Mutant Fruit Research Articles

Genome-wide identification of long non-coding RNA targets of the tomato MADS box transcription factor RIN and function analysis.

In recent years, increasing numbers of long non-coding RNAs (lncRNAs) have been identified in humans, animals and plants, and several of them have been shown to play important roles in diverse biological processes. However, little work has been performed on the regulation mechanism of lncRNA biogenesis and expression, especially in plants. Compared with studies of tomato MADS-box transcription factor RIPENING INHIBITOR (RIN) target coding genes, there are few reports on its relationship to non-coding RNAs. The aim of the present study was to identify and explore the specific role of RIN target lncRNAs in tomato fruit development and ripening. lncRNA targets of RIN were identified by chromatin immunoprecipitation sequencing (ChIP-seq) combined with RNA deep sequencing analysis. Six selected lncRNA targets were validated by quantitative real-time PCR, ChIP and electrophoretic mobility shift assays, and we further confirmed differential expression between wild-type and ripening-deficient mutant fruit, and RIN direct binding in the promoter regions. By means of virus-induced gene silencing (VIGS) assays and a clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) genome editing strategy, the ripening-related function of a specific target lncRNA (lncRNA2155) was studied. We identified 187 lncRNAs as direct RIN targets, which exhibited RIN binding sites in their promoters and showed different expression between the wild-type and rin mutant. Six target lncRNAs were shown to bind with RIN directly in their promoters in vivo and in vitro. Moreover, using CRISPR/Cas9 technology to knock out the locus of the target lncRNA2155 indicated that it delayed fruit ripening in tomato. Collectively, these findings provide new insight into RIN in the transcriptional regulation of lncRNAs and suggest that lncRNAs will contribute to a better understanding of the RIN regulatory network that controls fruit ripening.

Impaired Taste Associative Memory and Memory Enhancement by Feeding Omija in Parkinson's Disease Fly Model.

Neurodegeneration can result in memory loss in the central nervous system (CNS) and impairment of taste and smell in the peripheral nervous system (PNS). The neurodegeneration seen in Parkinson’s disease (PD) is characterized by functional loss of dopaminergic neurons. Recent studies have also found a role for dopaminergic neurons in regulating taste memory rewards in insects. To investigate how taste memories and sugar sensitivity can be affected in PD, we utilized the DJ-1β mutant fruit fly, DJ-1βex54, as a PD model. We performed binary choice feeding assays, electrophysiology and taste-mediated memory tests to explore the function of the DJ-1β gene in terms of sugar sensitivity as well as associative taste memory. We found that PD flies exhibited an impaired ability to discriminate sucrose across a range of sugar concentrations, with normal responses at only very high concentrations of sugar. They also showed an impairment in associative taste memory. We highlight that the taste impairment and memory defect in DJ-1βex54 can be recovered by the expression of wild-type DJ-1β gene in the dopaminergic neurons. We also emphasized the role of dopaminergic neurons in restoring taste memory function. This impaired memory property of DJ-1βex54 flies also allows them to be used as a model system for finding supplementary dietary foods that can improve memory function. Here we provide evidence that the associative taste memory of both control and DJ-1βex54 flies can be enhanced with dietary supplementation of the medicinal plant, omija.

Spectrum and Frequency of Mutations Induced by Gamma Radiations in Three Varieties of Nigerian Sesame (Sesamum indicum L.)

Insufficient genetic variability is one of the major problems of plant breeding programmes, especially in sesame. Gamma radiation has been reported to be very effective in creating genetic variability in plants. Three varieties of Nigerian sesame were assessed for spectrum and frequency of mutation induced by Gamma radiations in M1 and M2 generations. The varieties (NCRIBEN-04E, NCRIBEN-01M and NCRIBEN-03L) were treated with four different doses of gamma rays (250, 350, 450 and 550 Gy). The treated and untreated seeds (control) were sown in planting bags (under field condition) to raise M1 plants. Four treatments: V1D5, V2D3, V3D2 and V3D4 (from M1 plants) were selected and bulked to obtain M2 populations. The results of M1 revealed four mutant fruit traits: multicarpellate capsule, multiple capsule per leaf axil, indehiscent capsule and terminal capsules. The highest frequencies of the traits in M1 generation were 2.50×10-2, 9.17×10-2, 1.67×10-2and3.33×10-2 respectively. The highest branching (7) was from NCRIBEN-01M, while the least (2) was from NCRIBEN-04E. The M2 plants were grouped into eight M2 lines. The dose range (250-550 Gy) was proved to be effective in inducing viable mutations in sesame.

