Tomato Mutant Research Articles

The variegated canalized-1 tomato mutant is linked to photosystem assembly

The recently described canal-1 tomato mutant, which has a variegated leaf phenotype, has been shown to affect canalization of yield. The corresponding protein is orthologous to AtSCO2 -SNOWY COTYLEDON 2, which has suggested roles in thylakoid biogenesis. Here we characterize the canal-1 mutant through a multi-omics approach, by comparing mutant to wild-type tissues. While white canal-1 leaves are devoid of chlorophyll, green leaves of the mutant appear wild-type-like, despite an impaired protein function. Transcriptomic data suggest that green mutant leaves compensate for this impaired protein function by upregulation of transcription of photosystem assembly and photosystem component genes, thereby allowing adequate photosystem establishment, which is reflected in their wild-type-like proteome. White canal-1 leaves, however, likely fail to reach a certain threshold enabling this overcompensation, and plastids get trapped in an undeveloped state, while additionally suffering from high light stress, indicated by the overexpression of ELIP homolog genes. The metabolic profile of white and to a lesser degree also green tissues revealed upregulation of amino acid levels, that was at least partially mediated by transcriptional and proteomic upregulation. These combined changes are indicative of a stress response and suggest that white tissues behave as carbon sinks. In summary, our work demonstrates the relevance of the SCO2 protein in both photosystem assembly and as a consequence in the canalization of yield.

Tomato SlARF5 participate in the flower organ initiation process and control plant height

Plant height is a critical agronomic trait closely linked to yield, primarily regulated by Gibberellins (GA) and auxins, which interact in complex ways. However, the mechanism underlying their interactions remain incompletely understood. In this study, we identified a tomato mutant exhibiting significantly reduced plant height. Through gene cloning and bulked segregant analysis (BSA) sequencing, we found that the mutant gene corresponds to the tomato auxin response factor gene SlARF5/MP. Here, we show that overexpression of SlARF5/MP significantly enhances plant height. Additionally, treatment with GA3 restored the plant height of the mutant to wild-type (WT) levels, indicating that GA content is a key factor influencing plant height. We also observed significant upregulation of GA-biosynthesis genes, including GA2-oxidases GA20ox3 and GA20ox4, as well as the GA3 biosynthesis gene GA3ox1, in SlARF5-overexpressing plants. Furthermore, we demonstrated that SlARF5 directly binds to SlGA2ox3, which mediates the conversion of GA3 to inactive GA, therebyregulating its expression. Our findings suggest that SlARF5 modulates GA3 metabolism by regulating GA synthesis genes, ultimately leading to alterations in plant height.

Characterization of Low-Temperature Sensitivity and Chlorophyll Fluorescence in Yellow Leaf Mutants of Tomato

Leaf color mutants serve as valuable models for studying the regulation of plant photosynthesis, alternations in chloroplast structure and function, and the analysis of associated gene functions. A yellow leaf mutant, ylm, was separated from the wild tomato M82, with its yellowing intensity influenced by low temperature. To assess the low-temperature sensitivity of this mutant, the photosynthetic and chlorophyll fluorescence responses of ylm and M82 were examined under different temperature conditions. In this study, the ylm mutant and its wild type, M82, were exposed to three temperature levels, 16, 25, and 30 °C, for 48 h. The impact of these temperature treatments on leaf color change, chlorophyll content, photosynthetic performance, and chlorophyll fluorescence characteristics of mutant ylm was investigated. The results revealed the following: (1) After exposure to 16 °C, the ylm mutant exhibited significant yellowing, a marked reduction in chlorophyll content, and a notable increase in carotenoid content. At 25 °C, the differences were less pronounced, and at 30 °C, the differences between ylm and M82 were minimal. (2) The photosynthetic rate of the ylm mutant was lower than that of M82 at 16 °C, with the gap narrowing as temperature increased, eventually converging at higher temperatures. (3) The fluorescence transient curve (OJIP) of the ylm mutant differed significantly from that of M82 at 16 °C, with higher fluorescence intensity at the O point and lower intensity at the J, I, and P points. This difference was decreased at 25 °C and nearly disappeared at 30 °C. Additionally, the Fv/Fm, Fv/Fo, PIabs, PItotal, ABS/CSm, TRo/CSm, and ETo/CSm values of ylm were lower than those of M82 at 16 °C, while the ABS/RC and DIo/RC values were higher, with no significant differences observed at 30 °C. These findings suggest that the ylm mutant is highly sensitive to low temperature, with pronounced yellowing, reduced light energy absorption and capture efficiency, and impaired electron transport at lower temperature.

