Promoter variations of ClERF1 gene determines flesh firmness in watermelon.

BACKGROUND: Flesh firmness is a critical factor that influences fruit storability, shelf-life and consumer's preference as well. However, less is known about the key genetic factors that are associated with flesh firmness in fresh fruits like watermelon. RESULTS: In this study, through bulk segregant analysis (BSA-seq), we identified a quantitative trait locus (QTL) that influenced variations in flesh firmness among recombinant inbred lines (RIL) developed from cross between the Citrullus mucosospermus accession ZJU152 with hard-flesh and Citrullus lanatus accession ZJU163 with soft-flesh. Fine mapping and sequence variations analyses revealed that ethylene-responsive factor 1 (ClERF1) was the most likely candidate gene for watermelon flesh firmness. Furthermore, several variations existed in the promoter region between ClERF1 of two parents, and significantly higher expressions of ClERF1 were found in hard-flesh ZJU152 compared with soft-flesh ZJU163 at key developmental stages. DUAL-LUC and GUS assays suggested much stronger promoter activity in ZJU152 over ZJU163. In addition, the kompetitive allele-specific PCR (KASP) genotyping datasets of RIL populations and germplasm accessions further supported ClERF1 as a possible candidate gene for fruit flesh firmness variability and the hard-flesh genotype might only exist in wild species C. mucosospermus. Through yeast one-hybrid (Y1H) and dual luciferase assay, we found that ClERF1 could directly bind to the promoters of auxin-responsive protein (ClAux/IAA) and exostosin family protein (ClEXT) and positively regulated their expressions influencing fruit ripening and cell wall biosynthesis. CONCLUSIONS: Our results indicate that ClERF1 encoding an ethylene-responsive factor 1 is associated with flesh firmness in watermelon and provide mechanistic insight into the regulation of flesh firmness, and the ClERF1 gene is potentially applicable to the molecular improvement of fruit-flesh firmness by design breeding.

Comprehensive Profiling of Alternative Splicing and Alternative Polyadenylation during Fruit Ripening in Watermelon (Citrullus lanatus).

Fruit ripening is a highly complicated process that is accompanied by the formation of fruit quality. In recent years, a series of studies have demonstrated post-transcriptional control play important roles in fruit ripening and fruit quality formation. Till now, the post-transcriptional mechanisms for watermelon fruit ripening have not been comprehensively studied. In this study, we conducted PacBio single-molecule long-read sequencing to identify genome-wide alternative splicing (AS), alternative polyadenylation (APA) and long non-coding RNAs (lncRNAs) in watermelon fruit. In total, 6,921,295 error-corrected and mapped full-length non-chimeric (FLNC) reads were obtained. Notably, more than 42,285 distinct splicing isoforms were derived from 5,891,183 intron-containing full-length FLNC reads, including a large number of AS events associated with fruit ripening. In addition, we characterized 21,506 polyadenylation sites from 11,611 genes, 8703 of which have APA sites. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that fructose and mannose metabolism, starch and sucrose metabolism and carotenoid biosynthesis were both enriched in genes undergoing AS and APA. These results suggest that post-transcriptional regulation might potentially have a key role in regulation of fruit ripening in watermelon. Taken together, our comprehensive PacBio long-read sequencing results offer a valuable resource for watermelon research, and provide new insights into the molecular mechanisms underlying the complex regulatory networks of watermelon fruit ripening.

Clorf Encodes Carotenoid Isomerase and Regulates Orange Flesh Color in Watermelon (Citrullus lanatus L.).

