Search Results (135)

As an auxin-responsive gene, Gretchen Hagen 3 (GH3) maintains hormonal homeostasis by conjugating excess auxin with amino acids in plant stress-related signaling pathways. GH3 genes have been characterized in many plant species, but the characteristics of pepper (Capsicum annuum L.) GH3 (CaGH3) gene family members in response to multiple stimulants are largely unknown. In this study, we systematically identified the CaGH3 gene family at the genome level and identified eight members on four chromosomes in pepper. CaGH3s were divided into two groups (I and III) and shared conserved motifs, domains, and gene structures. Moreover, CaGH3s had close evolutionary relationships with tomato (Solanum lycopersicum L.), and the promoters of most CaGH3 genes contained hormone and abiotic stress response elements. A protein interaction prediction analysis demonstrated that the CaGH3-3/3-6/3-7/3-8 proteins were possibly core members of the CaGH3 family interaction. In addition, qRT-PCR results showed that CaGH3 genes were differentially expressed in pepper tissues and could be induced by phytohormones (IAA, ABA, and MeJA) and abiotic stresses (salt, low temperature, and drought) with different patterns. In addition, CaGH3-5 and CaGH3-7 were cloned, and the sequences showed a high degree of conservation. Moreover, the results of subcellular localization indicated that they were located in the membrane and chloroplast. Notably, after overexpressing CaGH3-7 in tomato, RNA-seq was performed on wild-type and transgenic lines, and the differentially expressed genes were mainly enriched in response to external stimuli. This study not only lays the foundation for a comprehensive understanding of the function of the CaGH3 gene family during plant growth and stress responses but also provides potential genetic resources for pepper resistance breeding. Full article

Crocin biosynthesis involves a complex network of enzymes with biosynthetic and modifier enzymes, and the manipulation of these pathways holds promise for improving human health through the broad exploitation of these bioactive metabolites. Crocins play a significant role in human nutrition and health, as they exhibit antioxidant and anti-inflammatory activity. Plants that naturally accumulate high levels of crocins are scarce, and the production of crocins is highly limited by the characteristics of the crops and their yield. The CCD2 enzyme, initially identified in saffron, is responsible for converting zeaxanthin into crocetin, which is further modified to crocins by aldehyde dehydrogenases and glucosyltransferase enzymes. Crops like tomato fruits, which naturally contain high levels of carotenoids, offer valuable genetic resources for expanding synthetic biology tools. In an effort to explore CCD2 enzymes with improved activity, two CCD2 alleles from saffron and Crocosmia were introduced into tomato, together with a UGT gene. Furthermore, in order to increase the zeaxanthin pool in the fruit, an RNA interference construct was introduced to limit the conversion of zeaxanthin to violaxanthin. The expression of saffron CCD2, CsCCDD2L, led to the creation of transgenic tomatoes with significantly high crocins levels, reaching concentrations of 4.7 mg/g dry weight. The Crocosmia allele, CroCCD2, also resulted in high crocins levels, reaching a concentration of 2.1 mg/g dry weight. These findings underscore the importance of enzyme variants in synthetic biology, as they enable the development of crops rich in beneficial apocarotenoids. Full article

Chaperonin 60 proteins plays an important role in plant growth and development as well as the response to abiotic stress. As part of the protein homeostasis system, molecular chaperones have attracted increasing attention in recent years due to their involvement in the folding and assembly of key proteins in photosynthesis. However, little is known about the function of maize chaperonin 60 protein. In the study, a gene encoding the chaperonin 60 proteins was cloned from the maize inbred line B73, and named ZmCpn60-3. The gene was 1, 818 bp in length and encoded a protein consisting of 605 amino acids. Phylogenetic analysis showed that ZmCpn60-3 had high similarity with OsCPN60-1, belonging to the β subunits of the chloroplast chaperonin 60 protein family, and it was predicted to be localized in chloroplasts. The ZmCpn60-3 was highly expressed in the stems and tassels of maize, and could be induced by exogenous plant hormones, mycotoxins, and pathogens; Overexpression of ZmCpn60-3 in Arabidopsis improved the resistance to Pst DC3000 by inducing the hypersensitive response and the expression of SA signaling-related genes, and the H₂O₂ and the SA contents of ZmCpn60-3-overexpressing Arabidopsis infected with Pst DC3000 accumulated significantly when compared to the wild-type controls. Experimental data demonstrate that flg22 treatment significantly upregulated transcriptional levels of the PR1 defense gene in ZmCpn60-3-transfected maize protoplasts. Notably, the enhanced resistance phenotype against Pseudomonas syringae pv. tomato DC3000 (Pst DC3000) in ZmCpn60-3-overexpressing transgenic lines was specifically abolished by pretreatment with ABT, a salicylic acid (SA) biosynthetic inhibitor. Our integrated findings reveal that this chaperonin protein orchestrates plant immune responses through a dual mechanism: triggering a reactive oxygen species (ROS) burst while simultaneously activating SA-mediated signaling cascades, thereby synergistically enhancing host disease resistance. Additionally, yeast two-hybrid assay preliminary data indicated that ZmCpn60-3 might bind to ZmbHLH118 and ZmBURP7, indicating ZmCpn60-3 might be involved in plant abiotic responses. The results provided a reference for comprehensively understanding the resistance mechanism of ZmCpn60-3 in plant responses to abiotic or biotic stress. Full article

