Natural resistance-associated macrophage proteins are involved in tolerance to heavy metal Cd2+ toxicity and resistance to bacterial wilt of peanut (Arachis hypogaea L.).

Peanut (Arachis hypogaea L.) is one of the most important oil and industrial crops. However, heavy-metal pollution and frequent soil diseases, poses a significant threat to the production of green and healthy peanuts. Herein, we investigated the effects of heavy metal Cd2+ toxicity to the peanuts, and screened out two peanut cultivars H108 and YZ 9102 with higher Cd2+-tolerance. RNA-seq revealed that Natural resistance-associated macrophage proteins (NRAMP)-like genes were involved in the Cd2+ stress tolerance in H108. Genome-wide identification revealed that 28, 13 and 9 Nramp-like genes existing in the A. hypogaea, A. duranensis and A. ipaensis, respectively. The 50 peanut NRAMP genes share conserved architectural characters, and they were classified into two groups. Expressions of AhNramps, particularly AhNramp4, AhNramp12, AhNramp19, and AhNramp25 could be greatly induced by not only cadmium toxicity, but also copper and zinc stresses. The expression profiles of AhNramp14, AhNramp16 and AhNramp25 showed significant differences in the H108 (resistance) and H107 (susceptible) under the infection of bacterial wilt. In addition, we found that the expression profiles of AhNramp14, AhNramp16, and AhNramp25 were greatly up- or down-regulated by the application of exogenous salicylic acid, methyl jasmonate, and abscisic acid. The AhNramp25, of which expression was affected by both heavy metal toxicity and bacterial wilt infection, were selected as strong candidate genes for peanut stress breeding. Our findings will provide an additional information required for further analysis of AhNramps involved in tolerance to heavy metal toxicity and resistance to bacterial wilt of peanut.

Unveiling the molecular regulatory mechanisms underlying sucrose accumulation and oil reduction in peanut kernels through genetic mapping and transcriptome analysis.

Sucrose content is a key factor for the flavor of edible peanut, which determines the sweet taste of fresh peanut and also attribute to pleasant flavor of roasted peanut. To explore the genetic mechanism of the sucrose content in peanut, an F2 population was created by crossing the sweet cultivar Zhonghuatian 1 (ZHT1) with Nanyangbaipi (NYBP). A genomic region spanning 28.26 kb on chromosome A06 was identified for the sucrose content through genetic mapping, elucidating 47.5% phenotypic variance explained. As the sucrose content had a significantly negative correlation with the oil content, this region was also found to be related to the oil content explaining 37.2% of phenotype variation. In this region, Arahy.42CAD1 was characterized as the most likely candidate gene through a comprehensive analysis. The nuclear localization of Arahy.42CAD1 suggests its potential involvement in the regulation of gene expression for sucrose and oil contents in peanut. Transcriptome analysis of the developing seeds in both parents revealed that genes involved in glycolysis and triacylglycerol biosynthesis pathways were not significantly down-regulated in ZHT1, indicating that the sucrose accumulation was not attributed to the suppression of triacylglycerol biosynthesis. Based on the WGCNA analysis, Arahy.42CAD1 was co-expressed with the genes involved in vesicle transport and oil body assembly, suggesting that the sucrose accumulation may be caused by disruptions in TAG transportation or storage mechanisms. These findings offer new insights into the molecular mechanisms governing sucrose accumulation in peanut, and also provide a potential gene target for enhancing peanut flavor.

Antioxidant Activities of Different Peanut (Arachis hypogaea L.) Market Types by Spectrophotometric Techniques Combined with Chemometrics.