Flesh color is a significant characteristic of watermelon. Although various flesh-color genes have been identified, the inheritance and molecular basis of the orange flesh trait remain relatively unexplored. In the present study, the genetic analysis of six generations derived from W1-1 (red flesh) and W1-61 (orange flesh) revealed that the orange flesh color trait was regulated by a single recessive gene, Clorf (orange flesh). Bulk segregant analysis (BSA) locked the range to ∼4.66 Mb, and initial mapping situated the Clorf locus within a 688.35-kb region of watermelon chromosome 10. Another 1,026 F2 plants narrowed the Clorf locus to a 304.62-kb region containing 32 candidate genes. Subsequently, genome sequence variations in this 304.62-kb region were extracted for in silico BSA strategy among 11 resequenced lines (one orange flesh and ten nonorange flesh) and finally narrowed the Clorf locus into an 82.51-kb region containing nine candidate genes. Sequence variation analysis of coding regions and gene expression levels supports Cla97C10G200950 as the most possible candidate for Clorf, which encodes carotenoid isomerase (Crtiso). This study provides a genetic resource for investigating the orange flesh color of watermelon, with Clorf malfunction resulting in low lycopene accumulation and, thus, orange flesh.

Transcriptomic and metabolic profiling of watermelon uncovers the role of salicylic acid and flavonoids in the resistance to cucumber green mottle mosaic virus.

Understanding the mechanisms underlying plant resistance to virus infections is crucial for viral disease management in agriculture. However, the defense mechanism of watermelon (Citrullus lanatus) against cucumber green mottle mosaic virus (CGMMV) infection remains largely unknown. In this study, we performed transcriptomic, metabolomic, and phytohormone analyses of a CGMMV susceptible watermelon cultivar 'Zhengkang No.2' ('ZK') and a CGMMV resistant wild watermelon accession PI 220778 (PI) to identify the key regulatory genes, metabolites, and phytohormones responsible for CGMMV resistance. We then tested several phytohormones and metabolites for their roles in watermelon CGMMV resistance via foliar application, followed by CGMMV inoculation. Several phenylpropanoid metabolism-associated genes and metabolites, especially those involved in the flavonoid biosynthesis pathway, were found to be significantly enriched in the CGMMV-infected PI plants compared with the CGMMV-infected 'ZK' plants. We also identified a gene encoding UDP-glycosyltransferase (UGT) that is involved in kaempferol-3-O-sophoroside biosynthesis and controls disease resistance, as well as plant height. Additionally, salicylic acid (SA) biogenesis increased in the CGMMV-infected 'ZK' plants, resulting in the activation of a downstream signaling cascade. SA levels in the tested watermelon plants correlated with that of total flavonoids, and SA pre-treatment up-regulated the expression of flavonoid biosynthesis genes, thus increasing the total flavonoid content. Furthermore, application of exogenous SA or flavonoids extracted from watermelon leaves suppressed CGMMV infection. In summary, our study demonstrates the role of SA-induced flavonoid biosynthesis in plant development and CGMMV resistance, which could be used to breed for CGMMV resistance in watermelon.

ClSnRK2.3 negatively regulates watermelon fruit ripening and sugar accumulation.

Watermelon (Citrullus lanatus) as non-climacteric fruit is domesticated from the ancestors with inedible fruits. We previously revealed that the abscisic acid (ABA) signaling pathway gene ClSnRK2.3 might influence watermelon fruit ripening. However, the molecular mechanisms are unclear. Here, we found that the selective variation of ClSnRK2.3 resulted in lower promoter activity and gene expression level in cultivated watermelons than ancestors, which indicated ClSnRK2.3 might be a negative regulator in fruit ripening. Overexpression (OE) of ClSnRK2.3 significantly delayed watermelon fruit ripening and suppressed the accumulation of sucrose, ABA and gibberellin GA4 . Furthermore, we determined that the pyrophosphate-dependent phosphofructokinase (ClPFP1) in sugar metabolism pathway and GA biosynthesis enzyme GA20 oxidase (ClGA20ox) could be phosphorylated by ClSnRK2.3 and thereby resulting in accelerated protein degradation in OE lines and finally led to low levels of sucrose and GA4 . Besides that, ClSnRK2.3 phosphorylated homeodomain-leucine zipper protein (ClHAT1) and protected it from degradation to suppress the expression of the ABA biosynthesis gene 9'-cis-epoxycarotenoid dioxygenase 3 (ClNCED3). These results indicated that ClSnRK2.3 negatively regulated watermelon fruit ripening by manipulating the biosynthesis of sucrose, ABA and GA4 . Altogether, these findings revealed a novel regulatory mechanism in non-climacteric fruit development and ripening.