Tomato bacterial wilt, caused by Ralstonia solanacearum (G⁻), is one of the most devastating plant diseases. Developing effective resistance against this pathogen remains a major challenge in plant disease management. In this study, we constructed a fusion gene BPI-LY by combining the gene encoding the lipophilic functional domains of human bactericidal/permeability-increasing protein (BPI) with the gene of human lysozyme (LY). The recombinant gene BPI-LY was heterologously expressed in yeast and tomato. Preliminary in vitro assays in yeast demonstrated that BPI enhances LY’s antibacterial activity against G⁻ bacteria. Furthermore, overexpression of BPI-LY in tomato delayed onset of the disease in the transgenic lines and lowered the degree of tissue damage and the number of bacteria present in the stems relative to those in the wild-type plant. Additionally, the expression levels of the SlSOD, SlPOD, SlPAL, SlPR5, SlPR10, and SlPR-NP24 genes were indirectly upregulated in the transgenic plants following R. solanacearum inoculation. Collectively, these findings demonstrate that BPI-LY enhances the resistance of transgenic tomato against bacterial wilt caused by R. solanacearum. Full article

Genome editing has revolutionized plant science, providing an unprecedented ability to precisely manipulate plant genomes. For this study, genome editing was utilized to target and modify the NF-YA8 transcription factor (TF) in tomato plants (Solanum lycopersicum L. var. Heinz 1706). The primary objective of this research was to introduce targeted mutations in a non-transgenic manner to the NF-YA8 gene, which encodes the alpha subunit of the Nuclear Factor-Y (NF-Y) heterotrimeric TF, and explore its potential for developing new and improved tomato varieties. Through the transient expression of custom-engineered zinc finger nucleases (ZFNs) in tomato seeds, mutations were successfully introduced in the target gene. The recovered mutant NF-YA8 coding sequences showed a significant level of similarity to the wild type, with a range of 86.9% to 98.21%. Genotyping M2 lines revealed monogenic mutations at or near the intended target site. Phenotypic changes were also evident in both vegetative and reproductive stages of plants. The research revealed that NF-YA8 functions as a high-level regulator, orchestrating a developmental cascade that influences key agronomic traits throughout the plant’s life cycle, including cotyledon development, stem architecture, inflorescence architecture, flowering time, and fruit size and shape. Full article

Lycopene, a natural pigment, is valuable for human health because of its strong antioxidant capacity. However, studies on the involvement of tomato miR394 in the regulation of lycopene have not been reported. The aim of this study was to reveal the molecular mechanism by which miR394 regulates lycopene synthesis by targeting ζ-carotene dehydrogenase (ZDS). The miR394-silenced transgenic tomato plants were constructed by short tandem target mimicry (STTM) technology, and the association between lycopene content and antioxidant capacity was analyzed by combining qRT-PCR, UV spectrophotometry, and a free radical scavenging assay. The targeting relationship between miR394 and ZDS was verified using a subcellular localization assay. The results showed that the silencing of miR394 significantly upregulated the expression of the ZDS gene and promoted lycopene accumulation. The antioxidant enzyme activities of STTM394 transgenic plants were significantly enhanced, and the free radical scavenging ability was obviously improved. Subcellular localization experiments confirmed that miR394 directly inhibited the chloroplast expression of ZDS. In conclusion, this study reveals for the first time that the miR394-ZDS module enhances the antioxidant capacity by regulating lycopene metabolism, which provides a new target for themolecular breeding of highly nutritious tomatoes. Full article