Peanut is rich in oil and protein and has a large content of bioactive constituents consisting of tocopherols, phytosterols, and so on. Generally, Virginia, Spanish, Valencia and Runner market types are grown of peanut. In this study, it is aimed to determine the antioxidant activity, total phenolic content and total flavonoid content of peanuts from four different market types, for the first time, and group them with principal component analysis (PCA) and hierarchical cluster analysis (HCA). For PCA, PC1 and PC2 explained 87.655 % of the total variation and, according to the HCA of peanut samples, two main groups were determined. The total phenolic content changed 1.556 to 2.899 mg GAE/g. The lowest value have seen at Spanish merket type to determine the antioxidant activities of peanut samples were maked FRAP and DPPH assay, the lowest FRAP value (8.136 µmol FeSO47H2O/g sample) was seen at Valencia market type, the highest (14.004 µmol FeSO47H2O/g sample) was seen at Virginia market type. It was determined that the total flavonoid, total phenolic content, and antioxidant activities of the Virginia, Valencia, Spanish, and Runner market types included in the study were different from each other, and the Virginia market type showed superior characteristics compared to the others. The results obtained suggest that Virginia market type may be preferred more especially in peanut cultivation for food uses. It is thought that this study can be a source for future studies by eliminating a deficiency in the literature.

Genome-wide analysis of the peanut CaM/CML gene family reveals that the AhCML69 gene is associated with resistance to Ralstonia solanacearum.

BACKGROUND: Calmodulins (CaMs)/CaM-like proteins (CMLs) are crucial Ca2+-binding sensors that can decode and transduce Ca2+ signals during plant development and in response to various stimuli. The CaM/CML gene family has been characterized in many plant species, but this family has not yet been characterized and analyzed in peanut, especially for its functions in response to Ralstonia solanacearum. In this study, we performed a genome-wide analysis to analyze the CaM/CML genes and their functions in resistance to R. solanacearum. RESULTS: Here, 67, 72, and 214 CaM/CML genes were identified from Arachis duranensis, Arachis ipaensis, and Arachis hypogaea, respectively. The genes were divided into nine subgroups (Groups I-IX) with relatively conserved exon‒intron structures and motif compositions. Gene duplication, which included whole-genome duplication, tandem repeats, scattered repeats, and unconnected repeats, produced approximately 81 pairs of homologous genes in the AhCaM/CML gene family. Allopolyploidization was the main reason for the greater number of AhCaM/CML members. The nonsynonymous (Ka) versus synonymous (Ks) substitution rates (less than 1.0) suggested that all homologous pairs underwent intensive purifying selection pressure during evolution. AhCML69 was constitutively expressed in different tissues of peanut plants and was involved in the response to R. solanacearum infection. The AhCML69 protein was localized in the cytoplasm and nucleus. Transient overexpression of AhCML69 in tobacco leaves increased resistance to R. solanacearum infection and induced the expression of defense-related genes, suggesting that AhCML69 is a positive regulator of disease resistance. CONCLUSIONS: This study provides the first comprehensive analysis of the AhCaM/CML gene family and potential genetic resources for the molecular design and breeding of peanut bacterial wilt resistance.

Covalent conjugation of Inca peanut albumin and polyphenols with different phenolic hydroxyl numbers through laccase catalysis to improve functional properties.

BACKGROUND: Enzymatic crosslinking is a method that can be used to modify Inca peanut albumin (IPA) using polyphenols, and provides desirable functionalities; however, the effect of polyphenol structures on conjugate properties is unclear. In this study, we selected four polyphenols with different numbers of phenolic hydroxyl groups [para-hydroxybenzoic acid (HBA), protocatechuic acid (PCA), gallic acid (GA), and epigallocatechin gallate (EGCG)] for covalent modification of IPA by enzymatic crosslinking, and explored the structure-function changes of the IPA-polyphenol conjugates. RESULTS: Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) analysis showed that laccase successfully promoted covalent crosslinking of IPA with polyphenols, with the order of degree of conjugation as EGCG > GA > PCA > HBA, the IPA-EGCG conjugate showed the highest polyphenol binding equivalents (98.35 g kg-1 protein), and a significant reduction in the content of free amino, sulfhydryl, and tyrosine group. The oxidation of polyphenols by laccase forms quinone or semiquinone radicals that are covalently crosslinked to the reactive groups of IPA, leading to significant changes in the secondary and tertiary structures of IPA, with spherical structures transforming into dense lamellar structures. The 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging ability and emulsification stability of IPA-EGCG conjugates improved by almost 6-fold and 2.7-fold, respectively, compared with those of unmodified IPA. CONCLUSION: These data suggest that the higher the number of polyphenol hydroxyl groups, the higher the degree of IPA-polyphenol conjugation; additionally, enzymatic crosslinking can significantly improve the functional properties of IPA. © 2024 Society of Chemical Industry.