FonTup1 functions in growth, conidiogenesis and pathogenicity of Fusarium oxysporum f. sp. niveum through modulating the expression of the tricarboxylic acid cycle genes.

The Tup1-Cyc8 complex is a highly conserved transcriptional corepressor that regulates intricate genetic network associated with various biological processes in fungi. Here, we report the role and mechanism of FonTup1 in regulating physiological processes and pathogenicity in watermelon Fusarium wilt fungus, Fusarium oxysporum f. sp. niveum (Fon). FonTup1 deletion impairs mycelial growth, asexual reproduction, and macroconidia morphology, but not macroconidial germination in Fon. The ΔFontup1 mutant exhibits altered tolerance to cell wall perturbing agent (congo red) and osmotic stressors (sorbitol or NaCl), but unchanged sensitivity to paraquat. The deletion of FonTup1 significantly decreases the pathogenicity of Fon toward watermelon plants through attenuating the ability to colonize and grow within the host. Transcriptome analysis revealed that FonTup1 regulates primary metabolic pathways, including the tricarboxylic acid (TCA) cycle, via altering the expression of corresponding genes. Downregulation of three malate dehydrogenase genes, FonMDH1-3, occurs in ΔFontup1, and disruption of FonMDH2 causes significant abnormalities in mycelial growth, conidiation, and virulence of Fon. These findings demonstrate that FonTup1, as a global transcriptional corepressor, plays crucial roles in different biological processes and pathogenicity of Fon through regulating various primary metabolic processes, including the TCA cycle. This study highlights the importance and molecular mechanism of the Tup1-Cyc8 complex in multiple basic biological processes and pathogenicity of phytopathogenic fungi.

Exogenous Brassinosteroid Enhances Zinc tolerance by activating the Phenylpropanoid Biosynthesis pathway in Citrullus lanatus L.

Zinc (Zn) is an important element in plants, but over-accumulation of Zn is harmful. The phytohormone brassinosteroids (BRs) play a key role in regulating plant growth, development, and response to stress. However, the role of BRs in watermelon (Citrullus lanatus L.) under Zn stress, one of the most important horticultural crops, remains largely unknown. In this study, we revealed that 24-epibrassinolide (EBR), a bioactive BR enhanced Zn tolerance in watermelon plants, which was related to the EBR-induced increase in the fresh weight, chlorophyll content, and net photosynthetic rate (Pn) and decrease in the content of hydrogen peroxide (H2O2), malondialdehyde (MDA), and Zn in watermelon leaves. Through RNA deep sequencing (RNA-seq), 350 different expressed genes (DEG) were found to be involved in the response to Zn stress after EBR treatment, including 175 up-regulated DEGs and 175 down-regulated DEGs. The up-regulated DEGs were significantly enriched in 'phenylpropanoid biosynthesis' pathway (map00940) using KEGG enrichment analysis. The gene expression levels of PAL, 4CL, CCR, and CCoAOMT, key genes involved in phenylpropanoid pathway, were significantly induced after EBR treatment. In addition, compared with Zn stress alone, EBR treatment significantly promoted the activities of PAL, 4CL, and POD by 30.90%, 20.69%, and 47.28%, respectively, and increased the content of total phenolic compounds, total flavonoids, and lignin by 23.02%, 40.37%, and 29.26%, respectively. The present research indicates that EBR plays an active role in strengthening Zn tolerance, thus providing new insights into the mechanism of BRs enhancing heavy metal tolerance.

Argonaute 1 and 5 proteins play crucial roles in the defence against cucumber green mottle mosaic virus in watermelon.

