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1.
J Exp Bot ; 74(1): 178-193, 2023 01 01.
Artículo en Inglés | MEDLINE | ID: mdl-36260406

RESUMEN

Pollen development is a crucial biological process indispensable for seed set in flowering plants and for successful crop breeding. However, little is known about the molecular mechanisms regulating pollen development in crop species. This study reports a novel male-sterile tomato mutant, pollen deficient 2 (pod2), characterized by the production of non-viable pollen grains and resulting in the development of small parthenocarpic fruits. A combined strategy of mapping-by-sequencing and RNA interference-mediated gene silencing was used to prove that the pod2 phenotype is caused by the loss of Solanum lycopersicum G-type lectin receptor kinase II.9 (SlG-LecRK-II.9) activity. In situ hybridization of floral buds showed that POD2/SlG-LecRK-II.9 is specifically expressed in tapetal cells and microspores at the late tetrad stage. Accordingly, abnormalities in meiosis and tapetum programmed cell death in pod2 occurred during microsporogenesis, resulting in the formation of four dysfunctional microspores leading to an aberrant microgametogenesis process. RNA-seq analyses supported the existence of alterations at the final stage of microsporogenesis, since we found tomato deregulated genes whose counterparts in Arabidopsis are essential for the normal progression of male meiosis and cytokinesis. Collectively, our results revealed the essential role of POD2/SlG-LecRK-II.9 in regulating tomato pollen development.


Asunto(s)
Arabidopsis , Fenómenos Biológicos , Solanum lycopersicum , Solanum lycopersicum/genética , Lectinas/genética , Lectinas/metabolismo , Proteínas Quinasas Dependientes de GMP Cíclico/genética , Proteínas Quinasas Dependientes de GMP Cíclico/metabolismo , Fitomejoramiento , Polen/metabolismo , Arabidopsis/genética , Regulación de la Expresión Génica de las Plantas
2.
Proc Natl Acad Sci U S A ; 117(14): 8187-8195, 2020 04 07.
Artículo en Inglés | MEDLINE | ID: mdl-32179669

RESUMEN

A dramatic evolution of fruit size has accompanied the domestication and improvement of fruit-bearing crop species. In tomato (Solanum lycopersicum), naturally occurring cis-regulatory mutations in the genes of the CLAVATA-WUSCHEL signaling pathway have led to a significant increase in fruit size generating enlarged meristems that lead to flowers with extra organs and bigger fruits. In this work, by combining mapping-by-sequencing and CRISPR/Cas9 genome editing methods, we isolated EXCESSIVE NUMBER OF FLORAL ORGANS (ENO), an AP2/ERF transcription factor which regulates floral meristem activity. Thus, the ENO gene mutation gives rise to plants that yield larger multilocular fruits due to an increased size of the floral meristem. Genetic analyses indicate that eno exhibits synergistic effects with mutations at the LOCULE NUMBER (encoding SlWUS) and FASCIATED (encoding SlCLV3) loci, two central players in the evolution of fruit size in the domestication of cultivated tomatoes. Our findings reveal that an eno mutation causes a substantial expansion of SlWUS expression domains in a flower-specific manner. In vitro binding results show that ENO is able to interact with the GGC-box cis-regulatory element within the SlWUS promoter region, suggesting that ENO directly regulates SlWUS expression domains to maintain floral stem-cell homeostasis. Furthermore, the study of natural allelic variation of the ENO locus proved that a cis-regulatory mutation in the promoter of ENO had been targeted by positive selection during the domestication process, setting up the background for significant increases in fruit locule number and fruit size in modern tomatoes.


Asunto(s)
Frutas/genética , Proteínas de Homeodominio/genética , Meristema/crecimiento & desarrollo , Proteínas de Plantas/metabolismo , Solanum lycopersicum/fisiología , Factores de Transcripción/metabolismo , Proliferación Celular/genética , Producción de Cultivos , Domesticación , Frutas/crecimiento & desarrollo , Regulación de la Expresión Génica de las Plantas , Genes de Plantas/genética , Meristema/citología , Mutación , Proteínas de Plantas/genética , Regiones Promotoras Genéticas , Sitios de Carácter Cuantitativo/genética , Células Madre/fisiología , Factores de Transcripción/genética
3.
New Phytol ; 234(3): 1059-1074, 2022 05.
Artículo en Inglés | MEDLINE | ID: mdl-35170044

RESUMEN

CRABS CLAW (CRC) orthologues play a crucial role in floral meristem (FM) determinacy and gynoecium formation across angiosperms, the key developmental processes for ensuring successful plant reproduction and crop production. However, the mechanisms behind CRC mediated FM termination are far from fully understood. Here, we addressed the functional characterization of tomato (Solanum lycopersicum) paralogous CRC genes. Using mapping-by-sequencing, RNA interference and CRISPR/Cas9 techniques, expression analyses, protein-protein interaction assays and Arabidopsis complementation experiments, we examined their potential roles in FM determinacy and carpel formation. We revealed that the incomplete penetrance and variable expressivity of the indeterminate carpel-inside-carpel phenotype observed in fruit iterative growth (fig) mutant plants are due to the lack of function of the S. lycopersicum CRC homologue SlCRCa. Furthermore, a detailed functional analysis of tomato CRC paralogues, SlCRCa and SlCRCb, allowed us to propose that they operate as positive regulators of FM determinacy by acting in a compensatory and partially redundant manner to safeguard the proper formation of flowers and fruits. Our results uncover for the first time the physical interaction of putative CRC orthologues with members of the chromatin remodelling complex that epigenetically represses WUSCHEL expression through histone deacetylation to ensure the proper termination of floral stem cell activity.


