ABA Signaling Research Articles

Overexpression of GiLEA5-2.1, a late embryogenesis abundant gene LEA3 from Glycyrrhiza inflata Bat., enhances the drought and salt stress tolerance of transgenic tobacco (Nicotiana benthamiana)

Abiotic stress, including drought and salt, reduces the yield of all major crop plants globally. Exploring the stress tolerance factors and mechanisms of dominant species is still an urgent task that is crucial for food and feed production. Glycyrrhiza inflata Bat., which is a halophytic plant, is economically important due to its medicinal, feeding, and ecological value. The plant is widely distributed in the salinized land of Northwest China. The plant demonstrates a robust capacity to withstand drought and salt stress, rendering it an excellent candidate for investigating the molecular mechanism behind drought and salt stress tolerance. Late embryogenesis abundant (LEA) proteins have been ascribed as significant stress tolerance factors attributed to their crucial role in improving the resilience against drought and salt stress. However, research on the salt tolerance mechanism of G. inflata is rare and limited to the physiological level. Furthermore, the role of the genes in G. inflata during abiotic stress conditions remains unclear. In this research, comprehensive knowledge of how G. inflata responds to salt stress was attained by investigating RNA-seq transcriptomes from the root of seedlings treated with 200 mM NaCl for three different time points (0 h, 3 h, and 48 h). Compared to the control, the roots exhibited a significant up-regulation in MDA and proline content as well as SOD and POD activity. The RNA-seq analysis revealed that there were 1938 DEGs that were common across various time points. GO and KEGG indicated that during salt stress, DEGs were found to be abundant in starch and sucrose metabolism, phenylpropanoid biosynthesis, glycolysis/gluconeogenesis, flavonoid biosynthesis, and the ABA signaling pathway. The correlation analysis suggested that Glyur000227s00015126.1 (GiLEA5–2.1) might be a candidate gene modulating the protection of G. inflata roots from salt stress. Apparently, the expression of GiLEA5–2.1 was triggered by drought and salt treatments. To investigate the role of GiLEA5–2.1 in drought and salt stress, plants overexpressed-GiLEA5–2.1 were generated. Overexpression of GiLEA5–2.1 in tobacco (N. benthamiana) could improve resistance against drought and salinity. Under drought and salty conditions, plants that overexpressed GiLEA5–2.1 exhibited enhanced activity of antioxidant enzymes and higher proline levels, along with decreased MDA content. Functional analyses demonstrate that GiLEA5–2.1 has a positive role in drought and salt treatments, it significantly increased the ability of tobacco to withstand drought and salt stress.

Comparative proteomic analysis of seed germination between allotetraploid cotton Gossypium hirsutum and Gossypium barbadense

Seed germination, a key initial event in the plant life cycle, directly affects cotton yield and quality. Gossypium barbadense and Gossypium hirsutum gradually evolved through polyploidization, resulting in different characteristics, and this interspecific variation lacks genetic and molecular explanation. This work aimed to compare the proteomes between G. barbadense and G. hirsutum during seed germination. Here, we identified 2740 proteins for G. barbadense and 3758 for G. hirsutum. In the initial state, proteins in two cotton involved similar bioprocess, such as sugar metabolism, DNA repairing, and ABA signaling pathway. However, in the post-germination stage, G. hirsutum expressed more protein related to redox homeostasis, peroxidase activity, and pathogen interactions. Analyzing the different expression patterns of 915 single-copy orthogroups between the two kinds of cotton indicated that most of the differentially expressed proteins in G. barbadense were related to carbon metabolism. In contrast, most proteins in G. hirsutum were associated with stress response. Besides that, by proteogenomic analysis, we found 349 putative non-canonical peptides, which may be involved in plant development. These results will help to understand the different characteristics of these two kinds of cotton, such as fiber quality, yield, and adaptability. Significance statementCotton is the predominant natural fiber crop worldwide; Gossypium barbadense and Gossypium hirsutum have evolved through polyploidization to produce differing traits. However, given their specific features, the divergence of mechanisms underlying seed germination between G. hirsutum and G. barbadense has not been discussed. Here, we explore what protein contributes to interspecific differences between G. barbadense and G. hirsutum during the seed germination period. This study helps to elucidate the evolution and domestication history of cotton polyploids and may allow breeders to understand their domestication history better and improve fiber quality and adaptability.