RNA silencing, a core part of plants' antiviral defence, requires the ARGONAUTE, DICER-like, and RNA-dependent RNA polymerase proteins. However, how these proteins contribute to watermelon's RNA interference (RNAi) pathway response to cucumber green mottle mosaic virus (CGMMV) has not been characterized. Here, we identify seven ClAGO, four ClDCL, and 11 ClRDR genes in watermelon and analyse their expression profiles when infected with CGMMV. ClAGO1 and ClAGO5 expression levels were highly induced by CGMMV infection. The results of ClAGO1 and ClAGO5 overexpression and silencing experiments suggest that these genes play central roles in watermelon's antiviral defence. Furthermore, co-immunoprecipitation and bimolecular fluorescence complementation experiments showed that ClAGO1 interacts with ClAGO5 in vivo, suggesting that ClAGO1 and ClAGO5 co-regulate watermelon defence against CGMMV infection. We also identified the ethylene response factor (ERF) binding site in the promoters of the ClAGO1 and ClAGO5 genes, and ethylene (ETH) treatment significantly increased ClAGO5 expression. Two ERF genes (Cla97C08G147180 and Cla97C06G122830) closely related to ClAGO5 expression were identified using co-expression analysis. Subcellular localization revealed that two ERFs and ClAGO5 predominantly localize at the nucleus, suggesting that enhancement of resistance to CGMMV by ETH is probably achieved through ClAGO5 but not ClAGO1. Our findings reveal aspects of the mechanisms underlying RNA silencing in watermelon against CGMMV.

Editorial for the Special Issue "Benefits of Supplementation with L-arginine, Citrulline and Watermelon on Vascular and Metabolic Health".

The endothelium is crucial in controlling blood pressure and preventing cardiovascular diseases [...].

Effect of microencapsulated watermelon (Citrullus Lanatus) rind on flow-mediated dilation and tissue oxygen saturation of young adults.

BACKGROUND/OBJECTIVES: Watermelon rind (usually discarded by consumers) presents a high L-citrulline content. Given that Lcitrulline is involved in nitric oxide (NO) synthesis, a crucial molecule that regulates vascular function, this study aimed to evaluate the effect of microencapsulated watermelon rind (MWR) on endothelial function and tissue oxygen saturation (StO2) assessed by flow-mediated dilation (FMD) and near-infrared spectroscopy, respectively. Plasma L-arginine and L-citrulline were also evaluated. SUBJECT/METHODS: Eleven participants ingested 30 g of MWR (containing 4 g of L-citrulline) and a placebo. Before and 30, 60, 90, and 120 min after ingestion, StO2 parameters were assessed, whereas FMD and plasma amino acids were analyzed 60 and 120 min after ingestion. RESULTS: The FMD improved 60 min after MWR without changes in StO2 parameters. Absolute plasma L-citrulline and relative change from baseline in plasma L-arginine increased 60 min after MWR ingestion. CONCLUSION: A single dose of microencapsulated watermelon rind containing 4 g of L-citrulline seems adequate to improve FMD response, but not StO2 parameters in healthy adults. (NCT04781595).

Comparative bioinformatics analysis and abiotic stress responses of expansin proteins in Cucurbitaceae members: watermelon and melon.

Watermelon and melon are members of the Cucurbitaceae family including economically significant crops in the world. The expansin protein family, which is one of the members of the cell wall, breaks down the non-covalent bonds between cell wall polysaccharides, causing pressure-dependent cell expansion. Comparative bioinformatics and molecular characterization analysis of the expansin protein family were carried out in the watermelon (Citrullus lanatus) and melon (Cucumis melo) plants in the study. Gene expression levels of expansin family members were analyzed in leaf and root tissues of watermelon and melon under ABA, drought, heat, cold, and salt stress conditions by quantitative real-time PCR analysis. After comprehensive searches, 40 expansin proteins (22 ClaEXPA, 14 ClaEXPLA, and 4 ClaEXPB) in watermelon and 43 expansin proteins (19 CmEXPA, 15 CmEXPLA, 3 CmEXPB, and 6 CmEXPLB) in melon were identified. The greatest orthologous genes were identified with soybean expansin genes for watermelon and melon. However, the latest divergence time between orthologous genes was determined with poplar expansin genes for watermelon and melon expansin genes. ClaEXPA-04, ClaEXPA-09, ClaEXPB-01, ClaEXPB-03, and ClaEXPLA-13 genes in watermelon and CmEXPA-12, CmEXPA-10, and CmEXPLA-01 genes in melon can be involved in tissue development and abiotic stress response of the plant. The current study combining bioinformatics and experimental analysis can provide a detailed characterization of the expansin superfamily which has roles in growth and reaction to the stress of the plant. The study ensures detailed data for future studies examining gene functions including the roles in plant growth and stress conditions.