Identification and in vitro Characterization of Novel Antidiabetic Peptides Released Enzymatically from Peanut Protein.

Bioactive peptides derived from proteins found in various foods provide significant health benefits, including regulating blood sugar levels by inhibiting carbohydrate-hydrolyzing enzymes. Hydrolysates of peanut protein were prepared using alcalase (AH) or trypsin (TH) to generate antidiabetic peptides with high activity against α-amylase (IC50 of 6.46 and 5.71 mg/mL) and α-glucosidase (IC50 of 6.30 and 5.57 mg/mL), as well as antiradical activity to scavenge DPPH• (IC50 of 4.18 and 3.12 mg/mL) and ABTS•+ (IC50 of 2.87 and 2.56 mg/mL), respectively. The bioactivities of hydrolysates were greatest in the ultrafiltration-generated F3 fraction (< 3 kDa). The most active fraction was TH-F3, which was purified by gel filtration chromatography to generate sub-fractions (SF). With IC50 values of 1.05 and 0.69 mg/mL, the F3-SF8 fraction was the most effective at inhibiting the activity of α-amylase and α-glucosidase, respectively. This fraction was further purified using RP-HPLC to generate sub-subfractions (SSF), the most active of which were F3-SF8-SSF9 and SSF10. The peptide sequences F3-SF8-SSF9 and SSF10 were determined using LC-MS/MS. Two novel antidiabetic peptides with the potential to inhibit α-amylase and α-glucosidase were identified, with the sequences Asp-Trp-Arg (476.22 Da, IC50 of 0.78, and 0.35 mg/mL) and Phe-Tyr (329.15 Da, IC50 of 0.91, and 0.41 mg/mL). These results suggest that peptides derived from peanut protein are attractive natural ingredients for diabetes management applications.

Metabolic profiles of peanut (Arachis hypogaea L.) in response to Puccinia arachidis fungal infection.

Background Puccinia arachidis fungus causes rust disease in the peanut plants (Arachis hypogaea L.), which leads to high yield loss. Metabolomic profiling of Arachis hypogaea was performed to identify the pathogen-induced production of metabolites involved in the defense mechanism of peanut plants. In this study, two peanut genotypes, one susceptible (JL-24) and one resistant (GPBD-4) were inoculated with Puccinia arachidis fungal pathogen. The metabolic response was assessed at the control stage (0 day without inoculation), 2 DAI (Day after inoculation), 4 DAI and 6 DAI by Gas Chromatography-Mass Spectrometry (GC-MS). Results About 61 metabolites were identified by NIST library, comprising sugars, phenols, fatty acids, carboxylic acids and sugar alcohols. Sugars and fatty acids were predominant in leaf extracts compared to other metabolites. Concentration of different metabolites such as salicylic acid, mannitol, flavonoid, 9,12-octadecadienoic acid, linolenic acid and glucopyranoside were higher in resistant genotype than in susceptible genotype during infection. Systemic acquired resistance (SAR) and hypersensitive reaction (HR) components such as oxalic acid was elevated in resistant genotype during pathogen infection. Partial least square-discriminant analysis (PLS-DA) was applied to GC-MS data for revealing metabolites profile between resistant and susceptible genotype during infection. Conclusion The phenol content and oxidative enzyme activity i.e. catalase, peroxidase and polyphenol oxidase were found to be very high at 4 DAI in resistant genotype (p-value < 0.01). This metabolic approach provides information about bioactive plant metabolites and their application in crop protection and marker-assisted plant breeding.

Alternative polyadenylation regulates acetyl-CoA carboxylase function in peanut.

