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A proteome analysis of freezing tolerance in red clover (Trifolium pratense L.).

ABSTRACT: Improvement of freezing tolerance of red clover (Trifolium pratense L.) would increase its persistence under cold climate. In this study, we assessed the freezing tolerance and compared the proteome composition of non-acclimated and cold-acclimated plants of two initial cultivars of red clover: Endure (E-TF0) and Christie (C-TF0) and of populations issued from these cultivars after three (TF3) and four (TF4) cycles of phenotypic recurrent selection for superior freezing tolerance. Through this approach, we wanted to identify proteins that are associated with the improvement of freezing tolerance in red clover.Freezing tolerance expressed as the lethal temperature for 50 % of the plants (LT50) increased markedly from approximately -2 to -16 °C following cold acclimation. Recurrent selection allowed a significant 2 to 3 °C increase of the LT50 after four cycles of recurrent selection. Two-dimensional difference gel electrophoresis (2D-DIGE) was used to study variations in protein abundance. Principal component analysis based on 2D-DIGE revealed that the largest variability in the protein data set was attributable to the cold acclimation treatment and that the two genetic backgrounds had differential protein composition in the acclimated state only. Vegetative storage proteins (VSP), which are essential nitrogen reserves for plant regrowth, and dehydrins were among the most striking changes in proteome composition of cold acclimated crowns of red clovers. A subset of proteins varied in abundance in response to selection including a dehydrin that increased in abundance in TF3 and TF4 populations as compared to TF0 in the Endure background.Recurrent selection performed indoor is an effective approach to improve the freezing tolerance of red clover. Significant improvement of freezing tolerance by recurrent selection was associated with differential accumulation of a small number of cold-regulated proteins that may play an important role in the determination of the level of freezing tolerance.

SUBMITTER: Bertrand A

PROVIDER: S-EPMC4787020 | biostudies-literature | 2016 Mar

REPOSITORIES: biostudies-literature

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A proteome analysis of freezing tolerance in red clover (Trifolium pratense L.).

Bertrand Annick A Bipfubusa Marie M Castonguay Yves Y Rocher Solen S Szopinska-Morawska Aleksandra A Papadopoulos Yousef Y Renaut Jenny J

BMC plant biology 20160310

<h4>Background</h4>Improvement of freezing tolerance of red clover (Trifolium pratense L.) would increase its persistence under cold climate. In this study, we assessed the freezing tolerance and compared the proteome composition of non-acclimated and cold-acclimated plants of two initial cultivars of red clover: Endure (E-TF0) and Christie (C-TF0) and of populations issued from these cultivars after three (TF3) and four (TF4) cycles of phenotypic recurrent selection for superior freezing tolera ...[more]

PMID: 26965047

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Project description:Improvement of persistency is an important breeding goal in red clover (Trifolium pratense L.). In areas with cold winters, lack of persistency is often due to poor winter survival, of which low freezing tolerance (FT) is an important component. We conducted a genome wide association study (GWAS) to identify loci associated with freezing tolerance in a collection of 393 red clover accessions, mostly of European origin, and performed analyses of linkage disequilibrium and inbreeding. Accessions were genotyped as pools of individuals using genotyping-by-sequencing (pool-GBS), generating both single nucleotide polymorphism (SNP) and haplotype allele frequency data at accession level. Linkage disequilibrium was determined as a squared partial correlation between the allele frequencies of pairs of SNPs and found to decay at extremely short distances (< 1 kb). The level of inbreeding, inferred from the diagonal elements of a genomic relationship matrix, varied considerably between different groups of accessions, with the strongest inbreeding found among ecotypes from Iberia and Great Britain, and the least found among landraces. Considerable variation in FT was found, with LT50-values (temperature at which 50% of the plants are killed) ranging from -6.0°C to -11.5°C. SNP and haplotype-based GWAS identified eight and six loci significantly associated with FT (of which only one was shared), explaining 30% and 26% of the phenotypic variation, respectively. Ten of the loci were found within or at a short distance (<0.5 kb) from genes possibly involved in mechanisms affecting FT. These include a caffeoyl shikimate esterase, an inositol transporter, and other genes involved in signaling, transport, lignin synthesis and amino acid or carbohydrate metabolism. This study paves the way for a better understanding of the genetic control of FT and for the development of molecular tools for the improvement of this trait in red clover through genomics assisted breeding.