Transcriptome Analysis Reveals the Vital Role of ABA Plays in Drought Tolerance of the ABA-Insensitive Alfalfa (Medicago sativa L.)

Drought stress severely affects alfalfa (Medicago sativa L.) growth and production. It is particularly important to analyze the key networks of drought in alfalfa through physiological and molecular levels. However, how to quickly screen drought-tolerant alfalfa germplasm and how to elucidate the molecular pathways of alfalfa responding to drought are less studied. In this study, based on our previous research, we further verified the association between the heritability of ABA sensitivity during seed germination and drought tolerance of plants and identified the key pathways of drought tolerance differences between ABA-sensitivity (S1-0) and -insensitivity (S1-50) plants via RNA-seq and analysis. The results showed that the sensitivity to ABA in alfalfa seeds can be inherited and that plants that are insensitive to ABA during germination show stronger drought tolerance. An analysis of the differentially expressed genes (DEGs) revealed that ABA biosynthesis and signaling, amino acid metabolism, LEA, and wax synthesis-related pathways may be the key pathways that can be used for drought tolerance improvement in alfalfa. DEGs such as NCED, PYR/PYL, and PP2C may contribute to drought tolerance in the S1-50 plant. The study further confirms that screening with ABA at the seed germination stage can select alfalfa lines with good drought tolerance, which provides a new theoretical basis for alfalfa drought tolerance breeding. The expression of the key genes of alfalfa in response to drought stress was also tested.

Potential Response Patterns of Endogenous Hormones in Cliff Species Opisthopappus taihangensis and Opisthopappus longilobus under Salt Stress.

When plants are exposed to salt stress, endogenous hormones are essential for their responses through biosynthesis and signal transduction pathways. However, the roles of endogenous hormones in two cliff species (Opisthopappus taihangensis and Opisthopappus longilobus (Opisthopappus genus)) in the Taihang Mountains under salt stress have not been investigated to date. Following different time treatments under 500 mM salt concentrations, 239 differentially expressed gene (DEG)-related endogenous hormones were identified that exhibited four change trends, which in Profile 47 were upregulated in both species. The C-DEG genes of AUX, GA, JA, BR, ETH, and ABA endogenous hormones were significantly enriched in Opisthopappus taihangensis (O. taihangensis) and Opisthopappus longilobus (O. longilobus). During the responsive process, mainly AUX, GA, and JA biosynthesis and signal transduction were triggered in the two species. Subsequently, crosstalk further influenced BR, EHT, ABA, and MAPK signal transduction pathways to improve the salt resistance of the two species. Within the protein-protein interactions (PPI), seven proteins exhibited the highest interactions, which primarily involved two downregulated genes (SAUR and GA3ox) and eight upregulated genes (ACX, MFP2, JAZ, BRI1, BAK1, ETR, EIN2, and SNRK2) of the above pathways. The more upregulated expression of ZEP (in the ABA biosynthesis pathway), DELLA (in the GA signaling pathway), ABF (in the ABA signaling pathway), and ERF1 (in the ETH signaling pathway) in O. taihangensis revealed that it had a relatively higher salt resistance than O. longilobus. This revealed that the responsive patterns to salt stress between the two species had both similarities and differences. The results of this investigation shed light on the potential adaptive mechanisms of O. taihangensis and O. longilobus under cliff environments, while laying a foundation for the study of other cliff species in the Taihang Mountains.

