Down-regulation of SlGT-26 gene confers dwarf plants and enhances drought and salt stress resistance in tomato.

Plant architecture, an important agronomic trait closely associated with yield, is governed by a highly intricate molecular network. Despite extensive research, many mysteries surrounding this regulation remain unresolved. Trihelix transcription factor family plays a crucial role in the development of plant morphology and abiotic stresses. Here, we identified a novel trihelix transcription factor named SlGT-26, and its down-regulation led to significant alterations in plant architecture, including dwarfing, reduced internode length, smaller leaves, and shorter petioles. The dwarf phenotype of SlGT-26 silenced transgenic plants could be recovered after spraying exogenous GA3, and the GA3 content were decreased in the RNAi plants. Additionally, the expression levels of gibberellin-related genes were affected in the RNAi lines. These results indicate that the dwarf of SlGT-26-RNAi plants may be a kind of GA3-sensitive dwarf. SlGT-26 was response to drought and salt stress treatments. SlGT-26-RNAi transgenic plants demonstrated significantly enhanced drought resistance and salt tolerance in comparison to their wild-type tomato counterparts. SlGT-26-RNAi transgenic plants grew better, had higher relative water content and lower MDA and H2O2 contents. The expression of multiple stress-related genes was also up-regulated. In summary, we have discovered a novel gene, SlGT-26, which plays a crucial role in regulating plant architecture and in respond to drought and salt stress.

Trihelix transcription factor SlGT31 regulates fruit ripening mediated by ethylene in tomato.

Trihelix proteins are plant-specific transcription factors that are classified as GT factors due to their binding specificity for GT elements, and they play crucial roles in development and stress responses. However, their involvement in fruit ripening and transcriptional regulatory mechanisms remains largely unclear. In this study, we cloned SlGT31, encoding a trihelix protein in tomato (Solanum lycopersicum), and determined that its relative expression was significantly induced by the application of exogenous ethylene whereas it was repressed by the ethylene-inhibitor 1-methylcyclopropene. Suppression of SlGT31 expression resulted in delayed fruit ripening, decreased accumulation of total carotenoids, and reduced ethylene content, together with inhibition of expression of genes related to ethylene and fruit ripening. Conversely, SlGT31-overexpression lines showed opposite results. Yeast one-hybrid and dual-luciferase assays indicated that SlGT31 can bind to the promoters of two key ethylene-biosynthesis genes, ACO1 and ACS4. Taken together, our results indicate that SlGT31 might act as a positive modulator during fruit ripening.

A tomato HD-zip I transcription factor, VAHOX1, acts as a negative regulator of fruit ripening.

Homeodomain-leucine zipper (HD-Zip) transcription factors are only present in higher plants and are involved in plant development and stress responses. However, our understanding of their participation in the fruit ripening of economical plants, such as tomato (Solanum lycopersicum), remains largely unclear. Here, we report that VAHOX1, a member of the tomato HD-Zip I subfamily, was expressed in all tissues, was highly expressed in breaker+4 fruits, and could be induced by ethylene. RNAi repression of VAHOX1 (VAHOX1-RNAi) resulted in accelerated fruit ripening, enhanced sensitivity to ethylene, and increased total carotenoid content and ethylene production. Conversely, VAHOX1 overexpression (VAHOX1-OE) in tomato had the opposite effect. RNA-Seq results showed that altering VAHOX1 expression affected the transcript accumulation of a series of genes involved in ethylene biosynthesis and signal transduction and cell wall modification. Additionally, a dual-luciferase reporter assay, histochemical analysis of GUS activity and a yeast one-hybrid (Y1H) assay revealed that VAHOX1 could activate the expression of AP2a. Our findings may expand our knowledge about the physiological functions of HD-Zip transcription factors in tomato and highlight the diversities of transcriptional regulation during the fruit ripening process.

The Promising Nanovectors for Gene Delivery in Plant Genome Engineering.

Highly efficient gene delivery systems are essential for genetic engineering in plants. Traditional delivery methods have been widely used, such as Agrobacterium-mediated transformation, polyethylene glycol (PEG)-mediated delivery, biolistic particle bombardment, and viral transfection. However, genotype dependence and other drawbacks of these techniques limit the application of genetic engineering, particularly genome editing in many crop plants. There is a great need to develop newer gene delivery vectors or methods. Recently, nanomaterials such as mesoporous silica particles (MSNs), AuNPs, carbon nanotubes (CNTs), and layer double hydroxides (LDHs), have emerged as promising vectors for the delivery of genome engineering tools (DNA, RNA, proteins, and RNPs) to plants in a species-independent manner with high efficiency. Some exciting results have been reported, such as the successful delivery of cargo genes into plants and the generation of genome stable transgenic cotton and maize plants, which have provided some new routines for genome engineering in plants. Thus, in this review, we summarized recent progress in the utilization of nanomaterials for plant genetic transformation and discussed the advantages and limitations of different methods. Furthermore, we emphasized the advantages and potential broad applications of nanomaterials in plant genome editing, which provides guidance for future applications of nanomaterials in plant genetic engineering and crop breeding.