Microbial necromass and glycoproteins for determining soil carbon formation under arbuscular mycorrhiza symbiosis

Arbuscular mycorrhizal fungi (AMF) form symbioses with most terrestrial plants and critically modulate soil organic carbon (C) dynamics. Whether AMF promote soil C storage and stability is, however, largely unknown. Since microbial necromass C (MNC) and glomalin-related soil protein (GRSP) are stable microbial-derived C in soils, we therefore evaluated how AMF symbiosis alters both soil C pools and their contributions to soil organic C (SOC) under nitrogen fertilization, based on a 16-weeks mesocosm experiment using a mutant tomato with highly reduced AMF symbiosis. Results showed that SOC content is 4.5 % higher following AMF symbiosis. Additionally, the content of MNC and total GRSP were 47.5 % and 22.3 % higher under AMF symbiosis than at AMF absence, respectively. The accumulations of GRSP and microbial necromass in soil were closely associated with mineral-associated organic C and the abundance of AMF. The increased soil living microbial biomass under AMF symbiosis was mainly derived from AMF biomass, and fungal necromass C significantly contributed to SOC accumulation, as evidenced by the higher fungal:bacterial necromass C ratio under AMF symbiosis. On the contrary, bacterial necromass was degraded to compensate for the increased microbial nutrient demand because of the aggravated nutrient limitation under AMF symbiosis, leading to a decrease in bacterial necromass. Redundancy analysis showing that bacterial necromass was negatively correlated with soil C:N ratio supported this argument. Moreover, the relative change rate of total GRSP was consistently greater in nitrogen-limited soil than that of microbial necromass. Our findings suggested GRSP accumulates faster and contributes more to SOC pools under AMF symbiosis than microbial necromass. The positive correlation between the contributions of GRSP and MNC to SOC further provided valuable information in terms of enhancing our understanding of mechanisms underlying the maintenance of SOC stocks through microbial-derived C.

Crossing a CRISPR/Cas9 transgenic tomato plant with a wild-type plant yields diverse mutations in the F1 progeny.

Generating CRISPR/Cas9-mediated mutants in tomato (Solanum lycopersicum L.) involves screening shoots regenerated from cultured cells transformed with a T-DNA harboring sequences encoding Cas9 and single guide RNAs (sgRNAs). Production of transformants can be inconsistent and obtaining transformants in large numbers may be difficult, resulting in a limited variety of mutations. Here, I report a method for generating various types of mutations from one transgenic plant harboring the CRISPR/Cas9 system. In this method, a wild-type plant was crossed with a T0 biallelic mutant expressing two sgRNAs targeting the RIPENING INHIBITOR (RIN) gene, and the resulting F1 seedlings were classified using a kanamycin resistance marker on the T-DNA. Genotyping of the RIN locus revealed that kanamycin-sensitive F1 seedlings, which carried no T-DNA, always harbored the wild-type allele and a mutant allele from the transgenic parent. Kanamycin-resistant F1 seedlings, which do carry the T-DNA, harbored a variety of novel mutant alleles, but not the wild-type allele, suggesting that it was mutated during crossing. The novel mutations included one-base insertions or short deletions at each target site, or large deletions across the two target sites. This method was also successfully applied to produce mutations in Geranylgeranyl pyrophosphate synthase 2 (GGPS2). Because this method involves crossing rather than transformation, it can be readily scaled up to produce numerous novel mutations, even in plant species or cultivars for which transformation is inefficient. Therefore, when initial transgene experiments fail to induce the desired mutation, this method provides additional opportunities for generating mutants.

Physiological characterization of the tomato cutin mutant cd1 under salinity and nitrogen stress.