The R2R3-MYB transcription factor GAMYB plays crucial roles in plant growth and development, but the biological functions of SlGAMYB1 in tomato remain poorly understood. Here, we investigated the roles of SlGAMYB1 by overexpressing a miR159-resistant version (35S:SlGAMYB1^m) in tomato. Transgenic plants exhibited a dwarf phenotype with reduced internode elongation, which was associated with decreased bioactive gibberellin (GA) levels due to transcriptional repression of SlGA3ox1 and activation of SlGA2ox1/2/4/5. Additionally, 35S:SlGAMYB1^m altered leaf morphology by inhibiting cell proliferation through downregulation of cell cycle genes, resulting in larger but fewer epidermal cells. Intriguingly, 35S:SlGAMYB1^m plants displayed increased floral organ number, a process likely mediated by the upregulation of SlWUS rather than GA signaling. These findings demonstrate that SlGAMYB1 regulates diverse aspects of tomato development through both GA-dependent and independent pathways, providing new insights into the functional diversification of GAMYB genes and potential strategies for genetic improvement of tomato architecture and yield. Full article

Tomato (Solanum lycopersicum L.) is one of the major vegetable crops worldwide. Research on the Janus kinase–signal transducer and activator of transcription (JAK–STAT) signaling pathway in tomatoes and other plant systems is extremely limited. In this study, the roles of STAT, a crucial element of the JAK–STAT signaling pathway in tomato seed germination and low-temperature stress responses are examined, employing gene family analysis and genetic transformation. The results indicate that the S. lycopersicum genome contains only one member of the STAT gene family, SlSTAT. Subcellular localization experiments reveal that SlSTAT is found in both the cytoplasm and nucleus, suggesting its potential involvement in biological functions within these cellular compartments. Among the 26 different tomato tissue/organs tested, SlSTAT exhibited higher expression levels in hypocotyl (8 days past germination; 8 DPG), and low expression of SlSTAT significantly reduced the germination rate and impacted biomass at 8 DPG. In addition, the SlSTAT gene was significantly downregulated during low-temperature treatment. Compared with the wild-type (WT) tomatoes, the SlSTAT-overexpressing plants showed more resistance to low-temperature conditions, whereas the downexpressing tomatoes exhibited increased sensitivity. The expressions of low-temperature marker genes (SlCBF1-3) and N⁶-methyladenosine (m⁶A)-modification-related genes (m⁶A writer, reader, and eraser genes) were detected to explore possible molecular mechanisms by which SlSTAT causes changes in tomato low-temperature stress resistance. The expression changes of SlCBF1-3 in transgenic plants do not merely follow a straightforward linear relationship with the changes in SlSTAT expression, suggesting a more complex molecular mechanism and a non-direct interaction between SlSTAT and the promoters of SlCBFs. On the other hand, SlSTAT also changes the expression levels of RNA m⁶A-modification-related genes, especially SlFIP37 (writer gene), SlYTP8/9 (reader genes), and SlALKBH8 (eraser gene), ultimately leading to changes in the levels of m⁶A modification. These research findings lay the groundwork for exploring functions of JAK–STAT pathway in tomato development and stress responses, expanding the scope of JAK–STAT signaling studies in plant systems. Full article

Research exploring involvement of RNA N⁶-methyladenosine (m⁶A) in tomato (Solanum lycopersicum) seed germination remains limited. There is also a lack of direct evidence supporting the interaction among tomato seed germination, microgravity, and m⁶A modification. In this study, Micro-Tom tomatoes are used as the experimental material to conduct tomato genetic transformation, seed germination assay, and m⁶A modification levels identification experiments. During tomato seed germination processes, the m⁶A modification level significantly increases under the mutual influence of various m⁶A methyltransferase subunits and multiple eraser proteins. As a m⁶A reader gene, SlYTP9 expression significantly affects the germination of tomato seeds, with promotion and inhibition in OE (overexpression) and RNAi (RNA interference) transgenic tomato plants, respectively. Microgravity promotes seed germination in the early germination period (0–3 days past germination; 0–3 DPG), but this promoting effect gradually disappears as the seedling grows (8–15 DPG). Further exploration revealed that this promoting effect is correlated with m⁶A modification, manifested as enhanced expression of most m⁶A writer genes; increased expression levels of overall reader genes; altered expression trends of some m⁶A eraser genes, particularly SlALKBH2; and enhanced m⁶A modification levels. The experimental results obtained in this study can provide a theoretical basis and evidence support for elucidating the role of m⁶A in tomato seed germination, as well as for exploring the interactions between seed germination, microgravity, and m⁶A modification. Full article

