Wild-type Plants Research Articles

GH3 Gene Family Identification in Chinese White Pear (Pyrus bretschneideri) and the Functional Analysis of PbrGH3.5 in Fe Deficiency Responses in Tomato

Iron (Fe) deficiency poses a major threat to pear (Pyrus spp.) fruit yield and quality. The Gretchen Hagen 3 (GH3) plays a vital part in plant stress responses. However, the GH3 gene family is yet to be characterized, and little focus has been given to the function of the GH3 gene in Fe deficiency responses. Here, we identified 15 GH3 proteins from the proteome of Chinese white pear (Pyrus bretschneideri) and analyzed their features using bioinformatics approaches. Structure domain and motif analyses showed that these PbrGH3s were relatively conserved, and phylogenetic investigation displayed that they were clustered into two groups (GH3 I and GH3 II). Meanwhile, cis-acting regulatory element searches of the corresponding promoters revealed that these PbrGH3s might be involved in ABA- and drought-mediated responses. Moreover, the analysis of gene expression patterns exhibited that most of the PbrGH3s were highly expressed in the calyxes, ovaries, and stems of pear plants, and some genes were significantly differentially expressed in normal and Fe-deficient pear leaves, especially for PbrGH3.5. Subsequently, the sequence of PbrGH3.5 was isolated from the pear, and the transgenic tomato plants with PbrGH3.5 overexpression (OE) were generated to investigate its role in Fe deficiency responses. It was found that the OE plants were more sensitive to Fe deficiency stress. Compared with wild-type (WT) plants, the rhizosphere acidification and ferric reductase activities were markedly weakened, and the capacity to scavenge reactive oxygen species was prominently impaired in OE plants under Fe starvation conditions. Moreover, the expressions of Fe-acquisition-associated genes, such as SlAHA4, SlFRO1, SlIRT1, and SlFER, were all greatly repressed in OE leaves under Fe depravation stress, and the free IAA level was dramatically reduced, while the conjugated IAA contents were notably escalated. Combined, our findings suggest that pear PbrGH3.5 negatively regulates Fe deficiency responses in tomato plants, and might help enrich the molecular basis of Fe deficiency responses in woody plants.

Invertase of the cell wall controls the growth of both the apical and vegetative organs of the tomato

Researchers have intensively studied how external factors like temperature, humidity, soil structure, and fertilizer affect tomato plant growth. However, little is known about how internal factors, particularly cell wall invertase, affect plant growth. This study aims to determine how the activity of the cell wall invertase enzyme regulates the apical and vegetative growth of plants. The study utilized four tomato plants, consisting of two transgenic and two wild-type plants. The transgenic plants in one set exhibit lower levels of the cell wall invertase, known as TLINA. Another set of transgenic plants, known as TINHI, exhibited higher levels of the cell wall invertase. We named both wild types of plants, WLINA and WINHI, respectively, and used them as the control plant. The study's results revealed that the transgenic plants (TLINA) ceased their apical shoot growth, leading to the growth of an axial shoot to take the place of the apical shoot. t. Also, the TLINA plants grew lower than wild-type plants. Interestingly, the vegetative organs of TINHI plants grew higher than both wild types and TLINA plants. This evidence suggested that cell wall invertase regulates the apical and vegetative organs of plants.

Analysis of the quadruple lsu mutant reveals molecular determinants of the role of LSU proteins in sulfur assimilation in Arabidopsis.

Because plants are immobile, they have developed intricate mechanisms to sense and absorb nutrients, adjusting their growth and development accordingly. Sulfur is an essential macroelement, but our understanding of its metabolism and homeostasis is limited. LSU (RESPONSE TO LOW SULFUR) proteins are plant-specific proteins with unknown molecular functions and were first identified during transcriptomic studies on sulfur deficiency in Arabidopsis. These proteins are crucial hubs that integrate environmental signals and are involved in the response to various stressors. Herein, we report the direct involvement of LSU proteins in primary sulfur metabolism. Our findings revealed that the quadruple lsu mutant, q-lsu-KO, which was grown under nonlimiting sulfate conditions, exhibited a molecular response resembling that of sulfur-deficient wild-type plants. This led us to explore the interactions of LSU proteins with sulfate reduction pathway enzymes. We found that all LSU proteins interact with ATPS1 and ATPS3 isoforms of ATP sulfurylase, all three isoforms of adenosine 5´ phosphosulfate reductase (APR), and sulfite reductase (SiR). Additionally, in vitro assays revealed that LSU1 enhances the enzymatic activity of SiR. These results highlight the supportive role of LSU proteins in the sulfate reduction pathway.