We identified tomato leaf cuticle and root suberin monomers that play a role in the response to nitrogen deficiency and salinity stress and discuss their potential agronomic value for breeding. The plant cuticle plays a key role in plant-water relations, and cuticle's agronomic value in plant breeding programs is currently under investigation. In this study, the tomato cutin mutant cd1, with altered fruit cuticle, was physiologically characterized under two nitrogen treatments and three salinity levels. We evaluated leaf wax and cutin load and composition, root suberin, stomatal conductance, photosynthetic rate, partial factor productivity from applied N, flower and fruit number, fruit size and cuticular transpiration, and shoot and root biomass. Both nitrogen and salinity treatments altered leaf cuticle and root suberin composition, regardless of genotype (cd1 or M82). Compared with M82, the cd1 mutant showed lower shoot biomass and reduced partial factor productivity from applied N under all treatments. Under N depletion, cd1 showed altered leaf wax composition, but was comparable to the WT under sufficient N. Under salt treatment, cd1 showed an increase in leaf wax and cutin monomers. Root suberin content of cd1 was lower than M82 under control conditions but comparable under higher salinity levels. The tomato mutant cd1 had a higher fruit cuticular transpiration rate, and lower fruit surface area compared to M82. These results show that the cd1 mutation has complex effects on plant physiology, and growth and development beyond cutin deficiency, and offer new insights on the potential agronomic value of leaf cuticle and root suberin for tomato breeding.

Mutation of YFT3, an isomerase in the isoprenoid biosynthetic pathway, impairs its catalytic activity and carotenoid accumulation in tomato fruit.

Tomato fruit colors are directly associated with their appearance quality and nutritional value. However, tomato fruit color formation is an intricate biological process that remains elusive. In this work we characterized a tomato yellow fruited tomato 3 (yft3, e9292, Solanum lycopersicum) mutant with yellow fruits. By the map-based cloning approach, we identified a transversion mutation (A2117C) in the YFT3 gene encoding a putative isopentenyl diphosphate isomerase (SlIDI1) enzyme, which may function in the isoprenoid biosynthetic pathway by catalyzing conversion between isopentenyl pyrophosphate (IPP) and dimethylallyl pyrophosphate (DMAPP). The mutated YFT3 (A2117C) (designated YFT3 allele) and the YFT3 genes did not show expression difference at protein level, and their encoded YFT3 allelic (S126R) and YFT3 proteins were both localized in plastids. However, the transcript levels of eight genes (DXR, DXS, HDR, PSY1, CRTISO, CYCB, CYP97A, and NCED) associated with carotenoid synthesis were upregulated in fruits of both yft3 and YFT3 knockout (YFT3-KO) lines at 35 and 47days post-anthesis compared with the red-fruit tomato cultivar (M82). In vitro and in vivo biochemical analyses indicated that YFT3 (S126R) possessed much lower enzymatic activities than the YFT3 protein, indicating that the S126R mutation can impair YFT3 activity. Molecular docking analysis showed that the YFT3 allele has higher ability to recruit isopentenyl pyrophosphate (IPP), but abolishes attachment of the Mg2+ cofactor to IPP, suggesting that Ser126 is a critical residue for YTF3 biochemical and physiological functions. As a result, the yft3 mutant tomato line has low carotenoid accumulation and abnormal chromoplast development, which results in yellow ripe fruits. This study provides new insights into molecular mechanisms of tomato fruit color formation and development.

Knock-out of SlDMR6-1 in tomato promotes a drought-avoidance strategy and increases tolerance to Late Blight

The DOWNY MILDEW RESISTANCE 6 (DMR6) protein is a 2-oxoglutarate (2OG) and Fe(II)-dependent oxygenase, involved in salicylic acid (SA) metabolism. SA is recognized as an abiotic stress-tolerance enhancer, and in tomato the inactivation of DMR6 was found increase its level and induce disease-resistance against several pathogens. By applying the CRISPR/Cas9 technique, we generated Sldmr6-1 tomato mutants and tested their tolerance to drought as well as to Late Blight. Wild-type tomato cultivar ‘San Marzano’ and its Sldmr6-1 mutants were subjected to water deprivation for 7 days. WT plants exhibited severe wilting, while T2Sldmr6-1 mutants showed turgid leaves and maintained higher Soil Relative Water Content. Eco-physiological measurements highlighted that Sldmr6-1 mutants adopted a water saving behavior reducing transpiration rate by decreasing stomatal conductance. Under drought stress the assimilation rate decreased as well, resulting in no alteration of the CO2 concentration in the sub-stomatal chamber and increasing the Water Use Efficiency. Furthermore, in Sldmr6-1 mutants the drought stress induced up-regulation of the anti-oxidant related genes SlAPX and SlGST as well as down-regulation of SlCYP707A2 gene, which is involved in ABA catabolism. At last we highlighted, for the first time in tomato, that Sldmr6-1 mutants exhibited a reduced susceptibility to Phytophthora infestans, the causal agent of Late Blight.