Integrated physiological, metabolomic, and transcriptomic analyses elucidate the regulation mechanisms of lignin synthesis under osmotic stress in alfalfa leaf (Medicago sativa L.)

Alfalfa, an essential forage crop known for its high yield, nutritional value, and strong adaptability, has been widely cultivated worldwide. The yield and quality of alfalfa are frequently jeopardized due to environmental degradation. Lignin, a constituent of the cell wall, enhances plant resistance to abiotic stress, which often causes osmotic stress in plant cells. However, how lignin responds to osmotic stress in leaves remains unclear. This study explored the effects of osmotic stress on lignin accumulation and the contents of intermediate metabolites involved in lignin synthesis in alfalfa leaves. Osmotic stress caused an increase in lignin accumulation and the alteration of core enzyme activities and gene expression in the phenylpropanoid pathway. We identified five hub genes (CSE, CCR, CADa, CADb, and POD) and thirty edge genes (including WRKYs, MYBs, and UBPs) by integrating transcriptome and metabolome analyses. In addition, ABA and ethylene signaling induced by osmotic stress regulated lignin biosynthesis in a contradictory way. These findings contribute to a new theoretical foundation for the breeding of high-quality and resistant alfalfa varieties.

Transcriptome Analysis Reveals the Molecular Mechanisms of BR Negative Regulatory Factor StBIN2 Maintaining Tuber Dormancy.

Potato is an important food crop. After harvest, these tubers will undergo a period of dormancy. Brassinosteroids (BRs) are a new class of plant hormones that regulate plant growth and seed germination. In this study, 500 nM of BR was able to break the dormancy of tubers. Additionally, exogenous BR also upregulated BR signal transduction genes, except for StBIN2. StBIN2 is a negative regulator of BR, but its specific role in tuber dormancy remains unclear. Transgenic methods were used to regulate the expression level of StBIN2 in tubers. It was demonstrated that the overexpression of StBIN2 significantly prolonged tuber dormancy while silencing StBIN2 led to premature sprouting. To further investigate the effect of StBIN2 on tuber dormancy, RNA-Seq was used to analyze the differentially expressed genes in OE-StBIN2, RNAi-StBIN2, and WT tubers. The results showed that StBIN2 upregulated the expression of ABA signal transduction genes but inhibited the expression of lignin synthesis key genes. Meanwhile, it was also found that StBIN2 physically interacted with StSnRK2.2 and StCCJ9. These results indicate that StBIN2 maintains tuber dormancy by mediating ABA signal transduction and lignin synthesis. The findings of this study will help us better understand the molecular mechanisms underlying potato tuber dormancy and provide theoretical support for the development of new varieties using related genes.

Transcriptome analysis reveals ABA involved in the detoxification mechanism of macroalga Gracilariopsis lemaneiformis to cadmium toxicity

IntroductionCadmium (Cd) is a significant threat environmental pollutant in the marine ecological environment offshore. The macroalga, Gracilariopsis lemaneiformis, of significant economic value, is widely cultivated along China’s coastline. Yet, little is known about the molecular mechanisms underlying Cd tolerance in macroalga.MethodsHere, we examined the transcriptome of G. lemaneiformis exposed to Cd to identify the responses to Cd stress.Results and discussionOur findings revealed that Cd led to the retardation of growth rate in G. lemaneiformis, accompanied by a notable reduction in the content of photosynthetic pigments and a decrease in the expression of genes associated with the photosynthetic system and nitrogen metabolism. When exposed to Cd, there was a rapid increase in Cd levels through the upregulation of genes encoding GlZIP6 and GlIRT1. Additionally, the expression of Cd efflux transporters, GlZIP1 and GlABCG22, and the ABCC7 transporter for compartmentation to the vacuole, was induced to mitigate Cd toxicity. Cd also activated crucial genes involved in the ABA biosynthesis and enhanced ABA content, thereby inducing ABA signaling pathway. Furthermore, exogenous ABA reduced the growth inhibition of G. lemaneiformis under Cd stress. Redox homeostasis was adjusted to adapt to Cd toxicity, with thioredoxin, glutaredoxin cycle and ascorbate-glutathione cycle identified as playing significant in maintaining reactive oxygen species homeostasis. Moreover, transcription factors such as several MYBs, signal transmission factors G protein and heat shock proteins (sHSPs, HSP 40, HSP 90, HSP101) were involved in the detoxification of Cd. Collectively, this study provided a comprehensive understanding of the molecular mechanisms underpinning the of responses of macroalga G. lemaneiformis to Cd exposure.