Birch (Betula platyphylla) BES/BZR transcription factor BpBZR1-6 improves salt tolerance in transgenic Arabidopsis thaliana

BackgroundSalt stress is one of the major environmental factors affecting plant growth and productivity. BRI1-EMS suppressor 1/brassinazole-resistant 1 ((BES1/BZR1) plays an important role in responding to abiotic stress in plants. Although the impacts of BES1/BZR1 on plant growth and resistance have been documented, the potential mechanisms are not fully elucidated in Betula platyphylla. This work contributes to the understanding of how BES1/BZR1 promotes stress tolerance in woody plants.ResultsSix BES1/BZR1 family members were identified from Betula platyphylla. Cis-element analysis showed that the promoters of six genes were rich in ABA-responsive element (ABRE), MYB and MBS cis-acting elements, which are reported to be involved in abiotic stress responses. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis indicated that BpBZR1-6 (BPChr10G06000) could be induced by salt stress, ABA and BRs. BpBZR1-6 was localized in the nucleus and had transactivation activity. Ectopic expression of BpBZR1-6 enhanced Arabidopsis tolerance and decreased abscisic acid (ABA) sensitivity under salt treatment. Specifically, the seed germination rate, root length, fresh weight and chlorophyll content were significantly higher in BpBZR1-6-overexpressing (OE) transgenic plants than in wild-type (WT) plants after salt stress (P < 0.05). Additionally, BpBZR1-6 overexpression showed enhanced the reactive oxygen species (ROS) scavenging capability under salt stress, including increasing the activities of antioxidant enzyme, resulting in a decrease in O2− and H2O2 accumulation, and reducing malondialdehyde (MDA) content. Meanwhile, the expression levels of six antioxidant enzyme genes were higher in OE plants than in WT plants after stress.ConclusionBpBZR1-6 overexpression enhanced the salt tolerance of transgenic plants by modulating antioxidant enzyme gene expression and ROS scavenging, which may provide underlying strategy for breeding of salt-tolerant plants.

Nitrate assimilation pathway is impacted in young tobacco plants overexpressing a constitutively active nitrate reductase or displaying a defective CLCNt2

BackgroundWe have previously shown that the expression of a constitutively active nitrate reductase variant and the suppression of CLCNt2 gene function (belonging to the chloride channel (CLC) gene family) in field-grown tobacco reduces tobacco-specific nitrosamines (TSNA) accumulation in cured leaves and cigarette smoke. In both cases, TSNA reductions resulted from a strong diminution of free nitrate in the leaf, as nitrate is a precursor of the TSNA-producing nitrosating agents formed during tobacco curing and smoking. These nitrosating agents modify tobacco alkaloids to produce TSNAs, the most problematic of which are NNN (N-nitrosonornicotine) and NNK (4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone). The expression of a deregulated nitrate reductase enzyme (DNR) that is no longer responsive to light regulation is believed to diminish free nitrate pools by immediately channeling incoming nitrate into the nitrate assimilation pathway. The reduction in nitrate observed when the two tobacco gene copies encoding the vacuolar nitrate transporter CLCNt2 were down-regulated by RNAi-mediated suppression or knocked out using the CRISPR-Cas technology was mechanistically distinct; likely attributable to the inability of the tobacco cell to efficiently sequester nitrate into the vacuole where this metabolite is protected from further assimilation. In this study, we used transcriptomic and metabolomic analyses to compare the nitrate assimilation response in tobacco plants either expressing DNR or lacking CLCNt2 function.ResultsWhen grown in a controlled environment, both DNR and CLCNt2-KO (CLCKO) plants exhibited (1) reduced nitrate content in the leaf; (2) increased N-assimilation into the amino acids Gln and Asn; and (3) a similar pattern of differential regulation of several genes controlling stress responses, including water stress, and cell wall metabolism in comparison to wild-type plants. Differences in gene regulation were also observed between DNR and CLCKO plants, including genes encoding nitrite reductase and asparagine synthetase.ConclusionsOur data suggest that even though both DNR and CLCKO plants display common characteristics with respect to nitrate assimilation, cellular responses, water stress, and cell wall remodeling, notable differences in gene regulatory patterns between the two low nitrate plants are also observed. These findings open new avenues in using plants fixing more nitrogen into amino acids for plant improvement or nutrition perspectives.