Asunto(s)
Proteínas de Arabidopsis , Solanum lycopersicum , Proteínas de Arabidopsis/metabolismo , Ensamble y Desensamble de Cromatina , Flores , Regulación de la Expresión Génica de las Plantas , Solanum lycopersicum/genética , Solanum lycopersicum/metabolismo , Meristema/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo
4.
Plant Cell Environ ; 43(7): 1722-1739, 2020 07.
Artículo en Inglés | MEDLINE | ID: mdl-32329086

RESUMEN

Increasing evidences highlight the importance of DEAD-box RNA helicases in plant development and stress responses. In a previous study, we characterized the tomato res mutant (restored cell structure by salinity), showing chlorosis and development alterations that reverted under salt-stress conditions. Map-based cloning demonstrates that RES gene encodes SlDEAD39, a chloroplast-targeted DEAD-box RNA helicase. Constitutive expression of SlDEAD39 complements the res mutation, while the silencing lines had a similar phenotype than res mutant, which is also reverted under salinity. Functional analysis of res mutant proved SlDEAD39 is involved in the in vivo processing of the chloroplast, 23S rRNA, at the hidden break-B site, a feature also supported by in vitro binding experiments of the protein. In addition, our results show that other genes coding for chloroplast-targeted DEAD-box proteins are induced by salt-stress, which might explain the rescue of the res mutant phenotype. Interestingly, salinity restored the phenotype of res adult plants by increasing their sugar content and fruit yield. Together, these results propose an unprecedented role of a DEAD-box RNA helicase in regulating plant development and stress response through the proper ribosome and chloroplast functioning, which, in turn, represents a potential target to improve salt tolerance in tomato crops.


Asunto(s)
ARN Helicasas DEAD-box/fisiología , Proteínas de Plantas/fisiología , Solanum lycopersicum/crecimiento & desarrollo , Northern Blotting , Cloroplastos/metabolismo , ARN Helicasas DEAD-box/metabolismo , Regulación de la Expresión Génica de las Plantas , Solanum lycopersicum/metabolismo , Solanum lycopersicum/fisiología , Proteínas de Plantas/metabolismo , Plantas Modificadas Genéticamente , Reacción en Cadena en Tiempo Real de la Polimerasa , Estrés Salino
5.
Plant J ; 96(2): 300-315, 2018 10.
Artículo en Inglés | MEDLINE | ID: mdl-30003619

RESUMEN

Pollen development is a crucial step in higher plants, which not only makes possible plant fertilization and seed formation, but also determines fruit quality and yield in crop species. Here, we reported a tomato T-DNA mutant, pollen deficient1 (pod1), characterized by an abnormal anther development and the lack of viable pollen formation, which led to the production of parthenocarpic fruits. Genomic analyses and the characterization of silencing lines proved that pod1 mutant phenotype relies on the tomato SlMED18 gene encoding the subunit 18 of Mediator multi-protein complex involved in RNA polymerase II transcription machinery. The loss of SlMED18 function delayed tapetum degeneration, which resulted in deficient microspore development and scarce production of viable pollen. A detailed histological characterization of anther development proved that changes during microgametogenesis and a significant delay in tapetum degeneration are associated with a high proportion of degenerated cells and, hence, should be responsible for the low production of functional pollen grains. Expression of pollen marker genes indicated that SlMED18 is essential for the proper transcription of a subset of genes specifically required to pollen formation and fruit development, revealing a key role of SlMED18 in male gametogenesis of tomato. Additionally, SlMED18 is able to rescue developmental abnormalities of the Arabidopsis med18 mutant, indicating that most biological functions have been conserved in both species.


Asunto(s)
Complejo Mediador/metabolismo , Solanum lycopersicum/genética , Arabidopsis/genética , Arabidopsis/crecimiento & desarrollo , Arabidopsis/fisiología , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Gametogénesis en la Planta/genética , Solanum lycopersicum/crecimiento & desarrollo , Solanum lycopersicum/fisiología , Complejo Mediador/genética , Mutación , Fenotipo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Polen/genética , Polen/crecimiento & desarrollo , Polen/fisiología
6.
BMC Plant Biol ; 19(1): 141, 2019 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-30987599