Research on a hybrid LSSVM intelligent algorithm in short term load forecasting

For enhancing the prediction accuracy of power load forecasting, a support vector machine (SVM) prediction model based on wavelet transform and the mutant fruit fly parameter optimization intelligent algorithm (WT-MFOA-LSSVM) was presented. The load data were pretreated by wavelet transform, and the load curves were decomposed into different scales, in order to strengthen the regularity and randomness of historical data. Aiming at overcoming the problems of low convergence precision and easily relapsing into local extreme in basic fruit fly optimization algorithm (FOA), judge whether the intelligent algorithm was trapped in local extreme from the fitness variance of the population and the current optimal. Then, it was conducted by optimal individual perturbation and Gauss mutation operation and the mutant fruit flies were second times optimized, which made the accuracy of prediction model be obviously enhanced. The next few days of historical load data of a certain area of Henan Province, China, in 2015 were predicted by using WT-MFOA-LSSVM, and then the prediction results were compared with the results predicted by the SVM model and by the SVM model based on particle swarm optimization model. The results showed that WT-MFOA-LSSVM had high precision in short term load forecasting, and it had a very good practical significance.

The RIN-MC Fusion of MADS-Box Transcription Factors Has Transcriptional Activity and Modulates Expression of Many Ripening Genes.

Fruit development and ripening is regulated by genetic and environmental factors and is of critical importance for seed dispersal, reproduction, and fruit quality. Tomato (Solanum lycopersicum) ripening inhibitor (rin) mutant fruit have a classic ripening-inhibited phenotype, which is attributed to a genomic DNA deletion resulting in the fusion of two truncated transcription factors, RIN and MC In wild-type fruit, RIN, a MADS-box transcription factor, is a key regulator of the ripening gene expression network, with hundreds of gene targets controlling changes in color, flavor, texture, and taste during tomato fruit ripening; MC, on the other hand, has low expression in fruit, and the potential functions of the RIN-MC fusion gene in ripening remain unclear. Here, overexpression of RIN-MC in transgenic wild-type cv Ailsa Craig tomato fruits impaired several ripening processes, and down-regulating RIN-MC expression in the rin mutant was found to stimulate the normal yellow mutant fruit to produce a weak red color, suggesting a distinct negative role for RIN-MC in tomato fruit ripening. By comparative transcriptome analysis of rin and rin 35S::RIN-MC RNA interference fruits, a total of 1,168 and 1,234 genes were identified as potential targets of RIN-MC activation and inhibition. Furthermore, the RIN-MC fusion gene was shown to be translated into a chimeric transcription factor that was localized to the nucleus and was capable of protein interactions with other MADS-box factors. These results indicated that tomato RIN-MC fusion plays a negative role in ripening and encodes a chimeric transcription factor that modulates the expression of many ripening genes, thereby contributing to the rin mutant phenotype.

Elucidating the Mechanisms of the Tomato ovate Mutation in Regulating Fruit Quality Using Proteomics Analysis.

The ovate mutation has frequently been used to study changes in fruit shape but not fruit quality. A deterioration in fruit quality associated with the ovate mutation was discovered in this study. To elucidate how ovate influences the quality of fruit, we performed a proteomics analysis of the fruits of the ovate mutant (LA3543) and wild-type ("Ailsa Craig", LA2838A) using tandem mass tag analysis. The results indicated that the ovate mutation significantly influences fruit quality in a number of ways, including by reducing the expression of 1-aminocyclopropane-1-carboxylic acid oxidase 3 (ACO3) in ethylene biosynthesis, improving firmness by reducing the amount of pectinesterase and polygalacturonase, reducing sugar accumulation by downregulating the abundance of mannan endo-1,4-β-mannosidase 4, β-galactosidase, and β-amylase, and reducing the malic acid content by downregulating the accumulation of malic enzymes and malate synthase. These findings could inform future improvements in fruit quality.