Two genes encoding a bacterial-type ABC transporter function in aluminum tolerance in soybean.

GmABCI5 and GmABCI13 enhance Al tolerance through regulating the composition of root cell wall, and in this process, GmABCI5 and GmABCI13 may act in the form of a complex. Aluminum (Al) toxicity is a major factor limiting plant growth in acidic soils. ATP-binding cassette (ABC) transporters are involved in plant tolerance to various environmental stresses. However, there are few reports on the ABC transporters implicated in soybean tolerance to Al toxicity. Here, we reported that two genes, GmABCI5 and GmABCI13, were involved in Al tolerance in soybean (Glycine max). GmABCI5 and GmABCI13 encode a nucleotide-binding domain and a transmembrane domain of a bacterial-type ABC transporter, respectively. The expression of both GmABCI5 and GmABCI13 was mainly induced by Al in the roots. GmABCI5 was localized at the plasma membrane and also in the cytoplasm and nucleus, while GmABCI13 was only localized at the plasma membrane. Furthermore, GmABCI5 could physically interact with GmABCI13. Overexpression of GmABCI5 or GmABCI13 in Arabidopsis reduced Al accumulation in roots and enhanced Al tolerance. However, expression of GmABCI5 and/or GmABCI13 in yeast cells did not affect Al uptake. Under Al stress, transgenic Arabidopsis lines expressing GmABCI5 or GmABCI13 had lower Al content in root cell walls than wild-type plants. Further analysis showed that Al content in cell wall fractions (pectin and hemicellulose 1) of transgenic lines was significantly lower than that of wild-type plants, which was coincident with the changes of pectin and hemicellulose 1 content under Al stress. These results indicate that GmABCI5 and GmABCI13 form an ABC transporter complex to regulate Al tolerance by affecting the modification of cell wall.

Overexpression of StCDPK13 in Potato Enhances Tolerance to Drought Stress

Calcium-dependent protein kinases (CDPKs), which are activated by transient changes in the Ca2+ concentration in plants, are important for various biological processes, such as growth, development, defense against biotic and abiotic stresses, and others. Mannitol is commonly used as an osmotic regulatory substance in culture medium or nutrient solutions to create water-deficit conditions. Here, we cloned the potato (Solanum tuberosum L.) StCDPK13 gene and generated stable transgenic StCDPK13-overexpression potato plants. To investigate the potential functions of StCDPK13 in response to drought stress, overexpression-transgenic (OE1, OE2, and OE7) and wild-type (WT) potato seedlings were cultured on MS solid media without or with mannitol, representing the control or drought stress, for 20 days; the elevated mannitol concentrations (150 and 200 mM) were the drought stress conditions. The StCDPK13 gene was consistently expressed in different tissues and was induced by drought stress in both OE and WT plants. The phenotypic traits and an analysis of physiological indicators revealed that the transgenic plants exhibited more tolerance to drought stress than the WT plants. The overexpression lines showed an increased plant height, number of leaves, dry shoot weight, root length, root number, root volume, number of root tips, fresh root weight, and dry root weight under drought stress. In addition, the activities of antioxidant enzymes (CAT, SOD, and POD) and the accumulation of proline and neutral sugars were significantly increased, whereas the levels of malondialdehyde (MDA) and reactive oxygen species (ROS), including hydrogen peroxide (H2O2) and O2•−, were significantly reduced in the OE lines compared to WT plants under drought stress. Moreover, the stomatal aperture of the leaves and the water loss rate in the leaves of the OE lines were significantly reduced under drought stress compared to the WT plants. In addition, the overexpression of StCDPK13 upregulated the expression levels of stress-related genes under drought stress. Collectively, these results indicate that the StCDPK13 gene plays a positive role in drought tolerance by reducing the stomatal aperture, promoting ROS scavenging, and alleviating oxidative damage under drought stress in potatoes.