Endomembrane-biased dimerization of ABCG16 and ABCG25 transporters determines their substrate selectivity in ABA-regulated plant growth and stress responses

ATP-binding cassette (ABC) transporters are integral membrane proteins that have evolved diverse functions fulfilled via the transport of various substrates. In Arabidopsis, the G subfamily of ABC proteins is particularly abundant and participates in multiple signaling pathways during plant development and stress responses. In this study, we revealed that two Arabidopsis ABCG transporters, ABCG16 and ABCG25, engage in ABA-mediated stress responses and early plant growth through endomembrane-specific dimerization-coupled transport of ABA and ABA-glucosyl ester (ABA-GE), respectively. We first revealed that ABCG16 contributes to osmotic stress tolerance via ABA signaling. More specifically, ABCG16 induces cellular ABA efflux in both yeast and plant cells. Using FRET analysis, we showed that ABCG16 forms obligatory homodimers for ABA export activity and that the plasma membrane-resident ABCG16 homodimers specifically respond to ABA, undergoing notable conformational changes. Furthermore, we demonstrated that ABCG16 heterodimerizes with ABCG25 at the endoplasmic reticulum (ER) membrane and facilitates the ER entry of ABA-GE in both Arabidopsis and tobacco cells. The specific responsiveness of the ABCG16-ABCG25 heterodimer to ABA-GE and the superior growth of their double mutant support an inhibitory role of these two ABCGs in early seedling establishment via regulation of ABA-GE translocation across the ER membrane. Our endomembrane-specific analysis of the FRET signals derived from the homo- or heterodimerized ABCG complexes allowed us to link endomembrane-biased dimerization to the translocation of distinct substrates by ABCG transporters, providing a prototypic framework for understanding the omnipotence of ABCG transporters in plant development and stress responses.

Downregulation of GhALKBH10B improves drought tolerance through increasing the stability of photosynthesis related- and ABA signaling pathway genes in cotton

N6-methyladenosine (m6A) is a post-transcriptional modification found in eukaryotic mRNAs. It is installed and removed by methyltransferases and demethylases, respectively. Recent studies have demonstrated that alpha-ketoglutarate-dependent dioxygenase homolog 10B (ALKBH10B) is an mRNA m6A eraser affecting floral transition and responding to abiotic stresses in Arabidopsis thaliana. However, the roles of m6A eraser proteins, especially cotton GhALKBH10B, in plant adaptation to abiotic stresses are largely unknown. In this study, we aimed to determine the role of GhALKBH10B in the response of cotton to drought stress. The m6A level significantly increased in GhALKBH10B-silenced plants compared to that in the control plants. Seedling growth and survival rate of GhALKBH10B-silenced plants were enhanced under drought stress, while heterologous expression of GhALKBH10B in Arabidopsis thaliana attenuates the drought tolerance. Notably, drought-tolerant phenotypes of GhALKBH10B-silenced plants were correlated with the increased mRNA stability of photosynthesis-related genes and GhSnRK2;3 which is a key regulator in ABA-dependent pathway under both normal and dehydration conditions. In addition, GhALKBH10B interacts with a RNA binding protein GhSERBP1(Hyaluronic / mRNA binding family HABP4_PAI-RBP1) both in vitro and in vivo and silencing GhSERBP1 decreased the drought tolerance in cotton. Collectively, these findings indicate that GhALKBH10B-mediated m6A demethylation affects the transcript levels of photosynthesis-related genes and GhSnRK2;3, which are important for survival of cotton in response to drought stress. This will provide important clues for revealing the novel molecular mechanism at the epi-transcriptomic level in drought response and provide key candidate genes for cotton drought-tolerant breeding in the future.