Re-evaluation of the rin mutation and the role of RIN in the induction of tomato ripening.

Tomato (Solanum lycopersicum) rin mutants completely fail to ripen: they do not produce red pigmentation, soften or induce an ethylene burst. Therefore, RIN has long been believed to function as a major regulator that is essential for the induction of ripening. Here, we provide evidence contradicting this concept of RIN function, showing induction of fruit ripening in the absence of RIN. A CRISPR/Cas9-mediated RIN-knockout mutation did not repress initiation of ripening and the mutant fruits showed moderate red colouring. Moreover, inactivation of the rin mutant allele partially restored the induction of ripening. Therefore, RIN is not required for the initiation of ripening and rin is not a null mutation, but rather is a gain-of-function mutation that produces a protein that actively represses ripening. Since the discovery of the rin mutant a half-century ago, many models have depicted RIN as indispensable for the induction of ripening; these models should be reconsidered in light of these results.

Critical roles of DNA demethylation in the activation of ripening-induced genes and inhibition of ripening-repressed genes in tomato fruit

DNA methylation is a conserved epigenetic mark important for genome integrity, development, and environmental responses in plants and mammals. Active DNA demethylation in plants is initiated by a family of 5-mC DNA glycosylases/lyases (i.e., DNA demethylases). Recent reports suggested a role of active DNA demethylation in fruit ripening in tomato. In this study, we generated loss-of-function mutant alleles of a tomato gene, SlDML2, which is a close homolog of the Arabidopsis DNA demethylase gene ROS1 In the fruits of the tomato mutants, increased DNA methylation was found in thousands of genes. These genes included not only hundreds of ripening-induced genes but also many ripening-repressed genes. Our results show that SlDML2 is critical for tomato fruit ripening and suggest that active DNA demethylation is required for both the activation of ripening-induced genes and the inhibition of ripening-repressed genes.

Targeted gene disruption coupled with metabolic screen approach to uncover the LEAFY COTYLEDON1-LIKE4 (L1L4) function in tomato fruit metabolism.

Functional analysis of tomato L1L4 master transcription factor resulted in important metabolic changes affecting tomato fruit quality. Tomato fruits from mutant lines bearing targeted disruption of the heterotrimeric nuclear transcription factor Y (NF-Y) transcription factor (TF) gene LEAFY-COTYLEDON1-LIKE4 (L1L4, NF-YB6), a master regulator of biosynthesis for seed storage proteins and fatty acids, were evaluated for metabolites content and morphology. Metabolic screens using LC-MS/MS-based analysis and physico-chemical methods in different L1L4 mutants of the fourth generation allowed a comparative assessment of the effects of the TF disruption. Mutagenesis resulted in fruits phenotypically similar to wild-type with subtle shape differences in the distal end protrusion and symmetry. Conversely, mutant fruits from independent lines had significant variation in moisture content, titratable acidity and overall metabolite profiles including oxalic and citric acid, fructose, β-carotene, total polyphenols and antioxidants. Lines 6, 7 and 9 were the richest in β-carotene and antioxidant activity, line 4 in ascorbic acid and lines 4 and 8 in succinic acid. The reduced content of the anti-nutrient oxalic acid in several mutant fruits suggests that L1L4 gene may regulate the accumulation of this compound during fruit development. Detailed LC-MS/MS analysis of mutant seeds showed substantial differences in bioactive compounds compared to wild-type seeds. Taken together, the results suggest that the L1L4 TF is a significant regulator of metabolites both in tomato fruit and seeds providing a molecular target for crop improvement. Elucidation of the candidate genes encoding key enzymes in the affected metabolic pathways aimed to facilitate the L1L4 gene network exploration and eventually lead to systems biology approaches in tomato fruit quality.