CCT39-COMT1/BGLU18-2 module promotes lignin biosynthesis in poplar

Lignin is a crucial constituent of cell walls and plays a pivotal role in plant growth and development. However, the transcriptional regulatory network governing lignin biosynthesis is not fully understood. In this study, we observed that PpnCCT39 overexpression resulted in greener stems, larger basal diameters, and increased stem dry weight. Additionally, the secondary xylem of lines overexpressing PpnCCT39 was wider, had larger xylem fiber cell areas, and thicker cell walls, compared to those of wild-type plants. Furthermore, PpnCCT39 overexpression led to elevated lignin content and enhanced the rigidity of secondary cell walls. RNA-seq and ChIP-seq association analyses identified 826 potential regulatory target genes of PpnCCT39 that were upregulated and expressed in 1-month-old PpnCCT39 overexpression lines. Gene enrichment analyses revealed enrichment in pathways related to cell wall formation, xylem and phloem development, and the phenylpropanoid pathway. Two genes involved in lignin biosynthesis, PagCOMT1 and PagBGLU18–2, exhibited significantly increased expression in stems of lines overexpressing PpnCCT39, as demonstrated by high FPKM values and RT-qPCR results. Further investigations using yeast one-hybrid, dual-luciferase assays, and electrophoretic mobility shift assays demonstrated that PpnCCT39 directly activates the transcription of PagCOMT1 and PagBGLU18–2, thereby promoting lignin biosynthesis. This study elucidated the transcriptional regulatory mechanism of PpnCCT39 in poplars and revealed its role in activating the expression of key lignin biosynthesis genes. PpnCCT39 facilitates lignin biosynthesis and secondary growth processes, offering a novel theoretical framework for modulating lignin biosynthesis and enhancing timber yield through molecular design.

Autophagy Regulates Plant Tolerance to Submergence by Modulating Photosynthesis.

The increase in global climate variability has increased the frequency and severity of floods, profoundly affecting agricultural production and food security worldwide. Autophagy is an intracellular catabolic pathway that is dispensable for plant responses to submergence. However, the physiological role of autophagy in plant response to submergence remains unclear. In this study, a multi-omics approach was applied by combining transcriptomics, proteomics, and lipidomics to characterize molecular changes in the Arabidopsis autophagy-defective mutant (atg5-1) responding to submergence. Our results revealed that submergence resulted in remarkable changes in the transcriptome, proteome, and lipidome of Arabidopsis. Under submerged conditions, the levels of chloroplastidic lipids, including monogalactosyldiacylglycerol (MGDG), digalactosyldiacylglycerol (DGDG), and phosphatidylglycerol (PG), were lower in atg5-1 than in wild-type, suggesting that autophagy may affect photosynthesis by regulating lipid metabolism. Consistently, photosynthesis-related proteins and photosynthetic efficiency decreased in atg5-1 under submergence conditions. Phenotypic analysis revealed that inhibition of photosynthesis resulted in a decreased tolerance to submergence. Compared to wild-type plants, atg5-1 plants showed a significant decrease in starch content after submergence. Collectively, our findings reveal a novel role for autophagy in plant response to submergence via the regulation of underwater photosynthesis and starch content.

PhNH10 Suppresses Low Temperature Tolerance in Petunia Through the Abscisic Acid-Dependent Pathway.