Basal ABA signaling balances transpiration and photosynthesis.

The balance between the CO2 entry for photosynthesis and transpiration water loss is crucial for plant growth, and ABA signaling can affect this equilibrium. To test how ABA balances plant growth and environmental adaptation, we performed molecular genetics studies in the biotech crop Nicotiana benthamiana under well-watered or drought conditions. Studies on ABA signaling in crops are complicated by the multigenic nature of the PYR/PYL/RCAR ABA receptor family and its functional redundancy, which is particularly challenging in polyploid plants. We have generated a pentuple pyl mutant in the allotetraploid Nicotiana benthamiana through CRISPR/Cas9 gene editing. The pentuple mutant is impaired in 2 NbPYL1-like and 3 NbPYL8-like receptors, affecting the regulation of transpiration and several ABA-dependent transcriptional processes. RNA-seq and metabolite analysis revealed that the synthesis of galactinol, an essential precursor for the osmoprotective raffinose family of oligosaccharides, is ABA-dependent and impaired in the mutant under osmotic stress. In contrast, our results show that, under well-watered conditions, partial inactivation of ABA signaling leads to higher CO2 entry and photosynthesis in the mutant than in WT. Photosynthesis analyses revealed an increased CO2 diffusion capacity mediated by higher stomatal and mesophyll conductances, and higher substomatal CO2 concentration in the pentuple mutant. RNA-seq analyses revealed that genes associated with cell wall loosening (e.g., expansins) and porosity were strongly downregulated by ABA in WT. In summary, a partial relief of the ABA control on transpiration mediated by ABA receptors positively affects photosynthesis when water is not limited, at the expense of reduced water use efficiency.

MsABCG1, ATP-Binding Cassette G transporter from Medicago Sativa, improves drought tolerance in transgenic Nicotiana Tabacum.

Drought has a devastating impact, presenting a formidable challenge to agricultural productivity and global food security. Among the numerous ABC transporter proteins found in plants, the ABCG transporters play a crucial role in plant responses to abiotic stress. In Medicago sativa, the function of ABCG transporters remains elusive. Here, we report that MsABCG1, a WBC-type transporter highly conserved in legumes, is critical for the response to drought in alfalfa. MsABCG1 is localized on the plasma membrane, with the highest expression observed in roots under normal conditions, and its expression is induced by drought, NaCl and ABA signalling. In transgenic tobacco, overexpression of MsABCG1 enhanced drought tolerance, evidenced by increased osmotic regulatory substances and reduced lipid peroxidation. Additionally, drought stress resulted in reduced ABA accumulation in tobacco overexpressing MsABCG1, demonstrating that overexpression of MsABCG1 enhanced drought tolerance was not via an ABA-dependent pathway. Furthermore, transgenic tobacco exhibited increased stomatal density and reduced stomatal aperture under drought stress, indicating that MsABCG1 has the potential to participate in stomatal regulation during drought stress. In summary, these findings suggest that MsABCG1 significantly enhances drought tolerance in plants and provides a foundation for developing efficient drought-resistance strategies in crops.

Apple E3 ligase MdPUB23 mediates ubiquitin-dependent degradation of MdABI5 to delay ABA-triggered leaf senescence.

