Identification of novel stress-responsive biomarkers from gene expression datasets in tomato roots

Published by CSIRO Publishing. This is the Author Accepted Manuscript. This article may be used for personal use only.Abiotic stresses such as heat, drought or salinity have been widely studied individually. Nevertheless, in the nature and in the field, plants and crops are commonly exposed to a different combination of stresses, which often result in a synergistic response mediated by the activation of several molecular pathways that cannot be inferred from the response to each individual stress. By screening microarray data obtained from different plant species and under different stresses, we identified several conserved stress-responsive genes whose expression was differentially regulated in tomato (Solanum lycopersicum L.) roots in response to one or several stresses. We validated 10 of these genes as reliable biomarkers whose expression levels are related to different signalling pathways involved in adaptive stress responses. In addition, the genes identified in this work could be used as general salt-stress biomarkers to rapidly evaluate the response of salt-tolerant cultivars and wild species for which sufficient genetic information is not yet available

Is it feasible to use mixed orchards to manage apple scab?

Apple scab, caused by the fungus Venturia inaequalis, is one of the most damaging diseases of cultivated apples (Malus x domestica) worldwide. It results in huge losses as it diminishes fruit quality and impacts tree growth. Current management revolves around application of fungicides, however the number of sprays required per season is unsustainable and expensive. Further to this, populations of V. inaequalis have developed fungicide resistance. Breeding new cultivars with higher levels of resistance to scab is a priority, however, this process is long, so introducing mixed cultivar orchards may be a faster solution. We reviewed the general principles of using mixtures to manage plant diseases, and then considered specifically using mixed cultivars to manage apple scab in commercial production. Limited field studies have demonstrated the potential of using mixture to suppress apple scab development; but scab super-races that could emerge from mixture can pose a significant risk to in commercial production. However, recent research on population genetics of apple scab in monoculture and mixed orchards suggests that the risk of super-race emergence is probably over-stated because assortative mating among lesions on the same leaves is likely to occur rather than commonly assumed random mating. Thus, we conclude that cultivar mixtures can contribute towards sustainable scab management, particularly in commercial cider apple production.Biotechnology and Biological Sciences Research Council (BBSRC) Collaborative Training Partnership for Fruit Crop Research (CTP-FCR

BIFURCATE FLOWER TRUSS: a novel locus controlling inflorescence branching in tomato contains a defective MAP kinase gene

A mutant line, bifurcate flower truss (bif), was recovered from a tomato breeding program. Plants from the control line LAM183 produced a mean of 0.16 branches per truss, whereas the value for bif plants was 4.1. This increase in branching was accompanied by a 3.3-fold increase in flower number and showed a significant interaction with exposure to low temperature during truss development. The LAM183 and bif genomes were resequenced and the bif gene was mapped to a 2.01 Mbp interval on chromosome 12; all coding region polymorphisms in the interval were surveyed and five candidate genes displaying altered protein sequences were detected. One of these genes, SlMAPK1, encoding a MAP kinase, contained a leucine-to-stop codon mutation predicted to disrupt kinase function. SlMAPK1 is an excellent candidate for bif because knock-out mutations of an Arabidopsis orthologue MPK6 were reported to have increased flower number. An introgression browser was used to demonstrate that the origin of the bif genomic DNA at the BIF locus was Solanum galapagense and that the SlMAPK1 null mutant is a naturally occurring allele widespread only on the Galápagos Islands. This work strongly implicates SlMAPK1 as part of the network of genes controlling inflorescence branching in tomato

Missense mutation of a class B heat shock factor is responsible for the tomato bushy root-2 phenotype