BACKGROUND: Polyadenylation is a crucial process that terminates mRNA molecules at their 3'-ends. It has been observed that alternative polyadenylation (APA) can generate multiple transcripts from a single gene locus, each with different polyadenylation sites (PASs). This leads to the formation of several 3' untranslated regions (UTRs) that vary in length and composition. APA has a significant impact on approximately 60-70% of eukaryotic genes and has far-reaching implications for cell proliferation, differentiation, and tumorigenesis. RESULTS: In this study, we conducted long-read, single-molecule sequencing of mRNA from peanut seeds. Our findings revealed that over half of all peanut genes possess over two PASs, with older developing seeds containing more PASs. This suggesting that the PAS exhibits high tissue specificity and plays a crucial role in peanut seed maturation. For the peanut acetyl-CoA carboxylase A1 (AhACCA1) gene, we discovered four 3' UTRs referred to UTR1-4. RT-PCR analysis showed that UTR1-containing transcripts are predominantly expressed in roots, leaves, and early developing seeds. Transcripts containing UTR2/3 accumulated mainly in roots, flowers, and seeds, while those carrying UTR4 were constitutively expressed. In Nicotiana benthamiana leaves, we transiently expressed all four UTRs, revealing that each UTR impacted protein abundance but not subcellular location. For functional validation, we introduced each UTR into yeast cells and found UTR2 enhanced AhACCA1 expression compared to a yeast transcription terminator, whereas UTR3 did not. Furthermore, we determined ACC gene structures in seven plant species and identified 51 PASs for 15 ACC genes across four plant species, confirming that APA of the ACC gene family is universal phenomenon in plants. CONCLUSION: Our data demonstrate that APA is widespread in peanut seeds and plays vital roles in peanut seed maturation. We have identified four 3' UTRs for AhACCA1 gene, each showing distinct tissue-specific expression patterns. Through subcellular location experiment and yeast transformation test, we have determined that UTR2 has a stronger impact on gene expression regulation compared to the other three UTRs.

Identification and functional characterization of REGULATORY PARTICLE NON-ATPASE 1a-like (AhRPN1a-like) in peanuts during aluminum-induced programmed cell death.

The toxicity of aluminum (Al) in acidic soil is a prevalent problem and causes reduced crop yields. In the plant response to Al toxicity, programmed cell death (PCD) appears to be one of the important mechanisms. However, the regulation of Al-induced PCD remains poorly understood. Here, we found that an uncharacterized protein REGULATORY PARTICLE NON-ATPASE 1a-like in peanut (AhRPN1a-like), located in the nucleus and cytoplasm, directly interacted with type I metacaspase in peanut (AhMC1). The overexpression of AhRPN1a-like in Arabidopsis strongly enhanced Al inhibition of root growth with a loss of root tip cell viability. Furthermore, in response to Al treatment, the VIGS knockdown line of AhRPN1a-like in peanut displayed decreased transcription of AhMC1, increased root growth, reduced Al-induced PCD and decreased 26S proteasomal activity. Taken together, these findings demonstrated that AhRPN1a-like interacted directly with AhMC1, and promotes the occurrence of Al-induced PCD via the 26S proteasome pathway, thereby reducing Al-resistance.

Lycopene alleviates food allergy by modulating the PI3K/AKT pathway in peanut-sensitized BALB/c mice.

Food allergies, which lead to life-threatening acute symptoms, are considered an important public health problem. Therefore, it is essential to develop efficient preventive and treatment measures. We developed a crude peanut protein extract (PPE)-induced allergy mouse model to investigate the effects of lycopene on peanut allergy. Mice were divided into four groups: 5 mg/kg lycopene, 20 mg/kg lycopene, no treatment, and control groups. Serum inflammatory factors were detected using enzyme-linked immunosorbent assay. In addition, pathology and immunohistochemistry analyses were used to examine the small intestine of mice. We found that lycopene decreased PPE-specific immunoglobulin E (IgE) and IL-13 levels in the serum, relieved small intestine inflammation, attenuated the production of histamine and mouse mast cell protease-1, and downregulated PI3K and AKT1 expression in the small intestine tissues of mice allergic to peanuts. Our results suggest that lycopene can ameliorate allergy by attenuating the PI3K/AKT pathway and the anaphylactic reactions mediated by PPE-specific IgE.

Purification, Identification, Chelation Mechanism, and Calcium Absorption Activity of a Novel Calcium-Binding Peptide from Peanut (Arachis hypogaea) Protein Hydrolysate.