ABSCISIC ACID-INSENSITIVE5 (ABI5) is a core regulatory factor that mediates the ABA signaling response and leaf senescence. However, the molecular mechanism underlying the synergistic regulation of leaf senescence by ABI5 with interacting partners and the homeostasis of ABI5 in the ABA signaling response remain to be further investigated. In this study, we found that the accelerated effect of MdABI5 on leaf senescence is partly dependent on MdbHLH93, an activator of leaf senescence in apple. MdABI5 directly interacted with MdbHLH93 and improved the transcriptional activation of the senescence-associated gene MdSAG18 by MdbHLH93. MdPUB23, a U-box E3 ubiquitin ligase, physically interacted with MdABI5 and delayed ABA-triggered leaf senescence. Genetic and biochemical analyses suggest that MdPUB23 inhibited MdABI5-promoted leaf premature senescence by targeting MdABI5 for ubiquitin-dependent degradation. In conclusion, our results verify that MdABI5 accelerates leaf senescence through the MdABI5-MdbHLH93-MdSAG18 regulatory module, and MdPUB23 is responsible for the dynamic regulation of ABA-triggered leaf senescence by modulating the homeostasis of MdABI5.

CaERF1- mediated ABA signal positively regulates camptothecin biosynthesis by activating the iridoid pathway in Camptotheca acuminata

Camptotheca acuminata is one of the primary sources of camptothecin (CPT), which is widely used in the treatment of human malignancies because of its inhibitory activity against DNA topoisomerase I. Although several transcription factors have been identified for regulating CPT biosynthesis in other species, such as Ophiorrhiza pumila, the specific regulatory components controlling CPT biosynthesis in C. acuminata have yet to be definitively determined. In this study, CaERF1, an DREB subfamily of the APETALA2/ethylene response factors (AP2ERFs), was identified in C. acuminata. The transient overexpression and silencing of CaERF1 in C. acuminata leaves confirmed that it positively regulates the accumulation of CPT by inducing the expression of CaCYC1 and CaG8O in the iridoid pathway. Results of transient transcriptional activity assay and yeast one-hybrid assays have showed that CaERF1 transcriptionally activates the expression of CaCYC1 and CaG8O by binding to RAA and CEI elements in the promoter regions of these two genes. Furthermore, the expression of CaCYC1 and CaG8O in CaERF1-silenced leaves was less sensitive to ABA treatment, indicating that CaERF1 is a crucial component involved in ABA-regulated CPT biosynthesis in C. acuminata.

GhAGL16 (AGAMOUS-LIKE16) Negatively Regulates Tolerance to Water Deficit in Transgenic Arabidopsis and Cotton.

Cotton is one of the most economically important crops in the world, and drought is a key abiotic factor that can significantly reduce cotton yield. MADS-box transcription factors play essential roles in various aspects of plant growth and development as well as responses to biotic and abiotic stress. However, the use of MADS-box transcription factors to regulate water stress responses has not been fully explored in cotton. Here, we showed that GhAGL16 acts as a negative regulator of water deficit in cotton, at least in part by regulating ABA signaling. GhAGL16-overexpressing (GhAGL16-OE) transgenic Arabidopsis had lower survival rates and relative water contents (RWCs) under water stress. Isolated leaves of GhAGL16-OE Arabidopsis had increased water loss rates, likely attributable to their increased stomatal density. GhAGL16-OE Arabidopsis also showed reduced primary root lengths in response to mannitol treatment and decreased sensitivity of seed germination to ABA treatment. By contrast, silencing GhAGL16 in cotton enhanced tolerance to water deficit by increasing proline (Pro) content, increasing superoxide dismutase (SOD) and peroxidase (POD) activities, and reducing malondialdehyde (MDA) and hydrogen peroxide (H2O2) contents under water stress. Subcellular localization and transcriptional activation assays confirmed that GhAGL16 is a nuclear protein that lacks transcriptional self-activation activity. The expression of ABA biosynthesis-related genes (GhNCED3/7/14), a catabolism-related gene (GhCYP707A), and a gene related to the ABA signaling pathway (GhABF4) was altered in GhAGL16-silenced plants. Taken together, our data demonstrate that GhAGL16 plays an important role in cotton resistance to water stress.