6-Benzyladenine alleviates NaCl stress in watermelon (Citrullus lanatus) seedlings by improving photosynthesis and upregulating antioxidant defences.

Soil salinity is a growing problem in agriculture, plant growth regulators (PGRs) can regulate plant response to stress. The objective of this study was to evaluate the effects of exogenous 6-benzyladenine (6-BA) on photosynthetic capacity and antioxidant defences in watermelon (Citrullus lanatus L.) seedlings under NaCl stress. Two watermelon genotypes were subjected to four different treatments: (1) normal water (control); (2) 20mgL-1 6-BA; (3) 120mmolL-1 NaCl; and (4) 120mmolL-1 NaCl+20mgL-1 6-BA. Our results showed that NaCl stress inhibited the growth of watermelon seedlings, decreased their photosynthetic capacity, promoted membrane lipid peroxidation, and lowered the activity of protective enzymes. Additionally the salt-tolerant Charleston Gray variety fared better than the salt-sensitive Zhengzi NO.017 variety under NaCl stress. Foliar spraying of 6-BA under NaCl stress significantly increased biomass accumulation, as well as photosynthetic pigment, soluble sugar, and protein content, while decreasing malondialdehyde levels, H2 O2 content, and electrolyte leakage. Moreover, 6-BA enhanced photosynthetic parameters, including net photosynthetic rate, stomatal conductance, intercellular CO2 concentration, and transpiration rate; activated antioxidant enzymes, such as superoxide dismutase, catalase, and peroxidase; and improved the efficiency of the ascorbate-glutathione cycle by stimulating glutathione reductase, dehydroascorbate reductase, and monodehydroascorbate reductase, as well as ascorbic acid and glutathione content. Principal component analysis confirmed that 6-BA improved salt tolerance of the two watermelon varieties, particularly Zhengzi NO.017, albeit through two different regulatory mechanisms. In conclusion, 6-BA treatment could alleviate NaCl stress-induced damage and improve salt tolerance of watermelons by regulating photosynthesis and osmoregulation, activating the ascorbate-glutathione cycle, and promoting antioxidant defences.

Identification of the GAPDH gene family in Citrullus lanatus and functional characteristics of ClGAPC2 in Arabidopsis thaliana.

Members of the GAPDH family play important roles in plant growth and development, as well as in stress responses. Our aim was to identify stress resistance genes through systematic analysis of the GAPDH family in watermelon. This could not only provide genetic resources for stress resistance breeding, but also form a basis for the study of plant stress resistance mechanisms. Eight GAPDHs representing four types of plant GAPDH in watermelon were identified (ClGAPA/B, ClGAPC1-3, ClGAPCp1-2 and ClGAPN). A comprehensive analysis of physicochemical properties, chromosome distribution, evolutionary relationships, exon-intron structure and conserved motifs of watermelon GAPDHs was performed using bioinformatics. Expression characteristics were assessed by RT-qPCR. Based on RT-qPCR results, ClGAPC2 was screened as a candidate for subcellular localization analysis and functional verification in Arabidopsis thaliana. Eight GAPDHs were classified into four subfamilies. GAPDHs in each subgroup were generally conserved and shared similarities in structure and conserved motifs. ClGAPDHs had notable tissue specificity and different expression patterns in response to H2 O2 , chilling, salt, osmotic stress, heat, salicylic acid, gibberellin, brassinosterol, ethylene and abscisic acid treatments. Three ClGAPC genes, especially ClGAPC2, were markedly induced by several treatments. ClGAPC2 was located in the nucleus and cytoplasm of tabacum epidermal cells. The ClGAPC2 transgenic Arabidopsis showed enhanced tolerance to salinity at the germination stage. We suggest that ClGAPC2 plays important roles in the adaptation of watermelon to salinity. Our findings provided candidate genes for further improving the salt tolerance of watermelon.