Circadian Expression of PERIOD and the Pigment-Dispersing Factor in the yellow white mutant Drosophila melanogaster

Daily behavioral andphysiological rhythms in most animals are regulated by a circadian clock. Inthe fruit fly, Drosophila melanogaster, this clock consists of a networkof about 150 cerebral neurons. Different parameters of the fruit fly circadianlocomotion are attributed to specific neuronal subsets and the molecularrhythms of clock genes and proteins within them. To understand the clockmachinery, many clock mutant flies have been used. yellow white (y w) mutation in D. melanogaster cause impairedmelanisation, eye pigmentation loss, and behavioral alterations includingchanges in circadian locomotion. This study investigates the possible molecularbackground for these circadian alterations. Results revealed that in the outputpathway of the clock, the Pigment-Dispersing Factor (PDF) expression wassuppressed in y w mutant fruit flies compared to Canton S (CS) wild-type in the PDF+ clock neurons. Onthe other hand, the degradation of the PERIOD (PER) protein was significantlydelayed in yw mutants and their levels was higher, especially atthe transition from dark to light. The combined effect of elevated PER levelsand suppressed PDF signaling provides an explanation for the delayed morninglocomotor activity peak (M) and advanced evening peak (E) of y w fliescompared to CS.It could be concluded that mutations affecting eye pigmentation like the y w mutationcould have profound effects on the circadian regulation of behavior and theirunderlying molecular oscillations in clock neurons. These effects reduce theplasticity and robustness of the circadian clock and expose the flies to higherlevels of the environmental risk of desiccation.

The Analysis of Physiological Variations in M2 Generation of Solanum melongena L. Mutagenized by Ethyl Methane Sulfonate.

The eggplant was mutagenized with ethyl methane sulfonate (EMS) to enhance its genetic variability in our previous paper. In this article, we further analyzed the phenotype of M2 generation of mutant eggplants. A total of 325 independent M2 families were investigated for phenotypic variation. In addition to the visible phenotypic variation, chlorogenic acid (CGA) concentrations were analyzed in 26 fruits of mutants with High Performance Liquid Chromatography assay. Seventeen fruits exhibited significantly higher concentrations of CGAs than those in wild-type. The anthocyanin concentration of S9-1, the purple black mutant, was higher than WT, meanwhile, the anthocyanin concentration of L6-4 and U36-1 was lower than WT. Furthermore, our RT-PCR result demonstrated that the expression levels of anthocyanin biosynthetic genes, except for SmPAL, were increased in S9-1, and the regulator SmMYB1 was decreased in L6-4 and U36-1 mutants. Together, our data indicated that, M2 generation showed abundant phenotypic variations and the strong potential usage for next step of breeding and molecular genetic mechanisms in eggplant.

Different People, Different Outcomes: Assessing Genetic Susceptibility to Lead Exposures.