Low-temperature stress limits plant growth, and reduces aesthetics of many ornamental plants. Plants have developed different adaptive mechanisms to cope with low-temperature stress, in which NAC transcription factor family members playing an important role in low-temperature tolerance. However, their roles in petunia in response to low temperature are still largely unknown. Here, we found that a NAC transcription factor, namely, PhNH10, negatively regulates low-temperature response in petunia. PhNH10-silenced and -CRISPR/Cas9 mutant plants displayed higher survival rate, anthocyanin content and abscisic acid concentration than PhNH10-overexpression and wild-type plants under low-temperature condition. PhNH10 can directly bind to the PhABA8ox promoter to active its expression, which further promotes the abscisic acid catabolism, while silencing of PhABA8ox increased the ABA concentration and low-temperature tolerance. In addition, PhNH10 interact with a low-temperature-related E2 ubiquitin-conjugating enzyme, PhUBC2-1, which in turn inhibited the binding capacity of PhNH10 on PhABA8ox promoter. Our research has elucidated an extensive mechanistic network underlying the PhNH10-mediated regulation of low-temperature response in petunia. This finding not only presents a new viewpoint in understanding the low-temperature tolerance mechanisms but also delineates a promising pathway for transgenic petunia with improved low-temperature resistance.

OsAMT1.1 knockout-induced decrease in cadmium absorption and accumulation by rice related to cadmium absorption-related gene downregulation

Cadmium (Cd) is a hazardous heavy metal that poses a serious risk to human health through the food chain, with rice being a significant vector because of its tendency to accumulate Cd. Nitrogen (N), an essential element for plant growth, also affects the Cd absorption and accumulation in crops. This study investigated the effects of N application on Cd absorption and accumulation in Cd-contaminated soils. Potting experiment showed that increasing N concentrations significantly increased the plant biomass and Cd contents in rice tissues. Ammonium (NH4+) transporter gene OsAMT1.1 knockout led to a substantial reduction in Cd absorption and accumulation in all rice tissues compared to that in the wild-type plants. Specifically, osamt1.1 mutants increased the Cd content in culm tissues, whereas it was reduced in brown rice. In addition, OsAMT1.1 knockout reduced Cd2+ influx in roots under NH4+-N addition, although OsAMT1.1 lacked Cd transport ability when expressed in yeast. Gene expression analysis revealed that OsAMT1.1 knockout reduced Cd absorption-related genes (OsIRT1, OsNRAMP1, and OsNRAMP5) expression levels. These finding highlight the critical role of N supply and OsAMT1.1 in regulating the Cd content in rice, offering insights into the molecular mechanisms of Cd transportation in plants.

Overexpression of MYB transcription factor LrAN2 from Lycium ruthenicum activated the anthocyanin biosynthesis in the leaf and fruit of Lycium barbarum

AN2 was thought to be the key gene inducing the fruit color differentiation of Lycium barbarum and Lycium ruthenicum, but there was no direct evidence. In this study, AN2 from L. ruhtenicum was overexpressed in L. barbarum to conform its function in regulating anthocyanin biosynthesis. The results showed that the stems, leaves, and fruits of transgenic plants were purple and contained significantly higher anthocyanin levels compared to the wild-type plants. Transcriptome analysis of leaf and fruit identified 5,832 differentially expressed genes (DEGs) in leaves and 2,029 DEGs in fruits, with 910 genes common to both tissues. GO and KEGG pathway enrichment analyses indicated a significant impact of LrAN2 on flavonoid biosynthesis and anthocyanin biosynthesis. The structural genes involved in the anthocyanin pathway, such as 4CL, CHS, F3H, F3’5’H, DFR, ANS, and UFGT, were up-regulated in transgenic lines, while the expression of transcription factors bHLH and WD40 were remained unchanged. These findings confirm the role of LrAN2 in the color differentiation of L. barbarum and L. ruthenicum and highlight its potential for the genetic enhancement of anthocyanin content in L. barbarum and other Solanaceae species.

Overexpression Analysis of PtrLBD41 Suggests Its Involvement in Salt Tolerance and Flavonoid Pathway in Populus trichocarpa.