A novel calcium-binding peptide was purified from peanut protein hydrolysate using gel filtration chromatography and identified using HPLC-MS/MS. Its amino acid sequence was determined as Phe-Pro-Pro-Asp-Val-Ala (FPPDVA, named as FA6) with the calcium-binding capacity of 15.67 ± 0.39 mg/g. Then, the calcium chelating characteristics of FPPDVA were investigated using ultraviolet-visible absorption spectroscopy, fluorescence spectroscopy, Fourier transform infrared spectroscopy, particle size, and zeta potential. The results showed that FPPDVA interacted with calcium ions, the chelation of calcium ions induced FPPDVA to fold and form a denser structure, the calcium-binding sites may mainly involve oxygen atoms from the carboxyl residues of Asp and Ala, and Phe possessed contact energy and carbonyl residues of Val. Microstructure analysis showed that FPPDVA-calcium chelate exhibited a regularly ordered and tightly aggregated sheets or block structures. Additionally, FPPDVA-calcium chelate had good gastrointestinal digestive stability and thermal stability. The results of everted rat intestinal sac and Caco-2 cell monolayer experiments showed that FPPDVA-calcium chelate could promote calcium absorption and transport through the Cav1.3 and TRPV6 calcium channels. These data suggest that FPPDVA-calcium chelate possesses the potential to be developed and applied as calcium supplement.

Novel Umami-, Salty-, and Kokumi-Enhancing γ-Glutamyl Tripeptides Synthesized with the Bitter Dipeptides from Defatted Peanut Meal Protein Hydrolysate.

Defatted peanut meal protein hydrolysates (DPMHs) usually have a bitter taste. γ-Glutamylation by Bacillus amyloliquefaciens l-glutaminase was introduced to DPMH to reduce its bitterness and generated a γ-glutamylated product (DPMH-G). Extra l-glutamine (l-Gln) (5% w/w) was added to DPMH, and the mixture was then γ-glutamylated (DPMH-G-Q). Results showed that γ-glutamylation decreased the bitterness of the products and also enhanced their kokumi, umami, and salty taste, especially for DPMH-G-Q. Bitter amino acids and bitter peptides were found to be substrates (acceptors) of the synthesized γ-[Glu](1,2)-AAs and γ-Glu-AA-AAs, respectively. The production yield of γ-[Glu](1,2)-AAs was only 0.69/100 g for DPMH-G and 2.30/100 g for DPMH-G-Q, which was much lower than that of γ-Glu-AA-AAs (5.73/100 g for DPMH-G and 18.72/100 g for DPMH-G-Q). The improvement in taste attributes of DPMH might mainly be due to the consumption of bitter dipeptides and the production of γ-Glu-AA-AAs. In DPMH-G-Q, eight γ-Glu-AA-AAs were identified, including γ-Glu-Ile-Lys, γ-Glu-Ala-Ile, γ-Glu-Leu-Leu, γ-Glu-Phe-Leu, γ-Glu-Thr-Leu, γ-Glu-Ile-Met, γ-Glu-Val-Leu, and γ-Glu-Ser-Tyr, which were first time reported. They all can enhance umami, salty, and kokumi taste with a threshold value between 1.61 ± 0.21-2.16 ± 0.19, 1.65 ± 0.19-2.23 ± 0.20, and 0.67 ± 0.21-1.00 ± 0.22 mM, respectively. Insufficient l-Gln restricted the formation of γ-glutamyl peptides, and this was why DPMH-G had a lower yield and variety than DPMH-G-Q. This also suggested that l-glutaminase is selective to different substrates. Overall, this study provides a new method to reduce the bitterness of protein hydrolysates and also improve the taste by synthesizing γ-glutamyl tripeptides.

Combining elicitor treatment of chitosan, methyl jasmonate, and cyclodextrin to induce the generation of immune response bioactive peptides in peanut hairy root culture.