Prenatal and early-life exposures to lead have long been known to cause an array of adverse developmental effects in children. 1 However, the variety of responses to lead, even with exposures, suggests that an individual’s genetic background might influence how lead toxicity manifests. 2 , 3 , 4 A new study in fruit flies scrutinizes susceptibility on the genetic level and highlights genes that might help shape individual responses. 5 “There are several problems in trying to find out variations and susceptibility to lead in human populations,” says study coauthor Robert Anholt, a professor of biological sciences at North Carolina State University. Uncontrolled genetic backgrounds, mixtures of contaminants in the environment, and additional factors such as diet and smoking all muddy the waters; in addition, lead’s effects may not be apparent until years after the exposure. 1 Similar lead exposures can cause very different responses in different people. An individual’s genetic makeup may be one reason why. To offset some of the complexity, model systems such as fruit flies (Drosophila melanogaster) are used to identify candidate genes, which can then be studied more closely in humans. 5 “Drosophila may seem a little far-fetched, but they are actually a really good system,” says Anholt. Many Drosophila genes have human orthologs; that is, they perform the same function in both species. 6 Anholt calls fruit flies “probably the most powerful, most versatile genetic model we have.” For the current study the researchers used the Drosophila melanogaster Genetic Reference Panel. This unique collection of more than 200 Drosophila lines (or varieties) represents varied genetic backgrounds, with all individuals within the same line being nearly identical genetically. 6 The researchers placed 50 larvae from each of 200 selected lines on control or lead-spiked medium. The timing of their development was based on how many days passed until adult flies emerged, and viability was assessed by counting the number of larvae that survived to adulthood. Twenty adult flies of each sex were then randomly selected from each line and individually placed in tubes. There, their activity was measured based on how many times they crossed an infrared beam. Based on differences in the traits of viability, development, and activity between the lines, the researchers conducted genome-wide association analyses to identify candidate genes potentially related to those traits. The researchers confirmed the results of the analyses by rearing 20 mutant fruit fly lines on control and lead-supplemented media. Each line contained a specific variant of a single candidate gene that had been implicated in altered activity and development time. Finally, human orthologs of many of the candidate Drosophila genes were identified. In general, lead-exposed flies developed more slowly and were less likely to survive than control flies, although these factors varied greatly among both control and lead-exposed lines. Of the 200 lines used, only 166 produced enough adults for activity measurements. Again, extensive variation occurred among lines and among the lead-exposed flies, with some becoming more active and others becoming less active. The majority of the implicated genes were associated with nervous system development and function. “Some genetic factors may modulate the sensitivity to lead, and there have been several genes that have been identified that can influence the accumulation and toxicokinetics of lead in humans 7 , 8 , 9 , 10 ,” says Jay Schneider, a professor of pathology, anatomy, and cell biology at Thomas Jefferson University, who was not involved in the current study. These genes are not targets of lead toxicity; rather, they influence the absorption, distribution, metabolism, and excretion of lead. 3 , 4 Other genes might further influence how cells respond to lead. 2 , 5 This study was not intended to gauge the level at which lead is toxic, and it did not reveal the mechanisms by which lead causes adverse effects. However, if these results are echoed in humans, they could help us understand why and how the effects from lead exposure are expressed differently in different people, says Schneider. “This kind of work shows how much we still have to learn about the potential effects that lead has on the body,” he says. “We know a lot, but there’s still a lot we don’t know.”

Effects of gravity on positive phototaxis in fruit fly Drosophila melanogaster

AbstractGeotaxis and phototaxis are movements in response to gravity and light, respectively, and are commonly observed in nature. The interactions between these two types of movement have been shown to confer ecological advantages to many taxa. Although several studies have been conducted on phototaxis and geotaxis in various organisms, reports on the interactions between positive phototaxis and negative geotaxis are lacking. In the fruit fly, Drosophila melanogaster, any direct interactions that exist between positive phototaxis and negative geotaxis are yet to be determined and the ecological significance of such interactions remains unclear. In the present study, the effects of gravity on positive phototaxis in a Y‐maze were investigated using the Canton‐S wild type and gravity‐sensing‐deficient pyx3 mutant fruit flies. Gravity sensing was not necessary for horizontal positive phototaxis, but was required for vertical positive phototaxis. These results suggest that gravitoreception may selectively modulate positive phototaxis depending on the vertical and horizontal movements of the fruit flies.

The Glycerol-3-Phosphate Acyltransferase GPAT6 from Tomato Plays a Central Role in Fruit Cutin Biosynthesis.

