overall, our results indicate that two splice variants of the CCA1 protein respond to environmental light and temperature signals and may, therefore, maintain the circadian rhythms and give individuals the ability to adapt to environment.

Circadian clock operates autonomously in each cell and drives the approximately 24-h rhythm in individual tissues and organs. It is known that the evening complex (EC) components and GI are required for ambient temperature perception and thermomorphogenesis in higher plants. Our previous study found that PRR9 and 7 are required for the lengthened period of the circadian rhythm in roots, and they are responsible for the temperature overcompensation in shoots. However, the molecular mechanism of the circadian clock, especially in different tissues, in response to temperature oscillations remains largely unknown. Here, we studied the transcript levels of EC genes and GI of the prr7 prr9 mutant shoots and roots in response to 22°C or 28°C, respectively. The results showed that PRR9 , 7 in roots inhibited the transcripts accumulation of ELF3, ELF4 , and LUX at 28°C. In addition, loss-of-function of both PRR9 and 7 caused an increase in GI expression at 22°C, but warm temperature of 28°C limited the negative effect of PRR9 , 7 on GI in roots. Our findings proposed a temperature-dependent molecular basis for root-specific circadian clock and indicated the critical role for PRR9 , 7 in negatively regulating ELF3, ELF4, LUX , and GI in the circadian gating of thermoresponse.

Trichomes are specialized epidermal appendages that serve as excellent models to study cell morphogenesis. Although the molecular mechanism underlying trichome morphogenesis in Arabidopsis has been well characterized, most of the regulators essential for multicellular trichome morphology remain unknown in tomato. In this study, we determined that the recessive hairless-2 (hl-2) mutation in tomato causes severe distortion of all trichome types, along with increased stem fragility. Using map-based cloning, we found that the hl-2 phenotype was associated with a 100 bp insertion in the coding region of Nck-associated protein 1, a component of the SCAR/WAVE complex. Direct protein-protein interaction was detected between Hl-2 and Hl (SRA1, specifically Rac1-associated protein) using yeast two-hybrid and co-immunoprecipitation assays, suggesting that these proteins may work together during trichome formation. In addition, knock-down of a HD-Zip IV transcription factor, HDZIPIV8, distorted trichomes similar to the hl-2 mutant. HDZIPIV8 regulates the expression of Hl-2 by binding to the L1-box in the Hl-2 promoter region, and is involved in organizing actin filaments. The brittleness of hl-2 stems was found to result from decreased cellulose content. Taken together, these findings suggest that the Hl-2 gene plays an important role in controlling multicellular trichome morphogenesis and mechanical properties of stems in tomato plants., (© The Author(s) 2020. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Abscisic acid (ABA) is the key phytohormone in plant drought tolerance and stress adaptation. The clade A protein phosphatase 2Cs (PP2Cs) like ABI1 (ABA-INSENSITIVE 1) work as coreceptors of ABA and regulate multiple ABA responses. Ubiquitination of ABI1 has been proven to play important regulatory roles in ABA signaling. However, the specific ubiquitin conjugating enzyme (E2) involved is unknown. Here, we report that UBC27 is an active E2 that positively regulates ABA signaling and drought tolerance. UBC27 forms the E2-E3 pair with the drought regulator RING E3 ligase AIRP3. Both UBC27 and AIRP3 interact with ABI1 and affect the ubiquitination and degradation of ABI1. ABA activates the expression of UBC27 , inhibits the proteasome degradation of UBC27, and enhances the interaction between UBC27 and ABI1 to increase its activity. These findings uncover a regulatory mechanism in ABA signaling and drought response and provide a further understanding of the plant ubiquitination system and ABA signaling pathway., Competing Interests: The authors declare no competing interest.