RESUMEN

BACKGROUND: Tomato mutants altered in leaf morphology are usually identified in the greenhouse, which demands considerable time and space and can only be performed in adequate periods. For a faster but equally reliable scrutiny method we addressed the screening in vitro of 971 T-DNA lines. Leaf development was evaluated in vitro in seedlings and shoot-derived axenic plants. New mutants were characterized in the greenhouse to establish the relationship between in vitro and in vivo leaf morphology, and to shed light on possible links between leaf development and agronomic traits, a promising field in which much remains to be discovered. RESULTS: Following the screening in vitro of tomato T-DNA lines, putative mutants altered in leaf morphology were evaluated in the greenhouse. The comparison of results in both conditions indicated a general phenotypic correspondence, showing that in vitro culture is a reliable system for finding mutants altered in leaf development. Apart from providing homogeneous conditions, the main advantage of screening in vitro lies in the enormous time and space saving. Studies on the association between phenotype and nptII gene expression showed co-segregation in two lines (P > 99%). The use of an enhancer trap also allowed identifying gain-of-function mutants through reporter expression analysis. These studies suggested that genes altered in three other mutants were T-DNA tagged. New mutants putatively altered in brassinosteroid synthesis or perception, mutations determining multiple pleiotropic effects, lines affected in organ curvature, and the first tomato mutant with helical growth were discovered. Results also revealed new possible links between leaf development and agronomic traits, such as axillary branching, flower abscission, fruit development and fruit cracking. Furthermore, we found that the gene tagged in mutant 2635-MM encodes a Sterol 3-beta-glucosyltransferase. Expression analysis suggested that abnormal leaf development might be due to the lack-off-function of this gene. CONCLUSION: In vitro culture is a quick, efficient and reliable tool for identifying tomato mutants altered in leaf morphology. The characterization of new mutants in vivo revealed new links between leaf development and some agronomic traits. Moreover, the possible implication of a gene encoding a Sterol 3-beta-glucosyltransferase in tomato leaf development is reported.


Asunto(s)
Glucosiltransferasas/genética , Solanum lycopersicum/genética , Flores/enzimología , Flores/genética , Flores/crecimiento & desarrollo , Frutas/enzimología , Frutas/genética , Frutas/crecimiento & desarrollo , Solanum lycopersicum/enzimología , Solanum lycopersicum/crecimiento & desarrollo , Mutación , Fenotipo , Hojas de la Planta/enzimología , Hojas de la Planta/genética , Hojas de la Planta/crecimiento & desarrollo , Proteínas de Plantas/genética
7.
Plant Physiol ; 176(2): 1676-1693, 2018 02.
Artículo en Inglés | MEDLINE | ID: mdl-29229696

RESUMEN

Characterization of a new tomato (Solanum lycopersicum) T-DNA mutant allowed for the isolation of the CALCINEURIN B-LIKE PROTEIN 10 (SlCBL10) gene whose lack of function was responsible for the severe alterations observed in the shoot apex and reproductive organs under salinity conditions. Physiological studies proved that SlCBL10 gene is required to maintain a proper low Na+/Ca2+ ratio in growing tissues allowing tomato growth under salt stress. Expression analysis of the main responsible genes for Na+ compartmentalization (i.e. Na+/H+ EXCHANGERs, SALT OVERLY SENSITIVE, HIGH-AFFINITY K+ TRANSPORTER 1;2, H+-pyrophosphatase AVP1 [SlAVP1] and V-ATPase [SlVHA-A1]) supported a reduced capacity to accumulate Na+ in Slcbl10 mutant leaves, which resulted in a lower uploading of Na+ from xylem, allowing the toxic ion to reach apex and flowers. Likewise, the tomato CATION EXCHANGER 1 and TWO-PORE CHANNEL 1 (SlTPC1), key genes for Ca2+ fluxes to the vacuole, showed abnormal expression in Slcbl10 plants indicating an impaired Ca2+ release from vacuole. Additionally, complementation assay revealed that SlCBL10 is a true ortholog of the Arabidopsis (Arabidopsis thaliana) CBL10 gene, supporting that the essential function of CBL10 is conserved in Arabidopsis and tomato. Together, the findings obtained in this study provide new insights into the function of SlCBL10 in salt stress tolerance. Thus, it is proposed that SlCBL10 mediates salt tolerance by regulating Na+ and Ca2+ fluxes in the vacuole, cooperating with the vacuolar cation channel SlTPC1 and the two vacuolar H+-pumps, SlAVP1 and SlVHA-A1, which in turn are revealed as potential targets of SlCBL10.


Asunto(s)
Calcineurina/metabolismo , Calcio/metabolismo , Intercambiadores de Sodio-Hidrógeno/metabolismo , Sodio/metabolismo , Solanum lycopersicum/genética , Calcineurina/genética , Homeostasis , Solanum lycopersicum/fisiología , Mutación , Fenotipo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Salinidad , Estrés Salino , Tolerancia a la Sal , Intercambiadores de Sodio-Hidrógeno/genética , Vacuolas/metabolismo
8.
J Exp Bot ; 70(20): 5731-5744, 2019 10 24.
Artículo en Inglés | MEDLINE | ID: mdl-31328220

RESUMEN

Arlequin (Alq) is a gain-of-function mutant whose most relevant feature is that sepals are able to become fruit-like organs due to the ectopic expression of the ALQ-TAGL1 gene. The role of this gene in tomato fruit ripening was previously demonstrated. To discover new functional roles for ALQ-TAGL1, and most particularly its involvement in the fruit set process, a detailed characterization of Alq yield-related traits was performed. Under standard conditions, the Alq mutant showed a much higher fruit set rate than the wild type. A significant percentage of Alq fruits were seedless. The results showed that pollination-independent fruit set in Alq is due to early transition from flower to fruit. Analysis of endogenous hormones in Alq suggests that increased content of cytokinins and decreased level of abscisic acid may account for precocious fruit set. Comparative expression analysis showed relevant changes of several genes involved in cell division, gibberellin metabolism, and the auxin signalling pathway. Since pollination-independent fruit set may be a very useful strategy for maintaining fruit production under adverse conditions, fruit set and yield in Alq plants under moderate salinity were assessed. Interestingly, Alq mutant plants showed a high yield under saline conditions, similar to that of Alq and the wild type under unstressed conditions.