The thick cuticle covering and embedding the epidermal cells of tomato (Solanum lycopersicum) fruit acts not only as a protective barrier against pathogens and water loss but also influences quality traits such as brightness and postharvest shelf-life. In a recent study, we screened a mutant collection of the miniature tomato cultivar Micro-Tom and isolated several glossy fruit mutants in which the abundance of cutin, the polyester component of the cuticle, was strongly reduced. We employed a newly developed mapping-by-sequencing strategy to identify the causal mutation underlying the cutin deficiency in a mutant thereafter named gpat6-a (for glycerol-3-phosphate acyltransferase6). To this end, a backcross population (BC1F2) segregating for the glossy trait was phenotyped. Individuals displaying either a wild-type or a glossy fruit trait were then pooled into bulked populations and submitted to whole-genome sequencing prior to mutation frequency analysis. This revealed that the causal point mutation in the gpat6-a mutant introduces a charged amino acid adjacent to the active site of a GPAT6 enzyme. We further showed that this mutation completely abolished the GPAT activity of the recombinant protein. The gpat6-a mutant showed perturbed pollen formation but, unlike a gpat6 mutant of Arabidopsis (Arabidopsis thaliana), was not male sterile. The most striking phenotype was observed in the mutant fruit, where cuticle thickness, composition, and properties were altered. RNA sequencing analysis highlighted the main processes and pathways that were affected by the mutation at the transcriptional level, which included those associated with lipid, secondary metabolite, and cell wall biosynthesis.

An INDEHISCENT-Controlled Auxin Response Specifies the Separation Layer in Early Arabidopsis Fruit

Seed dispersal is an important moment in the life cycle of a plant species. In Arabidopsis thaliana, it is dependent on transcription factor INDEHISCENT (IND)-mediated specification of a separation layer in the dehiscence zone found in the margin between the valves (carpel walls) and the central replum of the developing fruit. It was proposed that IND specifies the separation layer by inducing a local auxin minimum at late stages of fruit development. Here we show that morphological differences between the ind mutant and wild-type fruit already arise at early stages of fruit development, coinciding with strong IND expression in the valve margin. We show that IND-reduced PIN-FORMED3 (PIN3) auxin efflux carrier abundance leads to an increased auxin response in the valve margin during early fruit development, and that the concomitant cell divisions that form the dehiscence zone are lacking in ind mutant fruit. Moreover, IND promoter-driven ectopic expression of the AGC kinases PINOID (PID) and WAG2 induced indehiscence by expelling auxin from the valve margin at stages 14–16 of fruit development through increased PIN3 abundance. Our results show that IND, besides its role at late stages of Arabidopsis fruit development, functions at early stages to facilitate the auxin-triggered cell divisions that form the dehiscence zone.

The Cellular Compass

The article offers the authors' insight on the contribution of polarity proteins on the development processes in the animal kingdom. Authors discuss the study in 1980 which investigates tissue polarity in fruit flies to identify mutant fruit flies with polarity system defects. They mentions various studies conducted by several researchers to analyse the distribution of polarity proteins which could be used to regulate cytoskeleton formation.

Fruit Ripening Regulation of α-Mannosidase Expression by the MADS Box Transcription Factor RIPENING INHIBITOR and Ethylene.

α-Mannosidase (α-Man), a fruit ripening-specific N-glycan processing enzyme, is involved in ripening-associated fruit softening process. However, the regulation of fruit-ripening specific expression of α-Man is not well understood. We have identified and functionally characterized the promoter of tomato (Solanum lycopersicum) α-Man to provide molecular insights into its transcriptional regulation during fruit ripening. Fruit ripening-specific activation of the α-Man promoter was revealed by analysing promoter driven expression of beta-glucuronidase (GUS) reporter in transgenic tomato. We found that RIPENING INHIBITOR (RIN), a MADS box family transcription factor acts as positive transcriptional regulator of α-Man during fruit ripening. RIN directly bound to the α-Man promoter sequence and promoter activation/α-Man expression was compromised in rin mutant fruit. Deletion analysis revealed that a promoter fragment (567 bp upstream of translational start site) that contained three CArG boxes (binding sites for RIN) was sufficient to drive GUS expression in fruits. In addition, α-Man expression was down-regulated in fruits of Nr mutant which is impaired in ethylene perception and promoter activation/α-Man expression was induced in wild type following treatment with a precursor of ethylene biosynthesis, 1-aminocyclopropane-1-carboxylic acid (ACC). Although, α-Man expression was induced in rin mutant after ACC treatment, the transcript level was less as compared to ACC-treated wild type. Taken together, these results suggest RIN-mediated direct transcriptional regulation of α-Man during fruit ripening and ethylene may acts in RIN-dependent and -independent ways to regulate α-Man expression.