A myriad of abiotic stress responses in plants are controlled by abscisic acid (ABA) signaling. ABA receptors can be degraded by both the 26S proteasome pathway and vacuolar degradation pathway after processing via the endosomal sorting complex required for transport (ESCRT) proteins. Despite being essential for ABA signaling, the upstream regulators of ESCRTs remain unknown. Here, we report that the ESCRT-I component VPS23A is an unstable protein that is degraded via the ubiquitin-proteasome system (UPS). The UEV domain of VPS23A physically interacts with the two PSAP motifs of XBAT35, an E3 ubiquitin ligase, and this interaction results in the deposition of K48 polyubiquitin chains on VPS23A, marking it for degradation by 26S proteasomes. We showed that XBAT35 in plants is a positive regulator of ABA responses that acts via the VPS23A/PYL4 complex, specifically by accelerating VPS23A turnover and thereby increasing accumulation of the ABA receptor PYL4. This work deciphers how an ESCRT component is regulated in plants and deepens our understanding of plant stress responses by illustrating a mechanism whereby crosstalk between the UPS and endosome-vacuole-mediated degradation pathways controls ABA signaling., (Copyright © 2020 The Author. Published by Elsevier Inc. All rights reserved.)

Flowering plants and mammals contain imprinted genes that are primarily expressed in the endosperm and placenta in a parent-of-origin manner. In this study, we show that early activation of the geminivirus genes C2 and C3 in Arabidopsis ( Arabidopsis thaliana ) plants, encoding a viral suppressor of RNA interference and a replication enhancer protein, respectively, is correlated with the transient vegetative expression of VARIANT IN METHYLATION5 ( VIM5 ), an endosperm imprinted gene that is conserved in diverse plant species. VIM5 is a ubiquitin E3 ligase that directly targets the DNA methyltransferases MET1 and CMT3 for degradation by the ubiquitin-26S proteasome proteolytic pathway. Infection with Beet severe curly top virus induced VIM5 expression in rosette leaf tissues, possibly via the expression of the viral replication initiator protein, leading to the early activation of C2 and C3 coupled with reduced symmetric methylation in the C2-3 promoter and the onset of disease symptoms. These findings demonstrate how this small DNA virus recruits a host imprinted gene for the epigenetic activation of viral gene transcription. Our findings reveal a distinct strategy used by plant pathogens to exploit the host machinery in order to inhibit methylation-mediated defense responses when establishing infection., (© 2020 American Society of Plant Biologists. All rights reserved.)

Plants can be simultaneously exposed to multiple stresses. The interplay of abiotic and biotic stresses may result in synergistic or antagonistic effects on plant development and health. Temporary drought stress can stimulate plant immunity; however, the molecular mechanism of drought-induced immunity is largely unknown. In this study, we demonstrate that cysteine protease RD21A is required for drought-induced immunity. Temporarily drought-treated wild-type Arabidopsis plants became more sensitive to the bacterial pathogen-associated molecular pattern flg22, triggering stomatal closure, which resulted in increased resistance to Pseudomonas syringae pv. tomato DC3000 (Pst-DC3000). Knocking out rd21a inhibited flg22-triggered stomatal closure and compromised the drought-induced immunity. Ubiquitin E3 ligase SINAT4 interacted with RD21A and promoted its degradation in vivo. The overexpression of SINAT4 also consistently compromised the drought-induced immunity to Pst-DC3000. A bacterial type III effector, AvrRxo1, interacted with both SINAT4 and RD21A, enhancing SINAT4 activity and promoting the degradation of RD21A in vivo. Therefore, RD21A could be a positive regulator of drought-induced immunity, which could be targeted by pathogen virulence effectors during pathogenesis., (Published by Oxford University Press on behalf of the Society for Experimental Biology 2020.)

Small nuclear RNAs (snRNAs) are the basal components of the spliceosome and play crucial roles in splicing. Their biogenesis is spatiotemporally regulated. However, related mechanisms are still poorly understood. Defective in snRNA processing (DSP1) is an essential component of the DSP1 complex that catalyzes plant snRNA 3'-end maturation by cotranscriptional endonucleolytic cleavage of the primary snRNA transcripts (presnRNAs). Here, we show that DSP1 is subjected to alternative splicing in pollens and embryos, resulting in two splicing variants, DSP1 α and DSP1 β. Unlike DSP1α, DSP1β is not required for presnRNA 3'-end cleavage. Rather, it competes with DSP1α for the interaction with CPSF73-I, the catalytic subunit of the DSP1 complex, which promotes efficient release of CPSF73-I and the DNA-dependent RNA polymerease II (Pol II) from the 3' end of snRNA loci thereby facilitates snRNA transcription termination, resulting in increased snRNA levels in pollens. Taken together, this study uncovers a mechanism that spatially regulates snRNA accumulation., Competing Interests: The authors declare no competing interest.