SlMBP22 overexpression in tomato affects flower morphology and fruit development.

MADS-domain transcription factors have been identified as key regulators involved in proper flower and fruit development in angiosperms. As members of the MADS-box subfamily, Bsister (Bs) genes have been observed to play an important role during the evolution of the reproductive organs in seed plants. However, their effects on reproductive development in fruit crops, such as tomato (Solanum lycopersicum), remain unclear. Here, we found that SlMBP22 overexpression (SlMBP22-OE) resulted in considerable alterations in floral morphology and affected the expression levels of several floral homeotic genes. Further analysis by yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) assays demonstrated that SlMBP22 forms dimers with class A protein MACROCALYX (MC) and SEPALLATA (SEP) floral homeotic proteins TM5 and TM29, respectively. In addition, pollen viability and cross-fertilization assays suggested that the defect in female reproductive development was responsible for the infertility phenotype observed in the strong overexpression transgenic plants. Transgenic fruits with mild overexpression exhibited reduced size as a result of reduced cell expansion, rather than impaired cell division. Additionally, SlMBP22 overexpression in tomato not only affected proanthocyanidin (PA) accumulation but also altered seed dormancy. Taken together, these findings may provide new insights into the knowledge of Bs MADS-box genes in flower and fruit development in tomato.

Overexpression of SlMBP22 in Tomato Affects Plant Growth and Enhances Tolerance to Drought Stress.

MADS-box transcription factors play crucial and diverse roles in plant growth and development, and the responses to biotic and abiotic stresses. However, the implementation of MADS-box transcription factors in regulating plant architecture and stress responses has not been fully explored in tomato. Here, we found that a novel MADS-box transcription factor, SlMBP22, participated in the control of agronomical traits, tolerance to abiotic stress, and regulation of auxin and gibberellin signalling. Transgenic plants overexpressing SlMBP22 (SlMBP22-OE) displayed pleiotropic phenotypes, including reduced plant height and leaf size, by affecting auxin and/or gibberellin signalling. SlMBP22 was induced by dehydration treatment, and SlMBP22-OE plants were more tolerant to drought stress than wild-type (WT). Furthermore, SlMBP22 overexpression plants accumulated more chlorophyll, starch and soluble sugar than WT, indicating that the darker green leaves might be attributed to increased chlorophyll levels in the transgenic plants. RNA-Seq results showed that the transcript levels of a series of genes related to chloroplast development, chlorophyll metabolism, starch and sucrose metabolism, hormone signalling, and stress responses were altered. Collectively, our data demonstrate that SlMBP22 plays an important role in both regulating tomato growth and resisting drought stress.

Overexpression of SlOFP20 in Tomato Affects Plant Growth, Chlorophyll Accumulation, and Leaf Senescence.

Previous studies have shown that OVATE family proteins (OFPs) participate in various aspects of plant growth and development. How OFPs affect leaf chlorophyll accumulation and leaf senescence has not been reported yet. Here, we found that overexpression of SlOFP20 in tomato not only impacted plant architecture but also enhanced the leaf chlorophyll accumulation and retarded leaf senescence. Gene expression analysis of SlGLK1, SlGLK2, and HY5, encoding transcription factors that are putatively involved in chloroplast development and chlorophyll levels, were significantly up-regulated in SlOFP20-OE lines. Both chlorophyll biosynthesis and degradation genes were distinctly regulated in transgenic plants. Moreover, SlOFP20-OE plants accumulated more starch and soluble sugar than wild-type plants, indicating that an increased chlorophyll content conferred some higher photosynthetic performance in SlOFP20-OE plants. Furthermore, The levels of leaf senescence-related indexes, such as hydrogen peroxide, malondialdehyde, and antioxidant enzymes activities, were differently altered, too. SlOFP20 overexpression repressed the expression of senescence-related genes, SAG12, RAV1, and WRKY53. Moreover, abscisic acid and ethylene synthesis genes were down-regulated in transgenic lines. These results provide new insights into how SlOFP20 regulates chlorophyll accumulation and leaf senescence.

Overexpression of SlOFP20 affects floral organ and pollen development.