Asunto(s)
Flores/metabolismo , Flores/fisiología , Frutas/metabolismo , Frutas/fisiología , Proteínas de Plantas/metabolismo , Solanum lycopersicum/metabolismo , Solanum lycopersicum/fisiología , Ácido Abscísico/metabolismo , División Celular/genética , División Celular/fisiología , Citocininas/metabolismo , Flores/genética , Frutas/genética , Regulación de la Expresión Génica de las Plantas/genética , Regulación de la Expresión Génica de las Plantas/fisiología , Giberelinas/metabolismo , Solanum lycopersicum/genética , Proteínas de Plantas/genética , Plantas Modificadas Genéticamente/genética , Plantas Modificadas Genéticamente/metabolismo , Plantas Modificadas Genéticamente/fisiología , Polinización/genética , Polinización/fisiología
9.
Plant Biotechnol J ; 15(11): 1439-1452, 2017 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-28317264

RESUMEN

With the completion of genome sequencing projects, the next challenge is to close the gap between gene annotation and gene functional assignment. Genomic tools to identify gene functions are based on the analysis of phenotypic variations between a wild type and its mutant; hence, mutant collections are a valuable resource. In this sense, T-DNA collections allow for an easy and straightforward identification of the tagged gene, serving as the basis of both forward and reverse genetic strategies. This study reports on the phenotypic and molecular characterization of an enhancer trap T-DNA collection in tomato (Solanum lycopersicum L.), which has been produced by Agrobacterium-mediated transformation using a binary vector bearing a minimal promoter fused to the uidA reporter gene. Two genes have been isolated from different T-DNA mutants, one of these genes codes for a UTP-glucose-1-phosphate uridylyltransferase involved in programmed cell death and leaf development, which means a novel gene function reported in tomato. Together, our results support that enhancer trapping is a powerful tool to identify novel genes and regulatory elements in tomato and that this T-DNA mutant collection represents a highly valuable resource for functional analyses in this fleshy-fruited model species.


Asunto(s)
Elementos de Facilitación Genéticos , Genes de Plantas/genética , Genómica/métodos , Mutagénesis Insercional/métodos , Solanum lycopersicum/genética , Agrobacterium/genética , Secuencia de Bases , Mapeo Cromosómico , ADN Bacteriano/genética , ADN de Plantas/aislamiento & purificación , Frutas , Silenciador del Gen , Genes de Plantas/fisiología , Genes Reporteros , Fenotipo , Hojas de la Planta/crecimiento & desarrollo , Regiones Promotoras Genéticas
10.
Plant Cell Environ ; 40(5): 658-671, 2017 May.
Artículo en Inglés | MEDLINE | ID: mdl-27987209

RESUMEN

Excessive soil salinity diminishes crop yield and quality. In a previous study in tomato, we identified two closely linked genes encoding HKT1-like transporters, HKT1;1 and HKT1;2, as candidate genes for a major quantitative trait locus (kc7.1) related to shoot Na+ /K+ homeostasis - a major salt tolerance trait - using two populations of recombinant inbred lines (RILs). Here, we determine the effectiveness of these genes in conferring improved salt tolerance by using two near-isogenic lines (NILs) that were homozygous for either the Solanum lycopersicum allele (NIL17) or for the Solanum cheesmaniae allele (NIL14) at both HKT1 loci; transgenic lines derived from these NILs in which each HKT1;1 and HKT1;2 had been silenced by stable transformation were also used. Silencing of ScHKT1;2 and SlHKT1;2 altered the leaf Na+ /K+ ratio and caused hypersensitivity to salinity in plants cultivated under transpiring conditions, whereas silencing SlHKT1;1/ScHKT1;1 had a lesser effect. These results indicate that HKT1;2 has the more significant role in Na+ homeostasis and salinity tolerance in tomato.


Asunto(s)
Proteínas de Transporte de Catión/genética , Homeostasis , Proteínas de Plantas/genética , Brotes de la Planta/metabolismo , Potasio/metabolismo , Salinidad , Sodio/metabolismo , Solanum lycopersicum/genética , Solanum lycopersicum/metabolismo , Simportadores/genética , Alelos , Proteínas de Transporte de Catión/metabolismo , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Silenciador del Gen/efectos de los fármacos , Genes de Plantas , Sitios Genéticos , Homeostasis/efectos de los fármacos , Homeostasis/genética , Endogamia , Solanum lycopersicum/efectos de los fármacos , Solanum lycopersicum/crecimiento & desarrollo , Fenotipo , Hojas de la Planta/efectos de los fármacos , Hojas de la Planta/metabolismo , Proteínas de Plantas/metabolismo , Brotes de la Planta/efectos de los fármacos , Análisis de Componente Principal , Interferencia de ARN , ARN Mensajero/genética , ARN Mensajero/metabolismo , Cloruro de Sodio/farmacología , Simportadores/metabolismo
11.
Plant Mol Biol ; 91(4-5): 513-31, 2016 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-27125648