Watermelon domestication was shaped by stepwise selection and regulation of the metabolome.

Although crop domestication has greatly aided human civilization, the sequential domestication and regulation of most quality traits remain poorly understood. Here, we report the stepwise selection and regulation of major fruit quality traits that occurred during watermelon evolution. The levels of fruit cucurbitacins and flavonoids were negatively selected during speciation, whereas sugar and carotenoid contents were positively selected during domestication. Interestingly, fruit malic acid and citric acid showed the opposite selection trends during the improvement. We identified a novel gene cluster (CGC1, cucurbitacin gene cluster on chromosome 1) containing both regulatory and structural genes involved in cucurbitacin biosynthesis, which revealed a cascade of transcriptional regulation operating mechanisms. In the CGC1, an allele caused a single nucleotide change in ClERF1 binding sites (GCC-box) in the promoter of ClBh1, which resulted in reduced expression of ClBh1 and inhibition of cucurbitacin synthesis in cultivated watermelon. Functional analysis revealed that a rare insertion of 244 amino acids, which arose in C. amarus and became fixed in sweet watermelon, in ClOSC (oxidosqualene cyclase) was critical for the negative selection of cucurbitacins during watermelon evolution. This research provides an important resource for metabolomics-assisted breeding in watermelon and for exploring metabolic pathway regulation mechanisms.

The NF-Y Transcription Factor Family in Watermelon: Re-Characterization, Assembly of ClNF-Y Complexes, Hormone- and Pathogen-Inducible Expression and Putative Functions in Disease Resistance.

Nuclear factor Y (NF-Y) is a heterotrimeric transcription factor that binds to the CCAAT cis-element in the promoters of target genes and plays critical roles in plant growth, development, and stress responses. In the present study, we aimed to re-characterize the ClNF-Y family in watermelon, examine the assembly of ClNF-Y complexes, and explore their possible involvement in disease resistance. A total of 25 ClNF-Y genes (7 ClNF-YAs, 10 ClNF-YBs, and 8 ClNF-YCs) were identified in the watermelon genome. The ClNF-Y family was comprehensively characterized in terms of gene and protein structures, phylogenetic relationships, and evolution events. Different types of cis-elements responsible for plant growth and development, phytohormones, and/or stress responses were identified in the promoters of the ClNF-Y genes. ClNF-YAs and ClNF-YCs were mainly localized in the nucleus, while most of the ClNF-YBs were localized in the cytoplasm of cells. ClNF-YB5, -YB6, -YB7, -YB8, -YB9, and -YB10 interacted with ClNF-YC2, -YC3, -YC4, -YC5, -YC6, -YC7, and -YC8, while ClNF-YB1 and -YB3 interacted with ClNF-YC1. A total of 37 putative ClNF-Y complexes were identified, e.g., ClNF-YA1, -YA2, -YA3, and -YA7 assembled into 13, 8, 8, and 8 ClNF-Y complexes with different ClNF-YB/-YC heterodimers. Most of the ClNF-Y genes responded with distinct expression patterns to defense hormones such as salicylic acid, methyl jasmonate, abscisic acid, and ethylene precursor 1-aminocyclopropane-1-carboxylate, and to infection by the vascular infecting fungus Fusarium oxysporum f. sp. niveum. Overexpression of ClNF-YB1, -YB8, -YB9, ClNF-YC2, and -YC7 in transgenic Arabidopsis resulted in an earlier flowering phenotype. Overexpression of ClNF-YB8 in Arabidopsis led to enhanced resistance while overexpression of ClNF-YA2 and -YC2 resulted in decreased resistance against Botrytis cinerea. Similarly, overexpression of ClNF-YA3, -YB1, and -YC4 strengthened resistance while overexpression of ClNF-YA2 and -YB8 attenuated resistance against Pseudomonas syringae pv. tomato DC3000. The re-characterization of the ClNF-Y family provides a basis from which to investigate the biological functions of ClNF-Y genes in respect of growth, development, and stress response in watermelon, and the identification of the functions of some ClNF-Y genes in disease resistance enables further exploration of the molecular mechanism of ClNF-Ys in the regulation of watermelon immunity against diverse pathogens.