A Multi-Omics Analysis Revealed the Diversity of the MYB Transcription Factor Family's Evolution and Drought Resistance Pathways.

The MYB transcription factor family can regulate biological processes such as ABA signal transduction to cope with drought stress, but its evolutionary mechanism and the diverse pathways of response to drought stress in different species are rarely reported. In this study, a total of 4791 MYB family members were identified in 908,757 amino acid sequences from 12 model plants or crops using bioinformatics methods. It was observed that the number of MYB family members had a linear relationship with the chromosome ploidy of species. A phylogenetic analysis showed that the MYB family members evolved in subfamily clusters. In response to drought stress, the pathways of MYB transcription factor families exhibited species-specific diversity, with closely related species demonstrating a higher resemblance. This study provides abundant references for drought resistance research and the breeding of wheat, soybean, and other plants.

Genome-Wide Analysis of Homologous E6-AP Carboxyl-Terminal E3 Ubiquitin Ligase Gene Family in Populus trichocarpa

Proteins containing the homologous E6-AP carboxyl-terminal (HECT) domain are a class of E3 ubiquitin ligases involved in the ubiquitin–proteasome pathway, which plays an irreplaceable role in plant growth, development, and stress resistance. However, a phylogenetic analysis and expression profile of the HECT gene (PtrHECT) in the model plant Populus trichocarpa (Torr. & Gray) have not been reported. In this study, we identified 13 PtrHECT genes using genome-wide analysis, and then these were divided into four groups. The protein interaction networks showed that the PtrHECT protein may interact with PTR6 and participate in ABA signal regulation. Abiotic stress is the main environmental factor limiting plant growth and development. The qRT-PCR results showed that PtrHECT1, 4, 7, 8, and 9 were significantly up-regulated in leaves at each time point under drought stress, and most PtrHECT genes responded to both drought and high salt stress, consistent with their promoter sequence analysis, revealing the presence of an important number of phytohormone-responsive and stress-related cis-regulatory elements. This study provides useful information for further analysis of the functions of the HECT gene family in P. trichocarpa.

The zinc finger protein 3 of Arabidopsis thaliana regulates vegetative growth and root hair development.

Zinc finger protein 3 (ZFP3) and closely related C2H2 zinc finger proteins have been identified as regulators of abscisic acid signals and photomorphogenic responses during germination. Whether ZFP3 and related ZFP factors regulate plant development is, however, not known. ZFP3 overexpression reduced plant growth, limited cell expansion in leaves, and compromised root hair development. The T-DNA insertion zfp3 mutant and transgenic lines with silenced ZFP1, ZFP3, ZFP4, and ZFP7 genes were similar to wild-type plants or had only minor differences in plant growth and morphology, probably due to functional redundancy. RNAseq transcript profiling identified ZFP3-controlled gene sets, including targets of ABA signaling with reduced transcript abundance. The largest gene set that was downregulated by ZFP3 encoded regulatory and structural proteins in cell wall biogenesis, cell differentiation, and root hair formation. Chromatin immunoprecipitation confirmed ZFP3 binding to several target promoters. Our results suggest that ZFP3 and related ZnF proteins can modulate cellular differentiation and plant vegetative development by regulating the expression of genes implicated in cell wall biogenesis.

MKK3 Cascade Regulates Seed Dormancy Through a Negative Feedback Loop Modulating ABA Signal in Rice