The β-galactosidase gene AtrBGAL2 regulates Akebia trifoliata fruit cracking

Cracking of Akebia trifoliata fruit at maturity is problematic for the cultivation of the horticultural crop, shortening shelf-life quality and compromising commercial value. However, the molecular mechanisms underlying this feature of A. trifoliata are not known. Genes involved in cell wall metabolism were identified by genome and transcriptome sequencing, which may play important roles in fruit cracking. One of the galactose metabolism related genes, β-galactosidase (AtrBGAL2), was identified in A. trifoliata, and overexpression (OE) of AtrBGAL2 resulted in early fruit cracking, higher water-soluble pectin contents, and lower acid-soluble pectin, cellulose, and hemicellulose content compared to the wild type. Whereas silencing of AtrBGAL2 in trifoliata by virus induced gene silencing showed opposite trends. The levels of AtrBGAL2 transcripts were 24.6 and 66.0-fold higher in OE A. trifoliata and tomato fruits, respectively, and the cell wall-related genes were also gradually greater than in control plants during fruit ripening. Whereas the expression levels of AtrBGAL2 was significantly down-regulated by 54.1 % and 73.7 % in gene silenced A. trifoliata and CRISPR/Cas9 tomato mutant plants, respectively, and cell wall-related genes were also significantly reduced. These results demonstrate that AtrBGAL2 plays important roles in regulating fruit cracking during fruit ripening.

Tomato Mutants Reveal Root and Shoot Strigolactone Involvement in Branching and Broomrape Resistance.

The phytohormones strigolactones (SLs) control root and shoot branching and are exuded from roots into the rhizosphere to stimulate interaction with mycorrhizal fungi. The exuded SLs serve as signaling molecules for the germination of parasitic plants. The broomrape Phelipanche aegyptiaca is a widespread noxious weed in various crop plants, including tomato (Solanum lycopersicum). We have isolated three mutants that impair SL functioning in the tomato variety M82: SHOOT BRANCHING 1 (sb1) and SHOOT BRANCHING 2 (sb2), which abolish SL biosynthesis, and SHOOT BRANCHING 3 (sb3), which impairs SL perception. The over-branching phenotype of the sb mutants resulted in a severe yield loss. The isogenic property of the mutations in a determinate growth variety enabled the quantitative evaluation of the contribution of SL to yield under field conditions. As expected, the mutants sb1 and sb2 were completely resistant to infection by P. aegyptiaca due to the lack of SL in the roots. In contrast, sb3 was more susceptible to P. aegyptiaca than the wild-type M82. The SL concentration in roots of the sb3 was two-fold higher than in the wild type due to the upregulation of the transcription of SL biosynthesis genes. This phenomenon suggests that the steady-state level of root SLs is regulated by a feedback mechanism that involves the SL signaling pathway. Surprisingly, grafting wild-type varieties on sb1 and sb2 rootstocks eliminated the branching phenotype and yield loss, indicating that SL synthesized in the shoots is sufficient to control shoot branching. Moreover, commercial tomato varieties grafted on sb1 were protected from P. aegyptiaca infection without significant yield loss, offering a practical solution to the broomrape crisis.

Peptide REF1 is a local wound signal promoting plant regeneration

Plants frequently encounter wounding and have evolved an extraordinary regenerative capacity to heal the wounds. However, the wound signal that triggers regenerative responses has not been identified. Here, through characterization of a tomato mutant defective in both wound-induced defense and regeneration, we demonstrate that in tomato, a plant elicitor peptide (Pep), REGENERATION FACTOR1 (REF1), acts as a systemin-independent local wound signal that primarily regulates local defense responses and regenerative responses in response to wounding. We further identified PEPR1/2 ORTHOLOG RECEPTOR-LIKE KINASE1 (PORK1) as the receptor perceiving REF1 signal for plant regeneration. REF1-PORK1-mediated signaling promotes regeneration via activating WOUND-INDUCED DEDIFFERENTIATION 1 (WIND1), a master regulator of wound-induced cellular reprogramming in plants. Thus, REF1-PORK1 signaling represents a conserved phytocytokine pathway to initiate, amplify, and stabilize a signaling cascade that orchestrates wound-triggered organ regeneration. Application of REF1 provides a simple method to boost the regeneration and transformation efficiency of recalcitrant crops.