Influence of Enzymatic Hydrolysis and Molecular Weight Fractionation on the Antioxidant and Lipase / α-Amylase Inhibitory Activities In Vitro of Watermelon Seed Protein Hydrolysates.

This study aims to evaluate the potential in vitro antioxidant and anti-obesity activities of watermelon seed protein hydrolysates (WSPH) obtained using different combinations of enzymes alcalase−proteinase K (ALC-PK) and alcalase−actinidin (ALC-ACT). There was a direct relationship between the degree of hydrolysis (DH) and the biological activities of the WSPH, with the highest DPPH (approximately 85%) and lipase inhibitory activities (≈59%) appreciated at DH of 36−37% and 33−35% when using ALC-PK and ALC-ACT, respectively. Following molecular weight fractionation, the ALC-PK WSPH < 3 kDa (F1) assayed at 1 mg.mL−1 had the highest DPPH-radical scavenging (89.22%), ferrous chelating (FC) (79.83%), reducing power (RP) (A 0.51), lipase inhibitory (71.36%), and α-amylase inhibitory (62.08%) activities. The amino acid analysis of ALC-PK WSPH and its fractions revealed a relationship between the biological activity of the extracts and their composition. High contents of hydrophobic amino acids, arginine, and aromatic amino acids were related to high antioxidant, lipase inhibitory, and α-amylase inhibitory activities in the extracts, respectively. Overall, this study revealed that underutilized protein sources such as WSPH, using the appropriate combination of enzymes, could result in the generation of new ingredients and compounds with powerful antioxidant and anti-obesity activities with promising applications as nutraceuticals or functional foods.

Genome-wide comprehensive characterization and expression analysis of TLP gene family revealed its responses to hormonal and abiotic stresses in watermelon (Citrullus lanatus).

Thaumatin-like protein (TLP) is the well-known sweetest protein which plays a crucial role in diverse developmental processes and different stress conditions in plants, fungi and animals. The TLP gene family is extensively studied in different plant species including crop plants. Watermelon (Citrullus lanatus) is an important cucurbit crop cultivated worldwide; however, the comprehensive information about the TLP gene family is not available in watermelon. In the present study, we identified the 29 TLP genes as gene family members in watermelon using various computational methods to understand its role in different developmental processes and stress conditions. ClaTLP gene family members were not uniformly distributed on 22 chromosomes. Phylogenetic analysis revealed that the ClaTLP gene family members were grouped into 10 sub-groups. Further, gene duplication analysis showed thirteen gene duplication events which included one tandem and twelve segmental duplications. Amino acid sequence alignment has shown that ClaTLP proteins shared 16 conserved cysteine residues in their THN domain. Furthermore, cis-acting regulatory elements analysis also displayed that ClaTLP gene family members contain diverse phytohormone, various defense, and stress-responsive elements in their promoter region. The expression profile of the ClaTLP gene family revealed the differential expression of gene family members in different tissues and abiotic stresses conditions. Moreover, the expression profile of ClaTLP genes was further validated by semi-quantitative reverse transcriptase PCR. Taken together, these results indicate that ClaTLP genes might play an important role in developmental processes and diverse stress conditions. Therefore, the outcome of this study brings forth the valuable information for further interpret the precise role of ClaTLP gene family members in watermelon.

Disruption of REC8 in Meiosis I led to watermelon seedless.