Seed development and viviparous germination in one accession of a tomato rin mutant.

In an experimental field, seed vivipary occurred in one accession of tomato rin mutant fruit at approximately 45–50 days after pollination (DAP). In this study, the possible contributory factors to this viviparous germination were investigated. Firstly, developing seeds were freshly excised from the fruit tissue every 5 days from 25–60 DAP. Germination occurred when isolated seeds were incubated on water, but was inhibited when they remained ex situ in fruit mucilage gel. The effect of abscisic acid (ABA) and osmoticum, separate and together, on germination of developing seeds was investigated. Additionally, ABA content in the seed and mucilage gel, as well as fruit osmolality were measured. The results showed that ABA concentrations in seeds were low during early development and increased later, peaking at about 50 DAP. ABA concentrations in rin accession were similar to those of the control cultivar and thus are not directly associated with the occurrence of vivipary. Developing seeds of rin accession are more sensitive than control seeds to all inhibitory compounds. However, osmolality in rin fruit at later developmental stages becomes less negative that is required to permit germination of developing seeds. Hence, hypo-osmolality in rin fruit may be an important factor in permitting limited viviparous germination.

TOMATOMA Update: Phenotypic and Metabolite Information in the Micro-Tom Mutant Resource.

TOMATOMA (http://tomatoma.nbrp.jp/) is a tomato mutant database providing visible phenotypic data of tomato mutant lines generated by ethylmethane sulfonate (EMS) treatment or γ-ray irradiation in the genetic background of Micro-Tom, a small and rapidly growing variety. To increase mutation efficiency further, mutagenized M3 seeds were subjected to a second round of EMS treatment; M3M1 populations were generated. These plants were self-pollinated, and 4,952 lines of M3M2 mutagenized seeds were generated. We checked for visible phenotypes in the M3M2 plants, and 618 mutant lines with 1,194 phenotypic categories were identified. In addition to the phenotypic information, we investigated Brix values and carotenoid contents in the fruits of individual mutants. Of 466 samples from 171 mutant lines, Brix values and carotenoid contents were between 3.2% and 11.6% and 6.9 and 37.3 µg g(-1) FW, respectively. This metabolite information concerning the mutant fruits would be useful in breeding programs as well as for the elucidation of metabolic regulation. Researchers are able to browse and search this phenotypic and metabolite information and order seeds of individual mutants via TOMATOMA. Our new Micro-Tom double-mutagenized populations and the metabolic information could provide a valuable genetic toolkit to accelerate tomato research and potential breeding programs.

감귤 돌연변이체의 인간 암세포 증식 억제와 자연사멸 증강효과

Citrus mutant fruits were induced by irradiation of citrus budsticks with 120 Gy of cobalt ( 60 CO) gamma irradiation. The citrus mutant inhibited the growth and induced apoptosis in various human cancer cells, including A549, HepG2, HCT116, MCF-7, and Hela. The results of a trypan blue exclusion assay showed that citrus mutant fruits exhibited excellent antiproliferation activity in various human cancer cells and low cytotoxicity in normal 16HBE140and CHANG cells. In addition, the cell death induced by the citrus mutant fruits was associated with an increased population of cells in sub-G1 phase, and it caused DNA fragmentation in human lung adenocarcinoma A549 and hepatocellular carcinoma HepG2 cells. It also up-regulated the amount of cellular nitric oxide (NO • ) produced as a result of nitric oxide synthase (NOS) activation and suppressed the inhibitor of apoptosis protein (IAP) family in A549 and HepG2 cells. These findings indicate that the citrus mutant fruits activates the NO • -mediated apoptotic pathway in A549 and HepG2 cells. It may merit further investigation as a potential chemotherapeutic and chemopreventive agent for the treatment of various types of cancer cells. The results provide important major new insights into the mechanisms of the anticancer activity of citrus mutant fruits.