The endogenous peptides from peanut hairy root culture were induced upon elicitor treatment with chitosan (CHT), methyl jasmonate (MeJA), and cyclodextrin (CD): CHT+MeJA+CD. The peptides secreted into the liquid culture medium play an important role in plant signaling and stress responses. By performing gene ontology (GO) analysis, a number of plant proteins involved in biotic and abiotic defense responses were identified, such as endochitinase, defensin, antifungal protein, cationic peroxidase and Bowman-Birk type protease inhibitor A-II. The bioactivity of 14 peptides synthesized from secretome analysis was determined. Peptide BBP1-4, derived from the diverse region of Bowman-Birk type protease inhibitor, displayed high antioxidant activity and mimicked the property of chitinase and ß-1,3-glucanase enzymes. The antimicrobial activity against S. aureus, S. typhimurium, and E. coli was evidenced with different peptide concentrations. Additionally, peptide BBP1-4 has the potential to be a useful candidate for an immune response property, as it was found to increase the expression of some pathogenesis-related (PR) proteins and stilbene biosynthesis genes in peanut hairy root tissues. The findings indicate that secreted peptides may play a role in plant responses to both abiotic and biotic stresses. These peptides, which possess bioactive properties, could be considered as potential candidates for use in the pharmaceutical, agricultural, and food industries.

Effects of the polypeptide from peanut meal mixed fermentation on lipid metabolism and intestinal flora of hyperlipidemic mice.

BACKGROUND: Hyperlipidemia is one of the metabolic disorders posing great threat to human health. Our previous studies have shown that the nutritional properties of peanut meal after fermentation are markedly improved, and can effectively improve hyperlipidemia caused by high-fat diet in mice. In this study, in order to facilitate the further utilization of peanut meal, the effect of peanut polypeptide (PP) from peanut meal mixed fermentation on lipid metabolism in mice fed with high-fat diet (HFD) and its possible mechanism were investigated. Fifty male C57BL/6J mice were randomly divided into five groups: normal control group (N), high-fat model group (M), PP low-dose group (PL), PP high-dose group (PH), and atorvastatin positive control group (Y). RESULTS: The results show that PP supplementation can effectively reduce the body weight of mice, decrease the serum total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C) and leptin levels (P < 0.05), increase the high-density lipoprotein cholesterol (HDL-C) levels (P < 0.05), up-regulate the expression levels of ileal tight junction proteins ZO-1 and occludin (P < 0.05), reduce the hepatocyte injury and lipid accumulation caused by high-fat diet and increase the species richness of intestinal flora. CONCLUSION: PP can significantly improve hyperlipidemia and regulate intestinal flora disorders caused by hyperlipidemia. The possible mechanism may be related to the reduction of serum leptin levels and up-regulating the expression levels of the ileal tight junction proteins ZO-1 and occludin. This study provides evidence for its regulatory role in lipid metabolism and intestinal function, and provides a research basis for the potential nutritional benefits of underutilized food by-products. © 2023 Society of Chemical Industry.

Genome-Wide Identification and Expression Analysis Reveals Roles of the NRAMP Gene Family in Iron/Cadmium Interactions in Peanut.

The natural resistance-associated macrophage protein (NRAMP) family plays crucial roles in metal uptake and transport in plants. However, little is known about their functions in peanut. To understand the roles of AhNRAMP genes in iron/cadmium interactions in peanut, genome-wide identification and bioinformatics analysis was performed. A total of 15 AhNRAMP genes were identified from the peanut genome, including seven gene pairs derived from whole-genome duplication and a segmental duplicated gene. AhNRAMP proteins were divided into two distinct subfamilies. Subfamily I contains eight acid proteins with a specific conserved motif 7, which were predicted to localize in the vacuole membrane, while subfamily II includes seven basic proteins sharing specific conserved motif 10, which were localized to the plasma membrane. Subfamily I genes contained four exons, while subfamily II had 13 exons. AhNRAMP proteins are perfectly modeled on the 5m94.1.A template, suggesting a role in metal transport. Most AhNRAMP genes are preferentially expressed in roots, stamens, or developing seeds. In roots, the expression of most AhNRAMPs is induced by iron deficiency and positively correlated with cadmium accumulation, indicating crucial roles in iron/cadmium interactions. The findings provide essential information to understand the functions of AhNRAMPs in the iron/cadmium interactions in peanuts.

Identification and functional characterization of the xyloglucan endotransglucosylase/hydrolase 32 (AhXTH32) in peanut during aluminum-induced programmed cell death.