In triploid watermelon (Citrullus lanatus), the homologous chromosomes of germ cells are disorder during meiosis, resulting in the failure of seeds formation and producing seedless fruit. Therefore, mutating the genes specifically functioning in meiosis may be an alternative way to achieve seedless watermelon. REC8, as a key component of the cohesin complex in meiosis, is dramatically essential for sister chromatid cohesion and chromosome segregation. However, the role of REC8 in meiosis has not yet been characterized in watermelon. Here, we identified ClREC8 as a member of RAD21/REC8 family with a high expression in male and female flowers of watermelon. In situ hybridization analysis showed that ClREC8 was highly expressed at the early stage of meiosis during pollen formation. Knocking out ClREC8 in watermelon led to decline of pollen vitality. After pollinating with foreign normal pollen, the ovaries of ClREC8 knockout lines could inflate normally but failed to form seeds. We further compared the meiosis chromosomes of pollen mother cells in different stages between the knockout lines and the corresponding wild type. The results indicated that ClREC8 was required for the monopolar orientation of the sister kinetochores in Meiosis I. Additionally, transcriptome sequencing (RNA-seq) analysis between WT and the knockout lines revealed that the disruption of ClREC8 caused the expression levels of mitosis-related genes and meiosis-related genes to decrease. Our results demonstrated ClREC8 has a specific role in Meiosis I of watermelon germ cells, and loss-of-function of the ClREC8 led to seedless fruit, which may provide an alternative strategy to breed cultivars with seedless watermelon.

Induced oxidative equilibrium damage and reduced toxin synthesis in Fusarium oxysporum f. sp. niveum by secondary metabolites from Bacillus velezensis WB.

In this study, the antifungal mechanism of secondary metabolites from the WB strain against Fusarium oxysporum f. sp. niveum (Fon) was investigated. The WB strain induced the accumulation of reactive oxygen species in Fon hyphae and caused morphological changes, including surface subsidence and shrinkage deformation. The cell-free supernatants (CFSs) from WB treatment caused a significant increase in superoxide dismutase, catalase, peroxidase and glutathione reductase activities and the contents of soluble protein and malondialdehyde. Additionally, CFSs from WB decreased the fusaric acid concentration in Fon. Transcriptome analysis revealed that the expression of some antioxidant-related genes was upregulated and that the expression of mycotoxin-related genes was downregulated. Four polypeptide compounds from the WB strain, including iturin A, fengycin, surfactin and bacitracin, were identified by ultra-high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry analysis and complete genome mining. RT-qPCR and a quantitative analysis confirmed that the presence of Fon induced the expression of polypeptide genes and elevated polypeptide production. The combined minimum inhibitory concentration and quantitative analysis of four polypeptides revealed that iturin A, fengycin, surfactin and bacitracin might be responsible for inhibiting the growth of Fon. In conclusion, secondary metabolites from strain WB exhibited antifungal effects on Fon by triggering oxidative stress and decreasing toxin levels.

Metronidazole photocatalytic degradation by zinc oxide nanoparticles synthesized in watermelon peel extract; Advanced optimization, simulation and numerical models using machine learning applications.

Antibiotics in water systems and wastewater are among the greatest major public health problem and it is global environmental issues. Herein a novel approach for the photocatalytic degradation of metronidazole (MTZ) by using eco-green zinc oxide nanoparticles (EG-ZnO NPs) which biosynthesised using watermelon peels extracts has been investigated. Mathematical prediction models using an adaptive neuro-fuzzy inference system (ANFIS), artificial neural networks (ANN) and response surface methodology (RSM) were used to determine the optimal conditions for the degradation process. The FESEM analysis revealed that EG-ZnO NPs was white with a spherical shape and size between 40 and 88 nm. The simulation process for the mathematical prediction model revealed that the best validation performance was 55.35 recorded at epoch 2, the coefficient (R2) was 0.9967 for training data, as detected using ANN analysis. The best operating parameters for MTZ degradation was predicted using RSM to be: 170 mg L-1 of EG-ZnO NPs, 20.61 mg 100 mL-1 of MTZ, 10 min exposure time, and a pH of 5, with 77.48 vs 78.14% corresponding to the predicted and empirically measured respectively. The photocatalytic degradation of MTZ was fitted with pseudo-first-order kinetic (R2 > 0.90). MTZ lost the antimicrobial activity against Bacillus cereus (B. cereus) and Escherichia coli (E. coli) after degradation with EG-ZnO NPs at the optimal conditions as determined in the optimization process. These findings reflect the important role ANFIS and ANN in predicting and optimising the efficacy of engineered nanomaterials, including EG-ZnO NPs, for antibiotic degradation.