RESUMEN

Within the tomato MADS-box gene family, TOMATO AGAMOUS1 (TAG1) and ARLEQUIN/TOMATO AGAMOUS LIKE1 (hereafter referred to as TAGL1) are, respectively, members of the euAG and PLE lineages of the AGAMOUS clade. They perform crucial functions specifying stamen and carpel development in the flower and controlling late fruit development. To gain insight into the roles of TAG1 and TAGL1 genes and to better understand their functional redundancy and diversification, we characterized single and double RNAi silencing lines of these genes and analyzed expression profiles of regulatory genes involved in reproductive development. Double RNAi lines did show cell abnormalities in stamens and carpels and produced extremely small fruit-like organs displaying some sepaloid features. Expression analyses indicated that TAG1 and TAGL1 act together to repress fourth whorl sepal development, most likely through the MACROCALYX gene. Results also proved that TAG1 and TAGL1 have diversified their functions in fruit development: while TAG1 controls placenta and seed formation, TAGL1 participates in cuticle development and lignin biosynthesis inhibition. It is noteworthy that both TAG1 and double RNAi plants lacked seed development due to abnormalities in pollen formation. This seedless phenotype was not associated with changes in the expression of B-class stamen identity genes Tomato MADS-box 6 and Tomato PISTILLATA observed in silencing lines, suggesting that other regulatory factors should participate in pollen formation. Taken together, results here reported support the idea that both redundant and divergent functions of TAG1 and TAGL1 genes are needed to control tomato reproductive development.


Asunto(s)
Genes de Plantas , Proteínas de Dominio MADS/genética , Proteínas de Plantas/genética , Solanum lycopersicum/genética , Solanum lycopersicum/fisiología , Flores/genética , Flores/crecimiento & desarrollo , Regulación del Desarrollo de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Silenciador del Gen , Proteínas de Dominio MADS/metabolismo , Epidermis de la Planta/ultraestructura , Proteínas de Plantas/metabolismo , Plantas Modificadas Genéticamente , Reproducción/genética
12.
Plant Biotechnol J ; 14(6): 1345-56, 2016 06.
Artículo en Inglés | MEDLINE | ID: mdl-26578112

RESUMEN

A screening under salt stress conditions of a T-DNA mutant collection of tomato (Solanum lycopersicum L.) led to the identification of the altered response to salt stress 1 (ars1) mutant, which showed a salt-sensitive phenotype. Genetic analysis of the ars1 mutation revealed that a single T-DNA insertion in the ARS1 gene was responsible of the mutant phenotype. ARS1 coded for an R1-MYB type transcription factor and its expression was induced by salinity in leaves. The mutant reduced fruit yield under salt acclimation while in the absence of stress the disruption of ARS1 did not affect this agronomic trait. The stomatal behaviour of ars1 mutant leaves induced higher Na(+) accumulation via the transpiration stream, as the decreases of stomatal conductance and transpiration rate induced by salt stress were markedly lower in the mutant plants. Moreover, the mutation affected stomatal closure in a response mediated by abscisic acid (ABA). The characterization of tomato transgenic lines silencing and overexpressing ARS1 corroborates the role of the gene in regulating the water loss via transpiration under salinity. Together, our results show that ARS1 tomato gene contributes to reduce transpirational water loss under salt stress. Finally, this gene could be interesting for tomato molecular breeding, because its manipulation could lead to improved stress tolerance without yield penalty under optimal culture conditions.


Asunto(s)
Proteínas de Plantas/fisiología , Estomas de Plantas/fisiología , Cloruro de Sodio/metabolismo , Solanum lycopersicum/metabolismo , Factores de Transcripción/fisiología , Solanum lycopersicum/genética , Mutagénesis Insercional , Mutación , Fenotipo , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Transpiración de Plantas/genética , Alineación de Secuencia , Análisis de Secuencia de Proteína , Estrés Fisiológico , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Agua/metabolismo
13.
Plant Physiol ; 168(3): 1036-48, 2015 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-26019301

RESUMEN

Fruit development and ripening entail key biological and agronomic events, which ensure the appropriate formation and dispersal of seeds and determine productivity and yield quality traits. The MADS box gene Arlequin/tomato Agamous-like1 (hereafter referred to as TAGL1) was reported as a key regulator of tomato (Solanum lycopersicum) reproductive development, mainly involved in flower development, early fruit development, and ripening. It is shown here that silencing of the TAGL1 gene (RNA interference lines) promotes significant changes affecting cuticle development, mainly a reduction of thickness and stiffness, as well as a significant decrease in the content of cuticle components (cutin, waxes, polysaccharides, and phenolic compounds). Accordingly, overexpression of TAGL1 significantly increased the amount of cuticle and most of its components while rendering a mechanically weak cuticle. Expression of the genes involved in cuticle biosynthesis agreed with the biochemical and biomechanical features of cuticles isolated from transgenic fruits; it also indicated that TAGL1 participates in the transcriptional control of cuticle development mediating the biosynthesis of cuticle components. Furthermore, cell morphology and the arrangement of epidermal cell layers, on whose activity cuticle formation depends, were altered when TAGL1 was either silenced or constitutively expressed, indicating that this transcription factor regulates cuticle development, probably through the biosynthetic activity of epidermal cells. Our results also support cuticle development as an integrated event in the fruit expansion and ripening processes that characterize fleshy-fruited species such as tomato.