The toxicity of aluminum (Al) in acidic soil is a prevalent problem and causes reduced crop yields. In the plant response to Al toxicity, programmed cell death (PCD) appears to be an important mechanism. The plant cell wall of crop roots is the predominant site targeted by Al. Here, studies of the capacities of different cell wall constituents (pectin, hemicellulose 1 {HC1} and HC2) to adsorb Al indicated that HC1 has the greater ability to bind Al. The activity of xyloglucan endotransglucosylase (XET) was significantly inhibited by Al in the Al-tolerant peanut cultivar '99-1507' compared to that in 'ZH 2' (Al-sensitive). Results from qPCR analysis suggested that the suppression of XET activity by Al was transcriptionally regulated and that xyloglucan endotransglucosylase/hydrolase 32 (AhXTH32) was the major contributor to these changes. The overexpression of AhXTH32 in Arabidopsis strongly inhibited root growth with a loss of viability in root cells and the occurrence of typical hallmarks of PCD, while largely opposite effects were observed after xth32 suppression. AhXTH32 contributed to the modulation XET and xyloglucan endohydrolase (XEH) activity in vivo. Taken together, our results demonstrate that Al-tolerant peanut cultivar root tips cell walls bind Al predominantly in the HC1 fraction, which results in the inhibition of AhXTH32, with consequences to root growth, Al sensitivity, the occurrence of PCD and the XET/XEH activity ratio.

Introgression of a novel gene AhBINR differentially expressed during PGPR Brachybacterium saurashtrense-Arachis hypogaea interaction enhances plant performance under nitrogen starvation and salt stress in tobacco.

Plant growth-promoting rhizobacteria provide endurance during environmental stress conditions. Previously, we have shown that the interaction of the halotolerant diazotrophic bacteria Brachybacterium saurashtrense JG06 induces physio-biochemical and molecular changes in Arachis hypogaea under nitrogen starvation conditions. Here we deciphered the role of a novel gene AhBINR that was differentially overexpressed in A. hypogaea after interaction with B. saurashtrense JG06 under nitrogen deficit conditions. Overexpression of the AhBINR gene in the model plant (tobacco) showed higher growth parameters (root length, shoot length, fresh weight, and dry weight) under nitrogen starvation and salt stress in comparison to the wild type and vector control. Transgenic plants were enabled with a higher photosynthesis rate, which provides the support for better performance under N2 starvation and salt stress. Results showed that the transgenic plants overexpressing the AhBINR gene had better physiological status and lower ROS accumulation under adverse conditions. Microarray transcriptome analysis showed that the transcription factors, biotic and abiotic stress, photosynthesis, and metabolism-related genes were differentially expressed (total 736 and 6530 genes were expressed under nitrogen deficit and salt stress conditions, respectively at a 5-fold change level) in comparison to wild type plants. Overall results showed the involvement of the AhBINR gene in the activation of the abiotic stress (nitrogen starvation and salt stress) related pathways, which can be overexpressed after legume-rhizobacterial interaction.

Genome-wide exploration of sugar transporter (sweet) family proteins in Fabaceae for Sustainable protein and carbon source.

Sugar transporter proteins (STPs) are membrane proteins required for sugar transport throughout cellular membranes. They plays an imperative role in sugar transmission across the plant and determinants of crop yield. However, the analysis of these important STPs Sugars Will Eventually be Exported Transporters (SWEET) family in legumes is still not well-documented and remains unclear. Therefore, the in-silico analysis of STPs has been performed to unravel their cellular, molecular, and structural composition in legume species. This study conducted a systematic search for STPs in Cajanus cajan using the Blastp algorithm to understand its molecular basis. Here, we performed a comprehensive analysis of 155 identified SWEET proteins across 12 legumes species, namely (Cajanus cajan, Glycine max, Vigna radiate, Vigna angularis, Medicago truncatula, Lupinus angustifolius, Glycine soja, Spatholobus suberectus, Cicer arietinum, Arachis ipaensis, Arachis hypogaea, Arachis duranensis). The amino acid composition and motif analysis revealed that SWEET proteins are rich in essential amino acids such as leucine, valine, isoleucine, phenylalanine, and serine while less profuse in glutamine, tryptophan, cysteine, and histidine. A total of four main conserved motifs of SWEET proteins are also highly abundant in these amino acids. The present study deciphered the details on primary physicochemical properties, secondary, tertiary structure, and phylogenetic analysis of SWEETs protein. Majorities of SWEET proteins (72.26%) are in stable form with an average instability index of 36.5%, and it comprises a higher fraction of positively charged amino acid Arg + Lys residues. Secondary structure analysis shown that these proteins are richer in alpha-helix (40%) than extended strand (30%) and random coil (25%), respectively. Furthermore, to infer their mechanism at a structural and functional level which play an essential roles in growth, development, and stress responses. This study will be useful to examine photosynthetic productivity, embryo sugar content, seed quality, and yield enhancement in Fabaceae for a sustainable source of essential amino acids and carbon source.