Soil salinization is a significant environmental stress factor, threatening global agricultural yield and ecological security. Plants must effectively cope with the adverse effects of salt stress on survival and successful reproduction. Lateral Organ Boundaries (LOB) Domain (LBD) genes, a gene family encoding plant-specific transcription factors (TFs), play important roles in plant growth and development. Here, we identified and functionally characterized the LBD family TF PtrLBD41 from Populus trichocarpa, which can be induced by various abiotic stresses, including salt, dehydration, low temperature, and Abscisic Acid (ABA). Meanwhile, transgenic plants overexpressing PtrLBD41 showed a better phenotype and higher tolerance than the wild-type (WT) plants under salt stress treatment. Transcriptome analysis found that the differentially expressed genes (DEGs) between the WT and overexpression (OE) line were enriched in the flavonoid biosynthetic process, in which chalcone synthases (CHS), naringenin 3-dioxygenase (F3H), and chalcone isomerase (CHI) were significantly up-regulated under salt stress conditions through qRT-PCR analysis. Therefore, we demonstrate that PtrLBD41 plays an important role in the tolerance to salt stress in P. trichocarpa.

PP2C phosphatase Pic6 suppresses MAPK activation and disease resistance in tomato.

Type 2C protein phosphatases (PP2Cs) are essential for regulating plant immune responses to pathogens. Our study focuses on the tomato PP2C-immunity associated candidate 6 (Pic6), elucidating its role in negatively regulating pattern-triggered immunity (PTI) signaling pathways in tomato. Using reverse transcription quantitative polymerase chain reaction (RT-qPCR), we observed that treatment with microbe-associated molecular patterns (MAMPs)- flg22 and flgII-28-significantly increased Pic6 mRNA levels in wild-type (RG-PtoR) tomato plants. Pic6 features a conserved N-terminal kinase-interacting motif (KIM) and a C-terminal PP2C domain. We produced variants of Pic6 with mutations in these regions, demonstrating their involvements in negatively regulating tomato immunity. Agrobacterium-mediated transient overexpression of Pic6 resulted in enhanced growth of the bacterial pathogen Pseudomonas syringae pv. tomato (Pst) strain DC3000ΔhopQ1-1 compared to a YFP control. Additionally, Pic6 overexpression inhibited mitogen-activated protein kinase (MAPK) activation in response to flg22 and flgII-28 treatments. Importantly, Pic6 exhibited phosphatase activity and interacted with tomato Mkk1/Mkk2 proteins and dephosphorylated them in a KIM-dependent manner. Furthermore, we generated RG-pic6 loss-of-function mutants by CRISPR/Cas9, revealing that the absence of Pic6 heightened MAPK activity and increased resistance to Xanthomonas euvesicatoria strain 85-10 (Xe 85-10) when compared with the wild-type (RG-PtoR) plants. Transcript analyses showed that after flg22/flgII-28 treatment, PTI-reporter genes NAC and osmotin were significantly upregulated in RG-pic6 mutants in comparison to the wild-type (RG-PtoR) plants. Overall, our findings indicate that Pic6 acts as a negative regulator of MAPK signaling and playing a pivotal role in modulating tomato immunity against bacterial pathogens.

Functional Characterization of the 14-3-3 Gene Family in Alfalfa and the Role of MsGRF2 in Drought Response Mechanisms.

Drought stress affects crop growth and development, significantly reducing crop yield and quality. Alfalfa (Medicago sativa L.), the most widely cultivated forage crop, is particularly susceptible to drought. The general regulatory factor (GRF) protein 14-3-3, a highly conserved family in plants, specifically recognizes and binds to phosphoserine residues in target proteins, regulating both plant development and responses to environmental stressors. In this study, 66 alfalfa 14-3-3 proteins were identified, and the full-length MsGRF2 gene was cloned and functionally analyzed. The expression of MsGRF2 was highest in alfalfa inflorescences and lowest in roots. Transgenic tobacco overexpressing MsGRF2 exhibited increased tolerance to low temperature and drought stress, evidenced by physiological indicators including low levels of active oxygen species and increased activity of antioxidant enzymes and osmoregulatory substances. Under drought stress conditions, compared to wild-type plants, MsGRF2-overexpressing tobacco plants exhibited significantly increased expression of drought stress-related genes ERD10B and TIP, while the expression of BRI1, Cu/Zn-SOD, ERF2, and KC1 was significantly reduced. Together, these results provide new insights into the roles of the 14-3-3 protein MsGRF2 in plant drought response mechanisms.