Asunto(s)
Frutas/genética , Genes de Plantas , Proteínas de Dominio MADS/genética , Epidermis de la Planta/crecimiento & desarrollo , Solanum lycopersicum/crecimiento & desarrollo , Solanum lycopersicum/genética , Transcripción Genética , Fenómenos Biomecánicos , Vías Biosintéticas/genética , Flores/anatomía & histología , Flores/fisiología , Frutas/citología , Frutas/crecimiento & desarrollo , Regulación del Desarrollo de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Proteínas de Dominio MADS/metabolismo , Fenotipo , Epidermis de la Planta/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Interferencia de ARN , ARN Mensajero/genética , ARN Mensajero/metabolismo
14.
Physiol Plant ; 155(3): 296-314, 2015 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-25582191

RESUMEN

Jasmonic acid (JA) regulates a wide spectrum of plant biological processes, from plant development to stress defense responses. The role of JA in plant response to salt stress is scarcely known, and even less known is the specific response in root, the main plant organ responsible for ionic uptake and transport to the shoot. Here we report the characterization of the first tomato (Solanum lycopersicum) mutant, named res (restored cell structure by salinity), that accumulates JA in roots prior to exposure to stress. The res tomato mutant presented remarkable growth inhibition and displayed important morphological alterations and cellular disorganization in roots and leaves under control conditions, while these alterations disappeared when the res mutant plants were grown under salt stress. Reciprocal grafting between res and wild type (WT) (tomato cv. Moneymaker) indicated that the main organ responsible for the development of alterations was the root. The JA-signaling pathway is activated in res roots prior to stress, with transcripts levels being even higher in control condition than in salinity. Future studies on this mutant will provide significant advances in the knowledge of JA role in root in salt-stress tolerance response, as well as in the energy trade-off between plant growth and response to stress.


Asunto(s)
Ciclopentanos/metabolismo , Mutación , Oxilipinas/metabolismo , Raíces de Plantas/metabolismo , Solanum lycopersicum/genética , Solanum lycopersicum/metabolismo , Regulación de la Expresión Génica de las Plantas , Solanum lycopersicum/citología , Células Vegetales/metabolismo , Células Vegetales/ultraestructura , Hojas de la Planta/fisiología , Raíces de Plantas/genética , Potasio/metabolismo , Salinidad , Tolerancia a la Sal/fisiología , Transducción de Señal
15.
Physiol Plant ; 152(4): 700-13, 2014 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-24773242

RESUMEN

For salt tolerance to be achieved in the long-term plants must regulate Na(+)/K(+) homeostasis over time. In this study, we show that the salt tolerance induced by overexpression of the yeast HAL5 gene in tomato (Solanum lycopersicum) was related to a lower leaf Na(+) accumulation in the long term, by reducing Na(+) transport from root to shoot over time regardless of the severity of salt stress. Furthermore, maintaining Na(+)/K(+) homeostasis over time was associated with changes in the transcript levels of the Na(+) and K(+) transporters such as SlHKT1;2 and SlHAK5. The expression of SlHKT1;2 was upregulated in response to salinity in roots of transgenic plants but downregulated in the roots of wild-type (WT) plants, which seems to be related to the lower Na(+) transport rate from root to shoot in transgenic plants. The expression of the SlHAK5 increased in roots and leaves of both WT and transgenic plants under salinity. However, this increase was much higher in the leaves of transgenic plants than in those of WT plants, which may be associated with the ability of transgenic leaves to maintain Na(+)/K(+) homeostasis over time. Taken together, the results show that the salt tolerance mechanism induced by HAL5 overexpression in tomato is related to the appropriate regulation of ion transport from root to shoot and maintenance of the leaf Na(+)/K(+) homeostasis through modulation of SlHKT1 and SlHAK5 over time.


Asunto(s)
Adaptación Fisiológica , Regulación de la Expresión Génica de las Plantas , Proteínas Quinasas/genética , Proteínas de Saccharomyces cerevisiae/genética , Cloruro de Sodio/farmacología , Solanum lycopersicum/fisiología , Frutas/genética , Frutas/fisiología , Expresión Génica , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Transporte Iónico , Solanum lycopersicum/genética , Hojas de la Planta/genética , Hojas de la Planta/fisiología , Raíces de Plantas/genética , Raíces de Plantas/fisiología , Brotes de la Planta/genética , Brotes de la Planta/fisiología , Plantas Modificadas Genéticamente , Potasio/metabolismo , Proteínas Quinasas/metabolismo , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Salinidad , Tolerancia a la Sal , Sodio/metabolismo , Transgenes , Xilema/genética , Xilema/fisiología
16.
Plant Biotechnol J ; 11(6): 770-9, 2013 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-23581527