Differential Physio-Biochemical and Metabolic Responses of Peanut (Arachis hypogaea L.) under Multiple Abiotic Stress Conditions.

The frequency and severity of extreme climatic conditions such as drought, salinity, cold, and heat are increasing due to climate change. Moreover, in the field, plants are affected by multiple abiotic stresses simultaneously or sequentially. Thus, it is imperative to compare the effects of stress combinations on crop plants relative to individual stresses. This study investigated the differential regulation of physio-biochemical and metabolomics parameters in peanut (Arachis hypogaea L.) under individual (salt, drought, cold, and heat) and combined stress treatments using multivariate correlation analysis. The results showed that combined heat, salt, and drought stress compounds the stress effect of individual stresses. Combined stresses that included heat had the highest electrolyte leakage and lowest relative water content. Lipid peroxidation and chlorophyll contents did not significantly change under combined stresses. Biochemical parameters, such as free amino acids, polyphenol, starch, and sugars, significantly changed under combined stresses compared to individual stresses. Free amino acids increased under combined stresses that included heat; starch, sugars, and polyphenols increased under combined stresses that included drought; proline concentration increased under combined stresses that included salt. Metabolomics data that were obtained under different individual and combined stresses can be used to identify molecular phenotypes that are involved in the acclimation response of plants under changing abiotic stress conditions. Peanut metabolomics identified 160 metabolites, including amino acids, sugars, sugar alcohols, organic acids, fatty acids, sugar acids, and other organic compounds. Pathway enrichment analysis revealed that abiotic stresses significantly affected amino acid, amino sugar, and sugar metabolism. The stress treatments affected the metabolites that were associated with the tricarboxylic acid (TCA) and urea cycles and associated amino acid biosynthesis pathway intermediates. Principal component analysis (PCA), partial least squares-discriminant analysis (PLS-DA), and heatmap analysis identified potential marker metabolites (pinitol, malic acid, and xylopyranose) that were associated with abiotic stress combinations, which could be used in breeding efforts to develop peanut cultivars that are resilient to climate change. The study will also facilitate researchers to explore different stress indicators to identify resistant cultivars for future crop improvement programs.

Illegitimate Recombination between Duplicated Genes Generated from Recursive Polyploidizations Accelerated the Divergence of the Genus Arachis.

The peanut (Arachis hypogaea L.) is the leading oil and food crop among the legume family. Extensive duplicate gene pairs generated from recursive polyploidizations with high sequence similarity could result from gene conversion, caused by illegitimate DNA recombination. Here, through synteny-based comparisons of two diploid and three tetraploid peanut genomes, we identified the duplicated genes generated from legume common tetraploidy (LCT) and peanut recent allo-tetraploidy (PRT) within genomes. In each peanut genome (or subgenomes), we inferred that 6.8-13.1% of LCT-related and 11.3-16.5% of PRT-related duplicates were affected by gene conversion, in which the LCT-related duplicates were the most affected by partial gene conversion, whereas the PRT-related duplicates were the most affected by whole gene conversion. Notably, we observed the conversion between duplicates as the long-lasting contribution of polyploidizations accelerated the divergence of different Arachis genomes. Moreover, we found that the converted duplicates are unevenly distributed across the chromosomes and are more often near the ends of the chromosomes in each genome. We also confirmed that well-preserved homoeologous chromosome regions may facilitate duplicates' conversion. In addition, we found that these biological functions contain a higher number of preferentially converted genes, such as catalytic activity-related genes. We identified specific domains that are involved in converted genes, implying that conversions are associated with important traits of peanut growth and development.