The OVATE gene was initially identified in tomato and serves as a key regulator of fruit shape. There are 31 OFP members in the tomato genome. However, their roles in tomato growth and reproductive development are largely unknown. Here, we cloned the OFP transcription factor SlOFP20. Tomato plants overexpressing SlOFP20 displayed several phenotypic defects, including an altered floral architecture and fruit shape and reduced male fertility. SlOFP20 overexpression altered the expression levels of some brassinosteroid (BR)-associated genes, implying that SlOFP20 may play a negative role in the BR response, similar to its ortholog OsOFP19 in rice. Moreover, the transcript accumulation of gibberellin (GA)-related genes was significantly affected in the transgenic lines. SlOFP20 may play an important role in the crosstalk between BR and GA. The pollen germination assay suggested that the pollen germination rate of SlOFP20-OE plants was distinctly lower than that of WT plants. In addition, the tomato pollen-associated genes SlCRK1, SlPMEI, LePRK3, SlPRALF, and LAT52 were all suppressed in the transgenic lines. Our data imply that SlOFP20 may affect floral organ and pollen development by modulating BR and GA signaling in tomato.

The tomato MADS-box gene SlMBP9 negatively regulates lateral root formation and apical dominance by reducing auxin biosynthesis and transport.

KEY MESSAGE: Overexpression of SlMBP9 reduced auxin biosynthesis and transport, and negatively regulated lateral root formation and apical dominance. MADS-box transcription factors play a critical role in plant development. In this study, we describe SlMBP9, a novel MADS-box gene that is expressed in the roots of tomato plants. Tomato lines that over- or under-expressed SlMBP9 were generated using a transgenic approach. The number of lateral roots (LRs) were reduced in SlMBP9-overexpressing lines but slightly increased in SlMBP9-silenced lines. A physiological index revealed that the auxin content significantly decreased in the root maturation zone of the overexpression lines. In addition, gene expression analysis revealed that the expression of the polar auxin transporter genes PIN1 and ABCB19/MDR1 and genes involved in auxin biosynthesis was downregulated in the stems of overexpression lines, which is consistent with the reduced accumulation of auxin in the root maturation zone. Exogenous indole-3-acetic acid (auximone) rescued the lateral root phenotypes of the SlMBP9-overexpressing lines. Overexpression of SlMBP9 resulted in dwarf plants, enhanced lateral buds and reduced the gibberellin content in the stems. Together, these results suggest that SlMBP9 plays a negative role in the process of auxin biosynthesis and transport.

Manipulation of plant architecture and flowering time by down-regulation of the GRAS transcription factor SlGRAS26 in Solanum lycopersicum.

Previous studies suggest that GRAS transcription factors act as essential regulators, not only in plant growth and development but also in response to biotic and abiotic stresses. Recently, 53 GRAS proteins have been identified, but only a few of them have been functionally studied in tomato. Here, we isolated a novel GRAS transcription factor SlGRAS26, its down-regulation generated pleiotropic phenotypes, including reduced plant height with more lateral shoots, internode length, leaf size, even leaflets, accelerated flowering transition and decreased trichome number. Transcription analysis showed that down-regulation of SlGRAS26 altered vegetative growth by suppressing gibberellin (GA) biosynthesis genes and activating the GA inactivating genes, thereby reducing endogenous GA content in transgenic plants. SlGRAS26 may regulate the initiation of lateral buds by regulating the expression of Blind (BL) and BRC1b. The earlier initiation of flower buds in transgenic lines may be controlled by significant up-regulation of SFT, CO1, SBP3, SBP13, and SBP15 genes, related to flowering time. These results demonstrate that SlGRAS26 may play a vital role in the initiation of lateral and inflorescence meristems in tomato.

The Jasmonate ZIM-domain protein gene SlJAZ2 regulates plant morphology and accelerates flower initiation in Solanum lycopersicum plants.

JAZ (Jasmonate ZIM-domain) proteins are important repressors in JA signaling pathway. JAZs were proved taking part in various development processes and resistance to biotic and abiotic stresses in Arabiodopsis. However, in tomato, the functional study of JAZs is rare, especially on plant growth and development. Here, a typical tomato JAZ gene, SlJAZ2 was isolated. Tomato plants overexpressing SlJAZ2 exhibited quicker leaf initiation, reduced plant height and internode length, decreasing trichomes, earlier lateral bud emergence and advanced flowering transition. Further experiments showed that the pith cells in transgenic plant stem were much smaller than wild-type and the genes related to cell elongation and gibberellin biosynthesis were down-regulated. Genes mediating trichome formation were also inhibited in plant stem epidermis. In addition, the flower initiation of transgenic plants were earlier and genes controlling flowering time were up-regulated significantly after SlJAZ2 was overexpressed. Our research demonstrates that SlJAZ2 accelerates the transition from vegetative growth to reproductive growth.

Simultaneous functions of the installed DAS/DAK formaldehyde-assimilation pathway and the original formaldehyde metabolic pathways enhance the ability of transgenic geranium to purify gaseous formaldehyde polluted environment.