Efficient tobacco rattle virus-induced gene editing in tomato mediated by the CRISPR/Cas9 system.

Plant virus-based sgRNA delivery strategy has been widely applied for efficient genome editing across various plant species, leveraging its significant advantages in the rapid expression and expansion of sgRNA through virus replication and movement. However, the efficacy of the virus-induced gene editing (VIGE) tool in tomato has yet to be explored. In this paper, we established a TRV-mediated CRISPR/Cas9 genome editing system in the somatic cells of tomato, reporting the validation of VIGE and evaluating the mutagenesis efficiency in both tomato leaves and fruits using high-throughput sequencing. The results demonstrated an approximate 65% efficiency of VIGE in tomato leaves for the selected target genes, with VIGE efficiency reaching up to 50% in tomato fruits. This research not only introduces an efficient tool for reverse genetics but also reveals substantial potential of VIGE in surpassing traditional tissue culture techniques for creating heritable mutations in tomato.

Variability of fruit shelf-life of commercial tomatoes and their hybrids crossing with insensitivity ethylene mutant tomato Sletr1-2

Tomatoes are climacteric fruits that experience a surge in respiration rate and ethylene production. This condition affects the fruits’ physiological deterioration, shortens their shelf-life, and degrades the quality of tomatoes. The study aimed to enhance the shelf-life of commercial tomatoes with different genetic backgrounds by utilizing the Sletr1-2 mutant, which has a lower sensitivity to ethylene. A randomized block design (RBD) was applied to achieve this goal, where 12 different plant genotypes were considered as treatments, and each treatment was replicated four times. The results showed that fruit shelf-life was extended by crossing commercial tomatoes with Sletr1-2 mutant tomatoes. Genetic variability for all shelf-life characteristics were narrow. In contrast, there were broad phenotypic variability for fruit hardness and weight loss characteristics.

The effect of fruit maturation stage on seed germination of iaa9-3 and iaa9-5 tomato mutants

The effect of fruit maturation stage on seed germination of iaa9-3 and iaa9-5 tomato mutants

Characterization and RNA-Seq Analysis of Yellow-Green Leaf Mutants in Tomato

Characterization and RNA-Seq Analysis of Yellow-Green Leaf Mutants in Tomato

Knockout of SlDCL2b attenuates the resistance of tomato to potato spindle tuber viroid infection.

RNA interference, or RNA silencing, is an important defence mechanism against viroid infection in plants. Plants encode multiple DICER-LIKE (DCL) proteins that are key components of the RNA silencing pathway. However, the roles of different DCLs in defence responses against viroid infection remain unclear. Here, we determined the function of tomato DCL2b (SlDCL2b) in defence responses against potato spindle tuber viroid (PSTVd) infection using SlDCL2b loss-of-function tomato mutant plants. Compared with wild-type plants, mutant plants were more susceptible to PSTVd infection, developing more severe symptoms earlier and accumulating higher levels of PSTVd RNAs. Moreover, we verified the feedback mechanism for the regulation of SlDCL2b expression by miR6026. Functional blocking of tomato miR6026, by expressing its target mimics, can enhance resistance to PSTVd infection in tomato plants. These findings deepen the current understanding of RNAi-based resistance against viroid infection and provide a potentially new strategy for viroid control.

Interplay among photoreceptors determines the strategy of coping with excess light in tomato.