Altering endogenous cytokinin content by GmCKX13 as a strategy to develop drought-tolerant plants

Climate-resilient crops are essential to meet the growing food demands of an ever-increasing world population. The phytohormone cytokinins (CKs) have been well established to regulate plant adaptation to drought. Previously, we reported that GmCKX13, encoding a CK oxidase/dehydrogenase in soybean (Glycince max), was differentially expressed under drought. Here, we further characterized its in planta function to assess if GmCKX13 could be a candidate gene to develop drought-tolerant crops. Transgenic Arabidopsis 35S:GmCKX13 plants ectopically and constitutively expressing GmCKX13 using the 35S promoter had an increase in root length, while showing a reduction in shoot height and biomass. CK contents were reduced in 35S:GmCKX13 plants, coupled with the reduced expression of all AtCKX genes and increased expression of all Arabidopsis isopentenyl transferase (AtIPT) genes, except AtIPT2 and AtIPT9 that are responsible for the production of cis-zeatin-type CKs. The 35S:GmCKX13 plants showed improved tolerance to drought and more sensitivity to the treatment with exogenous abscisic acid (ABA). Under the control of the drought-responsive promoter RD29A, the transgenic RD29A:GmCKX13 Arabidopsis plants had a similar phenotype as that of the wild-type (WT) plants under normal conditions. Under dehydration, we found a significant increase in the expression of the transgene coupled with a higher leaf relative water content in RD29A:GmCKX13 plants when compared with WT plants. In consistence with this finding, the RD29A:GmCKX13 plants exhibited higher survival rates and 30 % higher seed yield than the WT plants under drought conditions. Taken together, our results demonstrated that GmCKX13 is an excellent gene for developing drought-tolerant crops by altering endogenous CK levels and ABA responsiveness when being driven by a drought-responsive promoter.

Unravelling the different components of nonphotochemical quenching using a novel analytical pipeline.

Photoprotection in plants includes processes collectively known as nonphotochemical quenching (NPQ), which quench excess excitation-energy in photosystem II. NPQ is triggered by acidification of the thylakoid lumen, which leads to PsbS-protein protonation and violaxanthin de-epoxidase activation, resulting in zeaxanthin accumulation. Despite extensive study, questions persist about the mechanisms of NPQ. We have set up a novel analytical pipeline to disentangle NPQ induction curves measured at many light intensities into a limited number of different kinetic components. To validate the method, we applied it to Chl-fluorescence measurements, which utilised the saturating-pulse methodology, on wild-type (wt) and zeaxanthin-lacking (npq1) Arabidopsis thaliana plants. NPQ induction curves in wt and npq1 can be explained by four components ( , , and ). The fastest two ( and ) correlate with pH difference formed across the thylakoid membrane in wt and npq1. In wt, the slower component ( ) appears to be due to the formation of zeaxanthin-related quenching whilst for npq1, this component is 'replaced' by a slower component ( ), which reflects a photoinhibition-like process that appears in the absence of zeaxanthin-induced quenching. Expanding this approach will allow the effects of mutations and other abiotic-stress factors to be directly probed by changes in these underlying components.

Molecular characterization of Pleiotropic Drug Resistance (PDR) genes involved in tolerance of cadmium in peanut (Arachis hypogaea L.)

Peanut (Arachis hypogaea L.) is one of the most important oil crops worldwide. Cadmium (Cd), a heavy metal that is nonessential and toxic, has the potential to significantly impacted the quality and safety of peanut. Despite the known importance of Pleiotropic Drug Resistance (PDR) genes in heavy metal accumulation and transport in plants, there is a lack of comprehensive research on the systematic identification and functional characterization of AhPDRs in peanut. In this study, a total of 38 AhPDR genes were discovered within the peanut genome. Among these, AhPDR24, AhPDR30, and AhPDR33 displayed notable variations in expression levels in response to Cd stress. Particularly noteworthy was the observation that AhPDR33, localized in the plasma membrane, exhibited a significant increase in expression (approximately 3.8-fold) and heightened promoter activity (approximately 4.1-fold) following exposure to Cd (75 μM CdCl2). Furthermore, the study found that the overexpression of AhPDR33 in Arabidopsis resulted in increased root elongation and decreased Cd accumulation (approximately 0.42-fold) compared to wild-type plants. This suggests that AhPDR33 may have a beneficial role in facilitating Cd efflux and tolerance in plants. Additionally, transient silencing of AhPDR33 in peanut demonstrated its positive regulation of Cd tolerance through the promotion of reactive oxygen species (ROS) scavenging and membrane permeability reduction. These findings contribute to the understanding of the molecular mechanisms involved in AhPDR33-mediated Cd tolerance and detoxification in peanut. Furthermore, this study provides comprehensive information to understand the AhPDR gene family, its features, and its expression, which will hold a promising utility as an excellent candidate in the genetic improvement of peanut Cd stress tolerance.