The bushy root-2 (brt-2) tomato mutant has twisting roots, and slower plant development. Here we used whole genome resequencing and genetic mapping to show that brt-2 is caused by a serine to cysteine (S75C) substitution in the DNA binding domain (DBD) of a heat shock factor class B (HsfB) encoded by SolycHsfB4a. This gene is orthologous to the Arabidopsis SCHIZORIZA gene, also known as AtHsfB4. The brt-2 phenotype is very similar to Arabidopsis lines in which the function of AtHsfB4 is altered: a proliferation of lateral root cap and root meristematic tissues, and a tendency for lateral root cap cells to easily separate. The brt-2 S75C mutation is unusual because all other reported amino acid substitutions in the highly conserved DBD of eukaryotic heat shock factors are dominant negative mutations, but brt-2 is recessive. We further show through reciprocal grafting that brt-2 exerts its effects predominantly through the root genotype even through BRT-2 is expressed at similar levels in both root and shoot meristems. Since AtHsfB4 is induced by root knot nematodes (RKN), and loss-of-function mutants of this gene are resistant to RKNs, BRT-2 could be a target gene for RKN resistance, an important trait in tomato rootstock breeding.Biotechnology and Biological Sciences Research Council (BBSRC): BB/L01954X/

Improving the tea withering process using ethylene or UV-C

Using a combination of biochemical, transcriptomic, and physiological analyses, we elucidated the mechanisms of physical and chemical withering of tea shoots subjected to UV-C and ethylene treatments. UV-C irradiation (15 kJ m–2) initiated oxidation of catechins into theaflavins, increasing theaflavin-3-monogallate and theaflavin digallate by 5- and 13.2–4.4-fold, respectively, at the end of withering. Concomitantly, a rapid change to brown/red, an increase in electrolyte leakage, and the upregulation of peroxidases (viz. Px2, Px4, and Px6) and polyphenol oxidases (PPO-1) occurred. Exogenous ethylene significantly increased the metabolic rate (40%) and moisture loss (30%) compared to control during simulated withering (12 h at 25 °C) and upregulated transcripts associated with responses to dehydration and abiotic stress, such as those in the ethylene signaling pathway (viz. EIN4-like, EIN3-FBox1, and ERFs). Incorporating ethylene during withering could shorten the tea manufacturing process, while UV-C could enhance the accumulation of flavor-related compounds

ABA-overproduction response under salinity

[SPA] Con el fin de comprender la influencia de la fitohormona ácido abscísico (ABA) en la adaptación al riego salino, dos líneas transgénicas independientes de tomate (Solanum lycopersicum L.), sp12 y sp5, que sobreexpresan constitutivamente el gen NCED1 (codifica para la enzima que cataliza un paso limitante en la biosíntesis de ABA) y la variedad silvestre Ailsa Craig, se han estudiado en experimentos o bien i) como planta entera o ii) como portainjerto bajo condiciones control y de estrés salino. Aunque la expresión constitutiva de NCED disminuye el crecimiento bajo condiciones control, minimiza los efectos producidos por la sal (planta completa) y mejora significativamente el crecimiento cuando se usa como portainjerto. El análisis de la savia xilemática de raíz mostró que los fenotipos resultantes bajo las diferentes condiciones de cultivo eran difíciles de explicar en términos de sobreproducción de ABA. Para intentar explicar estos resultados se llevó a cabo un análisis de expresión de un conjunto de genes relacionados con hormonas y estrés mediante PCR cuantitativa, así como un estudio transcriptómico mediante microarrays en la raíz. Los resultados sugieren que la sobreexpresión de NCED parece alterar diversas rutas de señalización, derivando en una respuesta adaptativa al estrés que podría ayudar a explicar los fenotipos observados. [ENG] With the aim of better understanding the influence of the plan hormone abscisic acid (ABA) in adaptation to saline irrigation, two independent transgenic tomato (Solanum lycopersicum L.) lines, sp12 and sp5, overexpressing constitutively NCED1 (the enzyme that catalyzes a key rate-limiting step in ABA biosynthesis) and the wild type Ailsa Craig, have been studied in experiments either i) as whole plants or ii) as rootstocks under control and salinity conditions. While NCED overexpression penalizes growth under control conditions, it minimized the effect of salinity (whole plants) or significantly improved plant growth and yield when used as rootstocks. The analysis of the root xylem sap revealed that the phenotypes resulting under the different conditions were difficult to explain in terms of ABA overproduction. With the aim of explaining these results, the expression of a set of hormone and stress associated genes (analysed by real time PCR) as well as a transcriptomic analysis (by using one-color microarray) were performed in roots. The results suggest that NCED overexpression seems to alter several signalling pathways leading to stress adaptive responses that could help to explain the observed phenotypes.The authors thank Andrew J. Thompson from Cranfield University, the NCED seeds set. This work was supported by CICYT-FEDER (project AGL2011-27996) and European Union’s Seventh Framework Programme for research, technological development and demonstration under grant agreement no 289365(ROOTOPOWER project)