This study investigates photoreceptor's role in the adaption of photosynthetic apparatus to high light (HL) intensity by examining the response of tomato wild type (WT) (Solanum lycopersicum L. cv. Moneymaker) and tomato mutants (phyA, phyB1, phyB2, cry1) plants to HL. Our results showed a photoreceptor-dependent effect of HL on the maximum quantum yield of photosystem II (Fv/Fm) with phyB1 exhibiting a decrease, while phyB2 exhibiting an increase in Fv/Fm. HL resulted in an increase in the efficient quantum yield of photosystem II (ΦPSII) and a decrease in the non-photochemical quantum yields (ΦNPQ and ΦN0) solely in phyA. Under HL, phyA showed a significant decrease in the energy-dependent quenching component of NPQ (qE), while phyB2 mutants showed an increase in the state transition (qT) component. Furthermore, ΔΔFv/Fm revealed that PHYB1 compensates for the deficit of PHYA in phyA mutants. PHYA signaling likely emerges as the dominant effector of PHYB1 and PHYB2 signaling within the HL-induced signaling network. In addition, PHYB1 compensates for the role of CRY1 in regulating Fv/Fm in cry1 mutants. Overall, the results of this research provide valuable insights into the unique role of each photoreceptor and their interplay in balancing photon energy and photoprotection under HL condition.

Dissection of mRNA ac4C acetylation modifications in AC and Nr fruits: insights into the regulation of fruit ripening by ethylene

N4-acetylcytidine (ac4C) modification of mRNA has been shown to be present in plant RNAs, but its regulatory function in plant remains largely unexplored. In this study, we investigated the differentially expressed mRNAs, lncRNAs and acetylation modifications of mRNAs in tomato fruits from both genotypes. By comparing wild-type (AC) tomato and the ethylene receptor-mutant (Nr) tomato from mature green (MG) to six days after the breaker (Br6) stage, we identified differences in numerous key genes related to fruit ripening and observed the corresponding lncRNAs positively regulated the target genes expression. At the post-transcriptional level, the acetylation level decreased and increased in AC and Nr tomatoes from MG to Br6 stage, respectively. The integrated analysis of RNA-seq and ac4C-seq data revealed the potential positive role of acetylation modification in regulating gene expression. Furthermore, we found differential acetylation modifications of certain transcripts (ACO, ETR, ERF, PG, CesA, β-Gal, GAD, AMY, and SUS) in AC and Nr fruits which may explain the differences in ethylene production, fruit texture, and flavor during their ripening processes. The present study provides new insights into the molecular mechanisms by which acetylation modification differentially regulates the ripening process of wild-type and mutant tomato fruits deficient in ethylene signaling.Graphical

Mediator kinase subunit CDK8 phosphorylates transcription factor TCP15 during tomato pollen development.

Pollen development in flowering plants has strong implications for reproductive success. Pollen DNA can be targeted to improve plant traits for yield and stress tolerance. In this study, we demonstrated that the Mediator subunit CYCLIN-DEPENDENT KINASE 8 (CDK8) is a key modulator of pollen development in tomato (Solanum lycopersicum). SlCDK8 knockout led to significant decreases in pollen viability, fruit yield, and fruit seed number. We also found that SlCDK8 directly interacts with transcription factor TEOSINTE BRANCHED1-CYCLOIDEA-PCF15 (SlTCP15) using yeast two-hybrid screens. We subsequently showed that SlCDK8 phosphorylates Ser 187 of SlTCP15 to promote SlTCP15 stability. Phosphorylated TCP15 directly bound to the TGGGCY sequence in the promoters of DYSFUNCTIONAL TAPETUM 1 (SlDYT1) and MYB DOMAIN PROTEIN 103 (SlMYB103), which are responsible for pollen development. Consistently, disruption of SlTCP15 resembled slcdk8 tomato mutants. In sum, our work identified a new substrate of Mediator CDK8 and revealed an important regulatory role of SlCDK8 in pollen development via cooperation with SlTCP15.

Rickettsia transmission from whitefly to plants benefits herbivore insects but is detrimental to fungal and viral pathogens.

Most insects are associated with symbiotic bacteria in nature. These symbionts play important roles in the life histories of herbivorous insects by impacting their development, survival, reproduction as well as stress tolerance. Rickettsia is one important symbiont to the agricultural pest whitefly Bemisia tabaci. Here, for the first time, we revealed that the persistence of Rickettsia symbionts in tomato leaves significantly changed the defense pattern of tomato plants. These changes benefit both sap-feeding and leaf-chewing herbivore insects, such as increasing the fecundity of whitefly adults, enhancing the growth and development of the noctuid Spodoptera litura, but reducing the pathogenicity of Verticillium fungi and TYLCV virus to tomato plants distinctively. Our study unraveled a new horizon for the multiple interaction theories among plant-insect-bacterial symbionts.