Characterization of a new tomato 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 (LeNHX3 and LeNHX4), SALT OVERLY SENSITIVE (SlSOS1 and SlSOS2), HIGH-AFFINITY K+ TRANSPORTER 1;2 (SlHKT1;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 (SlCAX1) 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 orthologue of the Arabidopsis 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.

Background: Nuña bean is a type of ancient common bean (Phaseolus vulgaris L.) native to the Andean region of South America, whose seeds possess the unusual property of popping. The nutritional features of popped seeds make them a healthy low fat and high protein snack. However, flowering of nuña bean only takes place under short-day photoperiod conditions, which means a difficulty to extend production to areas where such conditions do not prevail. Therefore, breeding programs of adaptation traits will facilitate the diversification of the bean crops and the development of new varieties with enhanced healthy properties. Although the popping trait has been profusely studied in maize (popcorn), little is known about the biology and genetic basis of the popping ability in common bean. To obtain insights into the genetics of popping ability related traits of nuña bean, a comprehensive quantitative trait loci (QTL) analysis was performed to detect single-locus and epistatic QTLs responsible for the phenotypic variance observed in these traits. Results: A mapping population of 185 recombinant inbred lines (RILs) derived from a cross between two Andean common bean genotypes was evaluated for three popping related traits, popping dimension index (PDI), expansion coefficient (EC), and percentage of unpopped seeds (PUS), in five different environmental conditions. The genetic map constructed included 193 loci across 12 linkage groups (LGs), covering a genetic distance of 822.1 cM, with an average of 4.3 cM per marker. Individual and multi-environment QTL analyses detected a total of nineteen single-locus QTLs, highlighting among them the co-localized QTLs for the three popping ability traits placed on LGs 3, 5, 6, and 7, which together explained 24.9, 14.5, and 25.3% of the phenotypic variance for PDI, EC, and PUS, respectively. Interestingly, epistatic interactions among QTLs have been detected, which could have a key role in the genetic control of popping. Conclusions: The QTLs here reported constitute useful tools for marker assisted selection breeding programs aimed at improving nuña bean cultivars, as well as for extending our knowledge of the genetic determinants and genotype x environment interaction involved in the popping ability traits of this bean crop.

Tomato (Solanum lycopersicum L.) is a major horticultural crop and a model species among eudicots, especially for traits related to reproductive development. Although considerable progress has been made since the tomato genome sequence project was completed, most of the genes identified remain predictions with an unknown or hypothetical function. This lack of functional characterization hampers the use of the huge amount of genomic information available to improve the quality and productivity of this crop. Reverse genetics strategies such as artificial mutagenesis and next-generation sequencing approaches build the perfect tandem for increasing knowledge on functional annotation of tomato genes. This work reports the phenotypic characterization of a tomato mutant collection generated from an EMS chemical mutagenesis program aimed to identify interesting agronomic mutants and novel gene functions. Tomato mutants were grouped into fourteen phenotypic classes, including vegetative and reproductive development traits, and the inheritance pattern of the identified mutations was studied. In addition, causal mutation of a selected mutant line was isolated through a mapping-by-sequencing approach as a proof of concept of this strategy’s successful implementation. Results support tomato mutagenesis as an essential tool for functional genomics in this fleshy-fruited model species and a highly valuable resource for future breeding programs of this crop species aimed at the development of more productive and resilient new varieties under challenging climatic and production scenarios.

The res (restored cell structure by salinity) mutant, recently identified as the first tomato mutant accumulating jasmonate in roots under non-stressful conditions, exhibits a remarkable growth inhibition and morphological alterations in roots and leaves, which are suppressed when the mutant plants are exposed to salinity. In order to understand the molecular basis of the phenotype recovery induced by salt stress in the res mutant, we carried out a comparative transcriptomic analysis in roots and leaves of wild-type and res plants in absence of stress (control) and when the phenotypic recovery of res mutant began to be observed upon salt stress (5 days of 200 mM NaCl). The number of differentially expressed genes was three times greater in roots than in leaves of res vs WT plants grown in control, and included the down-regulation of growth-promoting genes and the up-regulation of genes involved in Ca2+ signalling, transcription factors and others related to stress responses. However, these expression differences were attenuated under salt stress, coinciding with the phenotypic normalisation of the mutant. Contrarily to the attenuated response observed in roots, an enhanced response was found in leaves under salt stress. This included drastic expression changes in several circadian clock genes, such as GIGANTEA1, which was down-regulated in res vs WT plants. Moreover, the higher photosynthetic efficiency of res leaves under salt stress was accompanied by specific salt-upregulation of the genes RUBISCO ACTIVASE1 and ALTERNATIVE OXIDASE1A. Very few genes were found to be differentially expressed in both tissues (root and leaf) and conditions (control and salt), but this group included SlWRKY39 and SlMYB14 transcription factors, as well as genes related to protein homeostasis, especially protease inhibitors such as METALLOCARBOXYPEPTIDASE INHIBITOR, which also seem to play a role in the phenotype recovery and salt tolerance of res mutant. In summary, in this study we have identified genes which seem to have a prominent role in salt tolerance. Moreover, we think this work could contribute to future breeding of tomato crops with increased stress tolerance.

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. 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. 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.

Flower development is a crucial step towards the completion of the plant life cycle. Physiological processes and gene regulatory mechanisms underlying flower formation have been extensively characterized, and the implication of MADS-box transcription factors as primary regulators of flower morphology has been widely described, mainly due to the analysis of loss-of-function mutants in model species. Nevertheless, detailed characterization of allele variation in several MADS-box homologous genes from crop species remains undescribed. Here, we have characterized a tomato mutant with aberrant flower development. Mutant plants exhibit changes in petal cell identity, as well as homeotic transformations of stamens into carpelloid structures, which in most cases result in succulent organs. Molecular analysis proved that a loss-of-function mutation in the TOMATO MADS-BOX 6 (TM6) gene is responsible for this mutant phenotype. Furthermore, as a result of the loss of function of TM6, misregulation of the transcription and mRNA processing of other MADS-box genes involved in reproductive development has been detected. Our findings demonstrate that TM6 is a key player in the complex regulatory network of MADS-box genes controlling flower development and also provide a novel mutant that may be useful for generating male sterile lines in tomatoes.