Genes involved in auxin biosynthesis, transport and signalling underlie the extreme adventitious root phenotype of the tomato aer mutant

The use of tomato rootstocks has helped to alleviate the soaring abiotic stresses provoked by the adverse effects of climate change. Lateral and adventitious roots can improve topsoil exploration and nutrient uptake, shoot biomass and resulting overall yield. It is essential to understand the genetic basis of root structure development and how lateral and adventitious roots are produced. Existing mutant lines with specific root phenotypes are an excellent resource to analyse and comprehend the molecular basis of root developmental traits. The tomato aerial roots (aer) mutant exhibits an extreme adventitious rooting phenotype on the primary stem. It is known that this phenotype is associated with restricted polar auxin transport from the juvenile to the more mature stem, but prior to this study, the genetic loci responsible for the aer phenotype were unknown. We used genomic approaches to define the polygenic nature of the aer phenotype and provide evidence that increased expression of specific auxin biosynthesis, transport and signalling genes in different loci causes the initiation of adventitious root primordia in tomato stems. Our results allow the selection of different levels of adventitious rooting using molecular markers, potentially contributing to rootstock breeding strategies in grafted vegetable crops, especially in tomato. In crops vegetatively propagated as cuttings, such as fruit trees and cane fruits, orthologous genes may be useful for the selection of cultivars more amenable to propagation.The research was supported by BBSRC—UKRI funding; the RootLINK (BB/L01954X/1) and AdRoot (BB/S007970/1) projects

A loss-of-function allele of a TAC1-like gene (SlTAC1) located on tomato chromosome 10 is a candidate for the Erectoid leaf (Erl) mutation

The genetic basis of an erectoid leaf phenotype was investigated in distinct tomato breeding populations, including one derived from Solanum lycopersicum ‘LT05’ (with the erectoid leaf phenotype and uniform ripening, genotype uu) × S. pimpinellifollium ‘TO-937’ (with the wild-type leaf phenotype and green fruit shoulder, genotype UU). The erectoid leaf phenotype was inherited as a semi-dominant trait and it co-segregated with the u allele of gene SlGLK2 (Solyc10g008160). This genomic location coincides with a previously described semi-dominant mutation named as Erectoid leaf (Erl). The genomes of ‘LT05’, ‘TO-937’, and three other unrelated accessions (with the wild-type Erl+ allele) were resequenced with the aim of identifying candidate genes. Comparative genomic analyses, including the reference genome ‘Heinz 1706’ (Erl+ allele), identified an Erectoid leaf-specific single nucleotide polymorphism (SNP) in the gene Solyc10g009320. This SNP caused a change of a glutamine codon (present in all the wild-type genomes) to a TAA (= ochre stop-codon) in the Erl allele, resulting in a smaller version of the predicted mutant protein (221 vs. 279 amino acids). Solyc10g009320, previously annotated as an ‘unknown protein’, was identified as a TILLER ANGLE CONTROL1-like gene. Linkage between the Erl and Solyc10g009320 was confirmed via Sanger sequencing of the PCR amplicons of the two variant alleles. No recombinants were detected in 265 F2 individuals. Contrasting S7 near-isogenic lines were also homozygous for each of the alternate alleles, reinforcing Solyc10g009320 as a strong Erl candidate gene and opening the possibility for fine-tuning manipulation of tomato architecture in breeding programs