Tomato (Solanum lycopersicum L.) is a major economic vegetable crop globally, yet it is prone to gray mold disease caused by Botrytis cinerea infection during cultivation. Caffeoyl shikimate esterase (CSE) is a crucial component of the lignin biosynthesis pathway, which significantly contributes to plant stress resistance. Therefore, investigating the expression patterns of SlCSE after Botrytis cinerea infection may offer a theoretical foundation for breeding resistant tomato varieties. In this study, 11 SlCSE family members were identified from the tomato genome using bioinformatics analyses. Public transcriptome databases and RT-qPCR experiments were used to analyze gene expression in tomato tissues, responses to Botrytis cinerea infection, and the temporal characteristics of the response to 2-ethylfuran treatment during infection. These experiments resulted in the identification of the key gene SlCSE06. Transgenic tomato lines that overexpressed SlCSE06 were constructed to examine their resistance levels to gray mold disease. Many SlCSE genes were upregulated when tomato fruit were infected with Botrytis cinerea during the ripening stage. Furthermore, 24 h after treatment with 2-ethylfuran, most SlCSE genes exhibited increased expression levels compared with the control group, but they exhibited significantly lower levels at other time points. Thus, 2-ethylfuran treatment may enhance the responsiveness of SlCSEs. Based on this research, SlCSE06 was identified as the key gene involved in the response to Botrytis cinerea infection. The SlCSE06-overexpressing (OE6) tomato plants exhibited a 197.94% increase in expression levels compared to the wild type (WT). Furthermore, the lignin content in OE6 was significantly higher than in WT, suggesting that the overexpression of SlCSE06 enhanced lignin formation in tomato plants. At 5 days post-inoculation with Botrytis cinerea, the lesion diameter in OE6 decreased by 31.88% relative to the WT, whereas the lignin content increased by 370.90%. Furthermore, the expression level of SlCSE06 was significantly upregulated, showing a 17.08-fold increase compared with the WT. These findings suggest that 2-ethylfuran enhances the activation of the critical tomato disease resistance gene SlCSE06 in response to gray mold stress, thereby promoting lignin deposition to mitigate further infection by Botrytis cinerea. Full article

Alternative splicing is a critical post-transcriptional regulatory mechanism in eukaryotes. While infection with Ralstonia solanacearum GMI1000 significantly alters plant alternative splicing patterns, the underlying molecular mechanisms remain unclear. Herein, the effect of the GMI1000 Type III secretion system effectors on alternative splicing in the tomato cultivar Heinz 1706 was investigated. The RNA-seq analysis confirmed genome-wide alternative splicing changes induced by the Type III secretion system in tomato, including 1386 differential alternatively spliced events across 1023 genes, many of which are associated with plant defense. Seven nucleus-localized Type III effectors were transiently expressed in an RLPK splicing reporter system transgenic tobacco, identifying RipP2 as an effector that modulates alternative splicing levels. Sequence analysis, protein–protein interaction assays, and AlphaFold2 structural predictions revealed that RipP2 interacted with the tomato splicing factor SR34a. Furthermore, RipP2 acetylated a conserved lysine at position 132 within the SWQDLKD motif of SR34a, regulating its splicing pattern in defense-related genes and modulating plant immunity. This study elucidates how the “RipP2-SR34a module” influences plant immune responses by regulating the alternative splicing of immune-related genes, providing new insights into pathogen–plant interactions and splicing regulation. Full article

Background:Citrus Exocortis Viroid (CEVd) is a non-coding RNA pathogen capable of infecting a wide range of plant species, despite its lack of protein-coding ability. Viroid infections induce significant alterations in various physiological and biochemical processes, particularly impacting plant metabolism. This study shows the metabolic changes upon viroid infection in tomato plants (Solanum lycopersicum var. ‘MoneyMaker’) exhibiting altered levels of salicylic acid (SA), a key signal molecule involved in the plant defence against this pathogen. Methods: Transgenic RNAi_S5H lines, which have the salicylic acid 5-hydroxylase gene silenced to promote SA accumulation, and NahG lines, which overexpress a salicylate hydroxylase to degrade SA into catechol and prevent its accumulation, were used to establish different SA levels in plants, resulting in varying degrees of resistance to viroid infection. The analysis was performed by using gas chromatography–mass spectrometry (GC-MS) to explore the role of volatile organic compounds (VOCs) in plant immunity against this pathogen. Results: Our results revealed distinct volatile profiles associated with plant immunity, where RNAi_S5H-resistant plants showed significantly enhanced production of monoterpenoids upon viroid infection. Moreover, viroid-susceptible NahG plants emitted a broad range of VOCs, whilst viroid-tolerant RNAi_S5H plants exhibited less variation in VOC emission. Conclusions: This study demonstrates that SA levels significantly influence metabolic responses and immunity in tomato plants infected by CEVd. The identification of differential emitted VOCs upon CEVd infection could allow the development of biomarkers for disease or strategies for disease control. Full article