The Salt-Overly-Sensitive (SOS) signaling module, comprising the sodium-transport protein SOS1 and the regulatory proteins SOS2 and SOS3, is well known as the central salt excretion system, which helps protect plants against salt stress. Here we report that VPS23A, a component of the ESCRT (endosomal sorting complex required for transport), plays an essential role in the function of the SOS module in conferring plant salt tolerance. VPS23A enhances the interaction of SOS2 and SOS3. In the presence of salt stress, VPS23A positively regulates the redistribution of SOS2 to the plasma membrane, which then activates the antiporter activity of SOS1 to reduce Na + accumulation in plant cells. Genetic evidence demonstrated that plant salt tolerance achieved by the overexpression of SOS2 and SOS3 dependeds on VPS23A. Taken together, our results revealed that VPS23A is a crucial regulator of the SOS module and affects the localization of SOS2 to the cell membrane. Moreover, the strong salt tolerance of Arabidopsis seedlings conferred by the engineered membrane-bound SOS2 revealed the significance of SOS2 sorting to the cell membrane in achieving its function, providing a potential strategy for crop salt tolerance engineering., (Copyright © 2020 The Author. Published by Elsevier Inc. All rights reserved.)

Strigolactones (SLs) and karrikins (KARs) are related butenolide signaling molecules that control plant development. In Arabidopsis ( Arabidopsis thaliana ), they are recognized separately by two closely related receptors but use the same F-box protein MORE AXILLARY GROWTH2 (MAX2) for signal transduction, targeting different members of the SMAX1-LIKE (SMXL) family of transcriptional repressors for degradation. Both signals inhibit hypocotyl elongation in seedlings, raising the question of whether signaling is convergent or parallel. Here, we show that synthetic SL analog GR24 4DO enhanced the interaction between the SL receptor DWARF14 (D14) and SMXL2, while the KAR surrogate GR24 ent -5DS induced association of the KAR receptor KARRIKIN INSENSITIVE2 (KAI2) with SMAX1 and SMXL2. Both signals trigger polyubiquitination and degradation of SMXL2, with GR24 4DO dependent on D14 and GR24 ent -5DS dependent mainly on KAI2. SMXL2 is critical for hypocotyl responses to GR24 4DO and functions redundantly with SMAX1 in hypocotyl response to GR24 ent -5DS Furthermore, GR24 4DO induced response of D14-LIKE2 and KAR-UP F-BOX1 through SMXL2, whereas GR24 ent -5DS induced expression of these genes via both SMAX1 and SMXL2. These findings demonstrate that both SLs and KARs could trigger polyubiquitination and degradation of SMXL2, thus uncovering an unexpected but important convergent pathway in SL- and KAR-regulated gene expression and hypocotyl elongation., (© 2020 American Society of Plant Biologists. All rights reserved.)

The circadian pacemaker in plants is a hierarchical multioscillator system that directs and maintains a 24-hr oscillation required for organism homeostasis and environmental fitness. Molecular clockwork within individual tissues and organs acts cell autonomously, showing differences in circadian expression of core oscillators and their target genes; there are functional dominance and coupling in the complex regulatory network. However, molecular characteristics of organ-specific clocks are still unknown. Here, we showed the detached shoot and root possess dynamic circadian protein-protein interactions between clock core components, periodicity in organs exhibits a difference. The period length difference between shoot and root was not remarkable in prr7-3 and prr7-3 prr9-1 mutants. In addition, the phase transition curve indicated that shoot and root clock respond differently to the resetting cues of ambient temperature. PRR9 and PRR7 compensate circadian period between 22°C and 28°C in shoot, not in root. The circadian rhythms of PRR9 or PRR7 transcript accumulation showed no difference at 22°C and 28°C in shoot, but differences were observed in root. In summary, our results reveal the specificity of dynamic circadian protein-protein interactions in organ-autonomous clocks and the critical roles of PRR9 and PRR7 in mechanisms regulating temperature compensation in aerial shoot system., (© 2020 John Wiley & Sons Ltd.)