Functional Analysis of CsWOX4 Gene Mutation Leading to Maple Leaf Type in Cucumber (Cucumis sativus L.).

The leaf morphology is an important agronomic trait in crop production. Our study identified a maple leaf type (mlt) cucumber mutant and located the regulatory gene for leaf shape changes through BSA results. Hybrid F1 and F2 populations were generated by F1 self-crossing, and the candidate mlt genes were identified within the 2.8 Mb region of chromosome 2 using map cloning. Through the sequencing and expression analysis of genes within the bulk segregant analysis (BSA) region, we identified the target gene for leaf shape regulation as CsWOX4 (CsaV3_2G026510). The change from base C to T in the original sequence led to frameshift mutations and the premature termination of translation, resulting in shortened encoded proteins and conserved WUSCHEL (WUS) box sequence loss. The specific expression analysis of the CsWOX4/Cswox4 genes in the roots, stems, leaves and other tissue types of wild-type (WT) and mutant plants revealed that CsWOX4 was higher in the root, but Cswox4 (mutant gene) was significantly higher in the leaf. Subcellular localization analysis revealed that CsWOX4 was localized in the nucleus. RNA-seq analysis revealed that the differentially expressed genes were mainly enriched in the mitochondrial cell cycle phase transition, nucleosome and microtubule binding pathways. Simultaneously, the quantitative analysis of the expression trends of 25 typical genes regulating the leaf types revealed the significant upregulation of CsPIN3. In our study, we found that the conserved domain of CsWOX4 was missing in the mutant, and the transcriptome data revealed that the expression of some genes, such as CsPIN3, changed simultaneously, thereby jointly regulating changes in the cucumber leaf type.

Genome-wide analysis of the HSF family in Allium sativum L. and AsHSFB1 overexpression in Arabidopsis under heat stress

The heat shock transcription factor (HSF) family is one of the most widely studied transcription factor families in plants; HSFs can participate in the response to various stressors, such as heat stress, high salt, and drought stress. Based on garlic transcriptome data, we screened and identified 22 garlic HSFs. The HSF proteins of garlic and Arabidopsis can be divided into three (A, B, C) subfamilies. The phylogenetic relationship, chromosome localization, sequence characteristics, conserved motifs, and promoter analysis of the HSF family were analyzed through bioinformatics methods. RT-qPCR analysis showed that the nine selected genes had different degrees of response to heat stress. In addition, we isolated and identified a class B HSF gene, AsHSFB1, from garlic variety ‘Xusuan No.6’. Subsequently, the AsHSFB1 gene was overexpressed in Arabidopsis thaliana. Under heat stress, the germination rate and growth of wild-type plants were better than that of transgenic plants. Moreover, after heat treatment, the contents of peroxidase, catalase, and chlorophyll a and b of transgenic plants were lower, but the contents of malondialdehyde (MDA) and leaf conductivity were higher. Nitroblue tetrazolium (NBT) staining showed that the stained area of transgenic plant leaves was larger than that of the wild type. Further studies showed that AsHSFB1 overexpression inhibited the expression of related reverse resistance genes. These results indicate that AsHSFB1 might play a negative regulatory role in garlic resistance under high stress. Altogether, these findings provide valuable data for revealing the function of HSF genes and lay a foundation for the subsequent selection of heat-resistant garlic varieties.