BackgroundWith the increasing frequency of climatic anomalies, high temperatures and long-term rain often occur during the rice-harvesting period, especially for early rice crops in tropical and subtropical regions. Seed dormancy directly affects the resistance to pre-harvest sprouting (PHS). Therefore, in order to increase rice production, it is critical to enhance seed dormancy and avoid yield losses to PHS. The elucidation and utilization of the seed dormancy regulation mechanism is of great significance to rice production. Preliminary results indicated that the OsMKKK62-OsMKK3-OsMPK7/14 module might regulate ABA sensitivity and then control seed dormancy. The detailed mechanism is still unclear.ResultsThe overexpression of OsMKK3 resulted in serious PHS. The expression levels of OsMKK3 and OsMPK7 were upregulated by ABA and GA at germination stage. OsMKK3 and OsMPK7 are both located in the nucleus and cytoplasm. The dormancy level of double knockout mutant mkk3/mft2 was lower than that of mkk3, indicating that OsMFT2 functions in the downstream of MKK3 cascade in regulating rice seeds germination. Biochemical results showed that OsMPK7 interacted with multiple core ABA signaling components according to yeast two-hybrid screening and luciferase complementation experiments, suggesting that MKK3 cascade regulates ABA signaling by modulating the core ABA signaling components. Moreover, the ABA response and ABA responsive genes of mpk7/14 were significantly higher than those of wild-type ZH11 when subjected to ABA treatment.ConclusionMKK3 cascade mediates the negative feedback loop of ABA signal through the interaction between OsMPK7 and core ABA signaling components in rice.

Transcriptome analysis and identification of the low potassium stress-responsive gene SiSnRK2.6 in foxtail millet (Setaria italica L.).

The transcriptome is beneficial for dissecting the mechanism of millet in response to low potassium stress and SiSnRK2.6 was identified as a potential target for improving low potassium stress tolerance. Foxtail millet (Setaria italica L.), which originated in China, has high nutrient utilization character. Nevertheless, the molecular mechanism of its tolerance to low potassium stress is largely unclear. In this research, the low potassium tolerant variety "Yugu28" was screened out by low potassium stress treatment, and the transcriptome of "Yugu28" under low potassium stress was comprehensively analyzed. A total of 4254 differentially expressed genes (DEGs) were identified, including 1618 up-regulated and 2636 down-regulated genes, respectively. In addition, there were 302 transcription factor (TF) genes in the DEGs and MYB TFs accounted for the highest proportion, which was 14.9%. After functional analysis of all DEGs, a total of 7 genes involved in potassium transport and potassium ion channels and 50 genes corresponding to hormones were screened. The expression levels of randomly selected 17 DEGs were verified by qRT-PCR and the results coincided well with the RNA-seq analysis, indicating the reliability of our transcriptome data. Moreover, one of the ABA signaling pathway genes, SiSnRK2.6, was identified and selected for further functional verification. Compared with the wild type, transgenic rice with ecotopic expression of SiSnRK2.6 showed remarkably increased root length and root number, indicating that overexpression of SiSnRK2.6 can enhance the resistance of transgenic plants to low potassium stress.

Pepper SBP-box transcription factor, CaSBP13, plays a negatively role in drought response.

The SBP-box gene significantly influences plant growth, development, and stress responses, yet its function in pepper plants during drought stress remains unexplored. Using virus-induced gene silencing and overexpression strategies, we examined the role of CaSBP13 during drought stress in plants. The results revealed that the expression of CaSBP13 can be induced by drought stress. Silencing of CaSBP13 in pepper notably boosted drought resistance, as evident by decreased active oxygen levels. Furthermore, the water loss rate, relative electrical conductivity, malondialdehyde content, and stomatal density were reduced in CaSBP13-silenced plants compared to controls. In contrast, CaSBP13 overexpression in Nicotiana benthamiana decreased drought tolerance with elevated reactive oxygen levels and stomatal density. Additionally, ABA signaling pathway genes (CaPP2C, CaAREB) exhibited reduced expression levels in CaSBP13-silenced plants post drought stress, as compared to control plants. On the contrary, CaPYL9 and CaSNRK2.4 showed heightened expression in CaSBP13-sienced plants under the same conditions. However, a converse trend for NbAREB, NbSNRK2.4, and NbPYL9 was observed post-four day drought in CaSBP13-overexpression plants. These findings suggest that CaSBP13 negatively regulates drought tolerance in pepper, potentially via ROS and ABA signaling pathways.