RESUMEN

Fruit set and fruit development in tomato is largely affected by changes in environmental conditions, therefore autonomous fruit set independent of fertilization is a highly desirable trait in tomato. Here, we report the production and characterization of male-sterile transgenic plants that produce parthenocarpic fruits in two tomato cultivars (Micro-Tom and Moneymaker). We generated male-sterility using the cytotoxic gene barnase targeted to the anthers with the PsEND1 anther-specific promoter. The ovaries of these plants grew in the absence of fertilization producing seedless, parthenocarpic fruits. Early anther ablation is essential to trigger the developing of the transgenic ovaries into fruits, in the absence of the signals usually generated during pollination and fertilization. Ovaries are fully functional and can be manually pollinated to obtain seeds. The transgenic plants obtained in the commercial cultivar Moneymaker show that the parthenocarpic development of the fruit does not have negative consequences in fruit quality. Throughout metabolomic analyses of the tomato fruits, we have identified two elite lines which showed increased levels of several health promoting metabolites and volatile compounds. Thus, early anther ablation can be considered a useful tool to promote fruit set and to obtain seedless and good quality fruits in tomato plants. These plants are also useful parental lines to be used in hybrid breeding approaches.


Asunto(s)
Flores/metabolismo , Frutas/crecimiento & desarrollo , Partenogénesis , Solanum lycopersicum/crecimiento & desarrollo , Proteínas Bacterianas , Vías Biosintéticas/genética , Frutas/metabolismo , Regulación de la Expresión Génica de las Plantas , Genes de Plantas/genética , Genotipo , Giberelinas/metabolismo , Solanum lycopersicum/genética , Solanum lycopersicum/metabolismo , Metaboloma/genética , Plantas Modificadas Genéticamente , Ribonucleasas/metabolismo , Semillas/crecimiento & desarrollo , Semillas/metabolismo , Transformación Genética , Volatilización
17.
BMC Plant Biol ; 12: 156, 2012 Aug 31.
Artículo en Inglés | MEDLINE | ID: mdl-22935247

RESUMEN

BACKGROUND: Pelargonium is one of the most popular garden plants in the world. Moreover, it has a considerable economic importance in the ornamental plant market. Conventional cross-breeding strategies have generated a range of cultivars with excellent traits. However, gene transfer via Agrobacterium tumefaciens could be a helpful tool to further improve Pelargonium by enabling the introduction of new genes/traits. We report a simple and reliable protocol for the genetic transformation of Pelargonium spp. and the production of engineered long-life and male sterile Pelargonium zonale plants, using the pSAG12::ipt and PsEND1::barnase chimaeric genes respectively. RESULTS: The pSAG12::ipt transgenic plants showed delayed leaf senescence, increased branching and reduced internodal length, as compared to control plants. Leaves and flowers of the pSAG12::ipt plants were reduced in size and displayed a more intense coloration. In the transgenic lines carrying the PsEND1::barnase construct no pollen grains were observed in the modified anther structures, which developed instead of normal anthers. The locules of sterile anthers collapsed 3-4 days prior to floral anthesis and, in most cases, the undeveloped anther tissues underwent necrosis. CONCLUSION: The chimaeric construct pSAG12::ipt can be useful in Pelargonium spp. to delay the senescence process and to modify plant architecture. In addition, the use of engineered male sterile plants would be especially useful to produce environmentally friendly transgenic plants carrying new traits by preventing gene flow between the genetically modified ornamentals and related plant species. These characteristics could be of interest, from a commercial point of view, both for pelargonium producers and consumers.


Asunto(s)
Ingeniería Genética/métodos , Pelargonium/genética , Infertilidad Vegetal , Plantas Modificadas Genéticamente/fisiología , Agrobacterium tumefaciens/genética , Proteínas Bacterianas , Flores/genética , Flores/fisiología , Pelargonium/fisiología , Hojas de la Planta/genética , Hojas de la Planta/fisiología , Técnicas de Embriogénesis Somática de Plantas , Plantas Modificadas Genéticamente/genética , Ribonucleasas/genética , Transformación Genética
18.
Plant Biotechnol J ; 10(3): 341-52, 2012 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-22070155

RESUMEN

Edible fruits are inexpensive biofactories for human health-promoting molecules that can be ingested as crude extracts or partially purified formulations. We show here the production of a model human antibody for passive protection against the enteric pathogen rotavirus in transgenically labelled tomato fruits. Transgenic tomato plants expressing a recombinant human immunoglobulin A (hIgA_2A1) selected against the VP8* peptide of rotavirus SA11 strain were obtained. The amount of hIgA_2A1 protein reached 3.6 ± 0.8% of the total soluble protein in the fruit of the transformed plants. Minimally processed fruit-derived products suitable for oral intake showed anti-VP8* binding activity and strongly inhibited virus infection in an in vitro virus neutralization assay. In order to make tomatoes expressing hIgA_2A1 easily distinguishable from wild-type tomatoes, lines expressing hIgA_2A1 transgenes were sexually crossed with a transgenic tomato line expressing the genes encoding Antirrhinum majus Rosea1 and Delila transcription factors, which confer purple colour to the fruit. Consequently, transgenically labelled purple tomato fruits expressing hIgA_2A1 have been developed. The resulting purple-coloured extracts from these fruits contain high levels of recombinant anti-rotavirus neutralizing human IgA in combination with increased amounts of health-promoting anthocyanins.