A chromosome-level genome assembly of Solanum chilense, a tomato wild relative associated with resistance to salinity and drought

Introduction: Solanum chilense is a wild relative of tomato reported to exhibit resistance to biotic and abiotic stresses. There is potential to improve tomato cultivars via breeding with wild relatives, a process greatly accelerated by suitable genomic and genetic resources. Methods: In this study we generated a high-quality, chromosome-level, de novo assembly for the S. chilense accession LA1972 using a hybrid assembly strategy with ~180 Gbp of Illumina short reads and ~50 Gbp long PacBio reads. Further scaffolding was performed using Bionano optical maps and 10x Chromium reads. Results: The resulting sequences were arranged into 12 pseudomolecules using Hi-C sequencing. This resulted in a 901 Mbp assembly, with a completeness of 95%, as determined by Benchmarking with Universal Single-Copy Orthologs (BUSCO). Sequencing of RNA from multiple tissues resulting in ~219 Gbp of reads was used to annotate the genome assembly with an RNA-Seq guided gene prediction, and for a de novo transcriptome assembly. This chromosome-level, high-quality reference genome for S. chilense accession LA1972 will support future breeding efforts for more sustainable tomato production. Discussion: Gene sequences related to drought and salt resistance were compared between S. chilense and S. lycopersicum to identify amino acid variations with high potential for functional impact. These variants were subsequently analysed in 84 resequenced tomato lines across 12 different related species to explore the variant distributions. We identified a set of 7 putative impactful amino acid variants some of which may also impact on fruit development for example the ethylene-responsive transcription factor WIN1 and ethylene-insensitive protein 2. These variants could be tested for their ability to confer functional phenotypes to cultivars that have lost these variants.This work was jointly supported by the UK’s Biotechnology and Biological Sciences Research Council and the Indian Department of Biotechnology (BB/L011611/1)

A chromosome-level genome assembly of Solanum chilense, a tomato wild relative associated with resistance to salinity and drought

IntroductionSolanum chilense is a wild relative of tomato reported to exhibit resistance to biotic and abiotic stresses. There is potential to improve tomato cultivars via breeding with wild relatives, a process greatly accelerated by suitable genomic and genetic resources.MethodsIn this study we generated a high-quality, chromosome-level, de novo assembly for the S. chilense accession LA1972 using a hybrid assembly strategy with ~180 Gbp of Illumina short reads and ~50 Gbp long PacBio reads. Further scaffolding was performed using Bionano optical maps and 10x Chromium reads. ResultsThe resulting sequences were arranged into 12 pseudomolecules using Hi-C sequencing. This resulted in a 901 Mbp assembly, with a completeness of 95%, as determined by Benchmarking with Universal Single-Copy Orthologs (BUSCO). Sequencing of RNA from multiple tissues resulting in ~219 Gbp of reads was used to annotate the genome assembly with an RNA-Seq guided gene prediction, and for a de novo transcriptome assembly. This chromosome-level, high-quality reference genome for S. chilense accession LA1972 will support future breeding efforts for more sustainable tomato production. DiscussionGene sequences related to drought and salt resistance were compared between S. chilense and S. lycopersicum to identify amino acid variations with high potential for functional impact. These variants were subsequently analysed in 84 resequenced tomato lines across 12 different related species to explore the variant distributions. We identified a set of 7 putative impactful amino acid variants some of which may also impact on fruit development for example the ethylene-responsive transcription factor WIN1 and ethylene-insensitive protein 2. These variants could be tested for their ability to confer functional phenotypes to cultivars that have lost these variants