Salt tolerance in plants is mediated by Na+ extrusion from the cytosol by the plasma membrane Na+/H+ antiporter SOS1. This is activated in Arabidopsis root by the protein kinase complex SOS2-SOS3 and in Arabidopsis shoot by the protein kinase complex CBL10-SOS2, with SOS2 as a key node in the two pathways. The sos1 mutant is more sensitive than the sos2 mutant, suggesting that other partners may positively regulate SOS1 activity. Arabidopsis has 26 CIPK family proteins of which CIPK8 is the closest homolog to SOS2. It is hypothesized that CIPK8 can activate Na+ extrusion by SOS1 similarly to SOS2. The plasma membrane Na+/H+ exchange activity of transgenic yeast co-expressing CBL10, CIPK8, and SOS1 was higher than that of untransformed and SOS1 transgenic yeast, resulting in a lower Na+ accumulation and a better growth phenotype under salinity. However, CIPK8 could not interact with SOS3, and the co-expression of SOS3, CIPK8, and SOS1 in yeast did not confer a significant salt tolerance phenotype relative to SOS1 transgenic yeast. Interestingly, cipk8 displayed a slower Na+ efflux, a higher Na+ level, and a more sensitive phenotype than wild-type Arabidopsis, but grew better than sos2 under salinity stress. As expected, sos2cipk8 exhibited a more severe salt damage phenotype relative to cipk8 or sos2. Overexpression of CIPK8 in both cipk8 and sos2cipk8 attenuated the salt sensitivity phenotype. These results suggest that CIPK8-mediated activation of SOS1 is CBL10-dependent and SOS3-independent, indicating that CIPK8 and SOS2 activity in shoots is sufficient for regulating Arabidopsis salt tolerance., (© The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Root-derived abscisic acid (ABA) is known to regulate shoot physiology, such as stomata closure. Conversely, the basipetal regulatory effect of shoot-derived ABA is poorly understood. Herein, we report that simulation of shoot-ABA accumulation by exogenous application of ABA to shoots basipetally stimulates primary root (PR) growth. ABA applied to shoots accelerates root cell division, as evidenced by the increase in meristem size and cell number and the intensity of CYCB1;1::GFP (a mitosis marker). Root ABA content was not changed following shoot ABA application, although the ABA reporter line RAB18::GFP showed an increase in ABA in the cotyledons. Shoot-ABA application increases basipetal auxin transport by 114 %. Shoot-ABA-promoted PR growth can be abolished by attenuating basipetal auxin flux using 2,3,5-triiodobenzoic acid (TIBA, an auxin transport inhibitor), demonstrating that ABA promotes PR growth by increasing basipetal auxin transport. Root cell elongation, evaluated by the total length of the first 7 cells in the elongation zone (EZ), was increased by 56 % following shoot-ABA application. The cell walls of the root EZ were alkalinized by ABA, as exhibited by 8-hydroxypyrene-1,3,6-trisulfonic acid trisodium salt staining. Higher pH promotes both PR growth and cell elongation. Thus, shoot-ABA promotes cell elongation by alkalinizing the cell wall. In light of our results, we provide a representative detailed model of the basipetal regulatory effect of ABA on PR growth., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Small nuclear RNAs (snRNAs) play essential roles in spliceosome assembly and splicing. Most snRNAs are transcribed by the DNA-dependent RNA polymerase II (Pol II) and require 3'-end endonucleolytic cleavage. We have previously shown that the Arabidopsis ( Arabidopsis thaliana ) Defective in snRNA Processing 1 (DSP1) complex, composed of at least five subunits, is responsible for snRNA 3' maturation and is essential for plant development. Yet it remains unclear how DSP1 complex subunits act together to process snRNAs. Here, we show that DSP4, a member of the metallo-β-lactamase family, physically interacts with DSP1 through its β-Casp domain. Null dsp4-1 mutants have pleiotropic developmental defects, including impaired pollen development and reduced pre-snRNA transcription and 3' maturation, resembling the phenotype of the dsp1-1 mutant. Interestingly, dsp1-1 dsp4-1 double mutants exhibit complete male sterility and reduced pre-snRNA transcription and 3'-end maturation, unlike dsp1-1 or dsp4-1 In addition, Pol II occupancy at snRNA loci is lower in dsp1-1 dsp4-1 than in either single mutant. We also detected miscleaved pre-snRNAs in dsp1-1 dsp4-1, but not in dsp1-1 or dsp4-1 Taken together, these data reveal that DSP1 and DSP4 function is essential for pollen development, and that the two cooperatively promote pre-snRNA transcription and 3'-end processing efficiency and accuracy., (© 2019 The author(s). All Rights Reserved.)