Asunto(s)
Anticuerpos Neutralizantes/inmunología , Plantas Modificadas Genéticamente/inmunología , Proteínas de Unión al ARN/inmunología , Rotavirus/inmunología , Solanum lycopersicum/inmunología , Proteínas no Estructurales Virales/inmunología , Agrobacterium tumefaciens/genética , Agrobacterium tumefaciens/metabolismo , Antocianinas/metabolismo , Anticuerpos Monoclonales/inmunología , Anticuerpos Antivirales/inmunología , Antirrhinum/genética , Western Blotting , Cruzamientos Genéticos , Vectores Genéticos/genética , Vectores Genéticos/metabolismo , Humanos , Inmunoglobulina A/inmunología , Fragmentos de Inmunoglobulinas/genética , Fragmentos de Inmunoglobulinas/inmunología , Solanum lycopersicum/genética , Solanum lycopersicum/virología , Pruebas de Neutralización , Pigmentación , Extractos Vegetales/inmunología , Plantas Modificadas Genéticamente/genética , Plantas Modificadas Genéticamente/virología , Plásmidos/genética , Plásmidos/metabolismo , Proteínas de Unión al ARN/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/inmunología , Proteínas Recombinantes/metabolismo , Coloración y Etiquetado , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Transgenes , Proteínas no Estructurales Virales/genética
19.
Plant Physiol ; 157(4): 1650-63, 2011 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-21972266

RESUMEN

Fruit of tomato (Solanum lycopersicum), like those from many species, have been characterized to undergo a shift from partially photosynthetic to truly heterotrophic metabolism. While there is plentiful evidence for functional photosynthesis in young tomato fruit, the rates of carbon assimilation rarely exceed those of carbon dioxide release, raising the question of its role in this tissue. Here, we describe the generation and characterization of lines exhibiting a fruit-specific reduction in the expression of glutamate 1-semialdehyde aminotransferase (GSA). Despite the fact that these plants contained less GSA protein and lowered chlorophyll levels and photosynthetic activity, they were characterized by few other differences. Indeed, they displayed almost no differences in fruit size, weight, or ripening capacity and furthermore displayed few alterations in other primary or intermediary metabolites. Although GSA antisense lines were characterized by significant alterations in the expression of genes associated with photosynthesis, as well as with cell wall and amino acid metabolism, these changes were not manifested at the phenotypic level. One striking feature of the antisense plants was their seed phenotype: the transformants displayed a reduced seed set and altered morphology and metabolism at early stages of fruit development, although these differences did not affect the final seed number or fecundity. Taken together, these results suggest that fruit photosynthesis is, at least under ambient conditions, not necessary for fruit energy metabolism or development but is essential for properly timed seed development and therefore may confer an advantage under conditions of stress.


Asunto(s)
Frutas/crecimiento & desarrollo , Fotosíntesis/fisiología , Proteínas de Plantas/metabolismo , Semillas/crecimiento & desarrollo , Solanum lycopersicum/crecimiento & desarrollo , Ácido Aminolevulínico/metabolismo , Frutas/genética , Frutas/metabolismo , Frutas/fisiología , Perfilación de la Expresión Génica , Regulación de la Expresión Génica de las Plantas/fisiología , Glucuronidasa , Solanum lycopersicum/genética , Solanum lycopersicum/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Especificidad de Órganos , Fenotipo , Proteínas de Plantas/genética , Plantas Modificadas Genéticamente , Regiones Promotoras Genéticas/genética , Reproducción , Semillas/genética , Semillas/metabolismo
20.
Plant Cell Rep ; 30(10): 1865-79, 2011 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-21647638

RESUMEN

Salinity and drought have a huge impact on agriculture since there are few areas free of these abiotic stresses and the problem continues to increase. In tomato, the most important horticultural crop worldwide, there are accessions of wild-related species with a high degree of tolerance to salinity and drought. Thus, the finding of insertional mutants with other tolerance levels could lead to the identification and tagging of key genes responsible for abiotic stress tolerance. To this end, we are performing an insertional mutagenesis programme with an enhancer trap in the tomato wild-related species Solanum pennellii. First, we developed an efficient transformation method which has allowed us to generate more than 2,000 T-DNA lines. Next, the collection of S. pennelli T(0) lines has been screened in saline or drought conditions and several presumptive mutants have been selected for their salt and drought sensitivity. Moreover, T-DNA lines with expression of the reporter uidA gene in specific organs, such as vascular bundles, trichomes and stomata, which may play key roles in processes related to abiotic stress tolerance, have been identified. Finally, the growth of T-DNA lines in control conditions allowed us the identification of different development mutants. Taking into account that progenies from the lines are being obtained and that the collection of T-DNA lines is going to enlarge progressively due to the high transformation efficiency achieved, there are great possibilities for identifying key genes involved in different tolerance mechanisms to salinity and drought.


Asunto(s)
Mutagénesis Insercional/métodos , Solanum/genética , Estrés Fisiológico , ADN Bacteriano/genética , Sequías , Regulación de la Expresión Génica de las Plantas , Genes de Plantas , Ensayos Analíticos de Alto Rendimiento , Fenotipo , Salinidad , Plantas Tolerantes a la Sal/genética , Plantas Tolerantes a la Sal/fisiología , Solanum/fisiología , Transformación Genética
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