Project description:Red clover (Trifolium pratense L.) is an important cool-season legume plant, which is used as forage. Leaf senescence is a critical developmental process that negatively affects plant quality and yield. The regulatory mechanism of leaf senescence has been studied, and genes involved in leaf senescence have been cloned and characterized in many plants. However, those works mainly focused on model plants. Information about regulatory pathways and the genes involved in leaf senescence in red clover is very sparse. In this study, to better understand leaf senescence in red clover, transcriptome analysis of mature and senescent leaves was investigated using RNA-Seq. A total of about 35,067 genes were identified, and 481 genes were differentially expressed in mature and senescent leaves. Some identified differentially expressed genes showed similar expression patterns as those involved in leaf senescence in other species, such as Arabidopsis, Medicago truncatula and rice. Differentially expressed genes were confirmed by quantitative real-time PCR (qRT-PCR). Genes involved in signal transduction, transportation and metabolism of plant hormones, transcription factors and plant senescence were upregulated, while the downregulated genes were primarily involved in nutrient cycling, lipid/carbohydrate metabolism, hormone response and other processes. There were 64 differentially expressed transcription factor genes identified by RNA-Seq, including ERF, WRKY, bHLH, MYB and NAC. A total of 90 genes involved in biosynthesis, metabolism and transduction of plant hormones, including abscisic acid, jasmonic acid, cyokinin, brassinosteroid, salicylic acid and ethylene, were identified. Furthermore, 207 genes with direct roles in leaf senescence were demonstrated, such as senescence-associated genes. These genes were associated with senescence in other plants. Transcriptome analysis of mature and senescent leaves in red clover provides a large number of differentially expressed genes. Further analysis and identification of senescence-associated genes can provide new insight into the regulatory mechanisms of leaf development and senescence in legume plant and red clover.

Project description:IntroductionRed clover (Trifolium pratense) is a protein-rich, short-lived perennial forage crop that can achieve high yields, but suffers increasingly from drought in different cultivation areas. Breeding for increased adaptation to drought is becoming essential, but at this stage it is unclear which traits breeders should target to phenotype responses to drought that allow them to identify the most promising red clover genotypes. In this study, we assessed how prolonged periods of drought affected plant growth in field conditions, and which traits could be used to distinguish better adapted plant material.MethodsA diverse panel of 395 red clover accessions was evaluated during two growing seasons. We simulated 6-to-8-week drought periods during two consecutive summers, using mobile rain-out shelters, while an irrigated control field was established in an adjacent parcel. Plant growth was monitored throughout both growing seasons using multiple flights with a drone equipped with RGB and thermal sensors. At various observation moments throughout both growing seasons, we measured canopy cover (CC) and canopy height (CH). The crop water stress index (CWSI) was determined at two moments, during or shortly after the drought event.ResultsManual and UAV-derived measurements for CH were well correlated, indicating that UAV-derived measurements can be reliably used in red clover. In both years, CC, CH and CWSI were affected by drought, with measurable growth reductions by the end of the drought periods, and during the recovery phase. We found that the end of the drought treatment and the recovery phase of approximately 20 days after drought were suitable periods to phenotype drought responses and to distinguish among genotypes.DiscussionMultifactorial analysis of accession responses revealed interactions of the maturity type with drought responses, which suggests the presence of two independent strategies in red clover: 'drought tolerance' and 'drought recovery'. We further found that a large proportion of the accessions able to perform well under well-watered conditions were also the ones that were less affected by drought. The results of this investigation are interpreted in view of the development of breeding for adaptation to drought in red clover.

Dataset Information

A proteome analysis of freezing tolerance in red clover (Trifolium pratense L.).

Publications

A proteome analysis of freezing tolerance in red clover (Trifolium pratense L.).

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