Background: Na + extrusion from cells is important for plant growth in high saline environments. SOS1 (salt overly sensitive 1), an Na + /H + antiporter located in the plasma membrane (PM), functions in toxic Na + extrusion from cells using energy from an electrochemical proton gradient produced by a PM-localized H + -ATPase (AHA). Therefore, SOS1 and AHA are involved in plant adaption to salt stress., Results: In this study, the genes encoding SOS1 and AHA from the halophyte Sesuvium portulacastrum (SpSOS1 and SpAHA1, respectively) were introduced together or singly into Arabidopsis plants. The results indicated that either SpSOS1 or SpAHA1 conferred salt tolerance to transgenic plants and, as expected, Arabidopsis plants expressing both SpSOS1 and SpAHA1 grew better under salt stress than plants expressing only SpSOS1 or SpAHA1. In response to NaCl treatment, Na + and H + in the roots of plants transformed with SpSOS1 or SpAHA1 effluxed faster than wild-type (WT) plant roots. Furthermore, roots co-expressing SpSOS1 and SpAHA1 had higher Na + and H + efflux rates than single SpSOS1/SpAHA1-expressing transgenic plants, resulting in the former amassing less Na + than the latter. As seen from comparative analyses of plants exposed to salinity stress, the malondialdehyde (MDA) content was lowest in the co-transgenic SpSOS1 and SpAHA1 plants, but the K + level was the highest., Conclusion: These results suggest SpSOS1 and SpAHA1 coordinate to alleviate salt toxicity by increasing the efficiency of Na + extrusion to maintain K + homeostasis and protect the PM from oxidative damage induced by salt stress.

The ubiquitin 26S proteasome (26SP) system efficiently degrades many key regulators of plant development. 26SP consists of two subcomplexes: the catalytic 20S core particle (CP) and the 19S regulatory particle (RP). Previous studies have focused on 19S RP; whether there is a specific subunit in 20S CP that has a stress-related biological function in plants is unclear. PBE1, one of the β5 subunits of Arabidopsis proteasome CP, is essential for the assembly and proteolytic activity of 26SP in salt-stressed seedlings. The expression of PBE1 is stress-induced. During the transition from seed germination to autotrophic growth in salt-stressed seedlings, loss of PBE1 function results specifically in arrest in developmental transition but not in germination and post-germination growth. PBE1 is also important for other types of proteasome stress and Endoplasmic Reticulum (ER) stress. PBE1 modulates the protein level of the transcription factor ABI5 and thereby down-regulates the expression of several genes downstream of this key regulator which are known to be essential for plant growth under stress. Collectively, our results showed PBE1-mediated intact proteasome assembly that is essential for successful autotrophic growth, and revealed how PBE1 mediated stress proteasome functions to control both proteasome activity and abscisic acid (ABA)-mediated stress signaling in plants., (© 2018 The Authors. New Phytologist © 2018 New Phytologist Trust.)

Geminiviruses, such as beet severe curly top virus (BSCTV), are a group of DNA viruses that cause severe plant diseases and agricultural losses. The C4 protein is a major symptom determinant in several geminiviruses; however, its regulatory mechanism and molecular function in plant cells remain unclear. Here, we show that BSCTV C4 is S-acylated in planta, and that this post-translational lipid modification is necessary for its membrane localization and functions, especially its regulation of shoot development of host plants. Furthermore, the S-acylated form of C4 interacts with CLAVATA 1 (CLV1), an important receptor kinase in meristem maintenance, and consequentially affects the expression of WUSCHEL, a major target of CLV1. The abnormal development of siliques in Arabidopsis thaliana infected with BSCTV is also dependent on the S-acylation of C4, implying a potential role of CLAVATA signaling in this process. Collectively, our results show that S-acylation is essential for BSCTV C4 function, including the regulation of the CLAVATA pathway, during geminivirus infection.