The overexpression of dihydroxyacetone synthase (DAS) and dihydroxyacetone kinase (DAK) from methylotrophic yeasts in chloroplasts created a photosynthetic formaldehyde (HCHO)-assimilation pathway (DAS/DAK pathway) in transgenic tobacco. Geranium has abilities to absorb and metabolize HCHO. Results of this study showed that the installed DAS/DAK pathway functioning in chloroplasts greatly enhanced the role of the Calvin cycle in transgenic geranium under high concentrations of gaseous HCHO stress. Consequently, the yield of sugars from HCHO-assimilation increased approximately 6-fold in transgenic geranium leaves, and concomitantly, the role of three original HCHO metabolic pathways reduced, leading to a significant decrease in formic acid, citrate and glycine production from HCHO metabolism. Although the role of three metabolic pathways reduced in transgenic plants under high concentrations of gaseous HCHO stress, the installed DAS/DAK pathway could still function together with the original HCHO metabolic pathways. Consequently, the gaseous HCHO-resistance of transgenic plants was significantly improved, and the generation of H2O2 in the transgenic geranium leaves was significantly less than that in the wild type (WT) leaves. Under environmental-polluted gaseous HCHO stress for a long duration, the stomata conductance of transgenic plants remained approximately 2-fold higher than that of the WT, thereby increasing its ability to purify gaseous HCHO polluted environment.

Investigation of the role of the calvin cycle and C1 metabolism during HCHO metabolism in gaseous HCHO-treated petunia under light and dark conditions using 13C-NMR.

INTRODUCTION: It has been shown that formaldehyde (HCHO) absorbed by plants can be assimilated through the Calvin cycle or C1 metabolism. Our previous study indicated that Petunia hybrida could effectively eliminate HCHO from HCHO-polluted air. OBJECTIVE: To understand the roles of C1 metabolism and the Calvin cycle during HCHO metabolism and detoxification in petunia plants treated with gaseous H(13)CHO under light and dark conditions. METHODS: Aseptically grown petunia plants were treated with gaseous H(13)CHO under dark and light conditions. The metabolites generated from HCHO detoxification in petunia were investigated using (13)C-NMR. RESULTS: [2-(13)C]glycine (Gly) was generated via C1 metabolism and [U-(13)C]glucose (Gluc) was produced through the Calvin cycle simultaneously in petunia treated with low-level gaseous H(13)CHO under light conditions. Generation of [2-(13)C]Gly decreased whereas [U-(13) C]Gluc and [U-(13)C]fructose (Fruc) production increased greatly under high-level gaseous H(13)CHO stress in the light. In contrast, [U-(13)C]Gluc and [U-(13)C] Fruc production decreased greatly and [2-(13)C]Gly generation increased significantly under low-level and high-level gaseous H(13)CHO stress in the dark. CONCLUSION: C1 metabolism and the Calvin cycle contributed differently to HCHO metabolism and detoxification in gaseous H(13CHO-treated petunia plants. As the level of gaseous HCHO increased, the role of C1 metabolism decreased and the role of the Calvin cycle increased under light conditions. However, opposite changes were observed in petunia plants under dark conditions.

C1 metabolism plays an important role during formaldehyde metabolism and detoxification in petunia under liquid HCHO stress.

Petunia hybrida is a model ornamental plant grown worldwide. To understand the HCHO-uptake efficiency and metabolic mechanism of petunia, the aseptic petunia plants were treated in HCHO solutions. An analysis of HCHO-uptake showed that petunia plants effectively removed HCHO from 2, 4 and 6 mM HCHO solutions. The (13)C NMR analyses indicated that H(13)CHO was primarily used to synthesize [5-(13)C]methionine (Met) via C1 metabolism in petunia plants treated with 2 mM H(13)CHO. Pretreatment with cyclosporin A (CSA) or l-carnitine (LC), the inhibitors of mitochondrial permeability transition pores, did not affect the synthesis of [5-(13)C]Met in petunia plants under 2 mM H(13)CHO stress, indicating that the Met-generated pathway may function in the cytoplasm. Under 4 or 6 mM liquid H(13)CHO stress, H(13)CHO metabolism in petunia plants produced considerable amount of H(13)COOH and [2-(13)C]glycine (Gly) through C1 metabolism and a small amount of [U-(13)C]Gluc via the Calvin Cycle. Pretreatment with CSA or LC significantly inhibited the production of [2-(13)C]Gly in 6 mM H(13)CHO-treated petunia plants, which suggests that chloroplasts and peroxisomes might be involved in the generation of [2-(13)C]Gly. These results revealed that the C1 metabolism played an important role, whereas the Calvin Cycle had only a small contribution during HCHO metabolism and detoxification in petunia under liquid HCHO stress.