Heat stress is one of the most important environmental problems in agriculture, which severely restricts the growth and yield of plants. In plants, microRNA398 (miR398) negatively regulates the activity of superoxide dismutase (SOD) by modulating the expression of its coding genes (CSDs) post-transcriptionally, thereby regulating reactive oxygen species (ROS) homeostasis and stress resistance. In this study, the role of miR398 in heat stress tolerance in tomato was investigated. Under heat stress, the expression of miR398 was upregulated in tomato, while the expression of its target genes (CSD1 and CSD2) and SOD activity was downregulated. Furthermore, by comparing the heat stress response in wild type (WT) and a transgenic line overexpressing MIR398 (miR398-OE), the results showed that overexpression of miR398 promoted tomato growth and the expression of genes encoding heat shock factor (HSF, transcription factor) and heat shock protein (HSP) under heat stress. Meanwhile, downregulated activity of antioxidant enzymes, including SOD, catalase (CAT), peroxidase (POD), and ascorbate peroxidase (APX), and enhanced ROS accumulation was observed in miR398-OE compared with that in WT under heat stress. Further study using dimethylthiourea (DMTU, a ROS scavenger) indicated that the enhanced plant growth and expression of HSFs/HSPs was based on the promoted accumulation of ROS in miR398-OE. Overall, the results of this study revealed that the upregulated expression of miR398 in response to heat stress would modulate the antioxidant system and enhance ROS accumulation, thereby enhancing the expression of HSFs and HSPs and heat stress tolerance in tomato. Full article

Raspberry is a berry whose fruit is not tolerant to storage; breeding varieties with extended storage time and high comprehensive quality are significant for raspberries in cold regions. 1-Aminocyclopropane-1-carboxylic acid (ACC) synthase (ACS) is a limiting enzyme in the ethylene synthesis process, which plays essential roles in fruit ripening and softening in plants. In this study, the RiACS1 gene in raspberry (Rubus idaeus L.) variety ‘Polka’ was cloned. The RiACS1 gene overexpression vector was constructed and transformed into tomato plants using the Agrobacterium tumefaciens infection method to verify its function in their reproductive development. The RiACS1 gene, with a total length of 1476 bp, encoded a protein with 491 amino acids. The subcellular localization analysis of the RiACS1 protein in the tobacco transient expression system revealed that the RiACS1-GFP fusion protein was mainly located in the nucleus. Compared with the control, the flowering time and fruit color turning time of transgenic strains were advanced, and the fruit hardness was reduced. Overexpression of RiACS1 increased the activity of ACC synthase, ethylene release rate, and respiration rate during the transchromic phase. It changed the substance content, increased the content of vitamin C and anthocyanin in the fruit ripening process, and decreased the content of chlorophyll and titrable acid at the maturity stage. In addition, RiACS1 increased the relative expression levels of ethylene synthesis-related genes such as SlACS4, SlACO3, and SlACO1 in the fruit ripening process, while it decreased the expression levels of SlACS2 at the maturity stage. These results suggested that the RiACS1 gene could promote early flowering and fruit ripening in tomato plants. This study provided a basis for further modifying raspberry varieties using molecular biology techniques. Full article

Plants have large amounts of the late embryogenesis abundant protein (LEA) family of proteins, which is involved in osmotic regulation. The Korla Pear (Pyrus sinkiangensis Yu) is an uncommon pear species that thrives in Xinjiang and can survive below-freezing conditions. We found that the PsLEA4 gene was more expressed after cold treatment by looking at the transcriptome data of the Korla Pear. In order to evaluate the biological function of the PsLEA4 protein under low-temperature stress and its potential for use in agricultural breeding, we cloned the PsLEA4 gene from the Korla Pear, made a plant overexpression vector, and transformed it into a tomato via Agrobacterium transformation. When exposed to low temperatures, we found that PsLEA4 overexpression can regulate proline metabolism and antioxidant enzyme activity in tomatoes compared to wild tomatoes. Because of this, transgenic tomatoes are more resilient to cold temperatures and produce more than their wild counterparts. Thus, expressing PsLEA4 has multiple advantages: (1) Improving frost resistance and reducing plant damage. (2) Increasing crop yield. Therefore, this study provides a theoretical basis for the role of the PsLEA4 protein in plants’ resilience to low temperatures, as well as for its potential application in crop breeding. Full article