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Identification and analysis of MKK and MPK gene families in canola (Brassica napus L.).

ABSTRACT: BACKGROUND: Eukaryotic mitogen-activated protein kinase (MAPK/MPK) signaling cascades transduce and amplify environmental signals via three types of reversibly phosphorylated kinases to activate defense gene expression. Canola (oilseed rape, Brassica napus) is a major crop in temperate regions. Identification and characterization of MAPK and MAPK kinases (MAPKK/MKK) of canola will help to elucidate their role in responses to abiotic and biotic stresses. RESULTS: We describe the identification and analysis of seven MKK (BnaMKK) and 12 MPK (BnaMPK) members from canola. Sequence alignments and phylogenetic analyses of the predicted amino acid sequences of BnaMKKs and BnaMPKs classified them into four different groups. We also examined the subcellular localization of four and two members of BnaMKK and BnaMPK gene families, respectively, using green fluorescent protein (GFP) and, found GFP signals in both nuclei and cytoplasm. Furthermore, we identified several interesting interaction pairs through yeast two-hybrid (Y2H) analysis of interactions between BnaMKKs and BnaMPKs, as well as BnaMPK and BnaWRKYs. We defined contiguous signaling modules including BnaMKK9-BnaMPK1/2-BnaWRKY53, BnaMKK2/4/5-BnaMPK3/6-BnaWRKY20/26 and BnaMKK9-BnaMPK5/9/19/20. Of these, several interactions had not been previously described in any species. Selected interactions were validated in vivo by a bimolecular fluorescence complementation (BiFC) assay. Transcriptional responses of a subset of canola MKK and MPK genes to stimuli including fungal pathogens, hormones and abiotic stress treatments were analyzed through real-time RT-PCR and we identified a few of BnaMKKs and BnaMPKs responding to salicylic acid (SA), oxalic acid (OA), Sclerotinia sclerotiorum or other stress conditions. Comparisons of expression patterns of putative orthologs in canola and Arabidopsis showed that transcript expression patterns were generally conserved, with some differences suggestive of sub-functionalization. CONCLUSIONS: We identified seven MKK and 12 MPK genes from canola and examined their phylogenetic relationships, transcript expression patterns, subcellular localization, and protein-protein interactions. Not all expression patterns and interactions were conserved between canola and Arabidopsis, highlighting the limitations of drawing inferences about crops from model species. The data presented here provide the first systematic description of MKK-MPK-WRKY signaling modules in canola and will further improve our understanding of defense responses in general and provide a basis for future crop improvement.

SUBMITTER: Liang W

PROVIDER: S-EPMC3701561 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Identification and analysis of MKK and MPK gene families in canola (Brassica napus L.).

Liang Wanwan W Yang Bo B Yu Bao-Jun BJ Zhou Zili Z Li Cui C Jia Ming M Sun Yun Y Zhang Yue Y Wu Feifei F Zhang Hanfeng H Wang Boya B Deyholos Michael K MK Jiang Yuan-Qing YQ

BMC genomics 20130611

<h4>Background</h4>Eukaryotic mitogen-activated protein kinase (MAPK/MPK) signaling cascades transduce and amplify environmental signals via three types of reversibly phosphorylated kinases to activate defense gene expression. Canola (oilseed rape, Brassica napus) is a major crop in temperate regions. Identification and characterization of MAPK and MAPK kinases (MAPKK/MKK) of canola will help to elucidate their role in responses to abiotic and biotic stresses.<h4>Results</h4>We describe the iden ...[more]

PMID: 23758924

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Project description:BackgroundCanola (Brassica napus L.) is one of the most important oil-producing crops in China and worldwide. The yield and quality of canola is frequently threatened by environmental stresses including drought, cold and high salinity. Calcium is a ubiquitous intracellular secondary messenger in plants. Calcineurin B-like proteins (CBLs) are Ca2+ sensors and regulate a group of Ser/Thr protein kinases called CBL-interacting protein kinases (CIPKs). Although the CBL-CIPK network has been demonstrated to play crucial roles in plant development and responses to various environmental stresses in Arabidopsis, little is known about their function in canola.ResultsIn the present study, we identified seven CBL and 23 CIPK genes from canola by database mining and cloning of cDNA sequences of six CBLs and 17 CIPKs. Phylogenetic analysis of CBL and CIPK gene families across a variety of species suggested genome duplication and diversification. The subcellular localization of three BnaCBLs and two BnaCIPKs were determined using green fluorescence protein (GFP) as the reporter. We also demonstrated interactions between six BnaCBLs and 17 BnaCIPKs using yeast two-hybrid assay, and a subset of interactions were further confirmed by bimolecular fluorescence complementation (BiFC). Furthermore, the expression levels of six selected BnaCBL and 12 BnaCIPK genes in response to salt, drought, cold, heat, ABA, methyl viologen (MV) and low potassium were examined by quantitative RT-PCR and these CBL or CIPK genes were found to respond to multiple stimuli, suggesting that the canola CBL-CIPK network may be a point of convergence for several different signaling pathways. We also performed a comparison of interaction patterns and expression profiles of CBL and CIPK in Arabidospsis, canola and rice, to examine the differences between orthologs, highlighting the importance of studying CBL-CIPK in canola as a prerequisite for improvement of this crop.ConclusionsOur findings indicate that CBL and CIPK family members may form a dynamic complex to respond to different abiotic or hormone signaling. Our comparative analyses of the CBL-CIPK network between canola, Arabidopsis and rice highlight functional differences and the necessity to study CBL-CIPK gene functions in canola. Our data constitute a valuable resource for CBL and CPK genomics.

Project description:Genomic selection (GS) is a modern breeding approach where genome-wide single-nucleotide polymorphism (SNP) marker profiles are simultaneously used to estimate performance of untested genotypes. In this study, the potential of genomic selection methods to predict testcross performance for hybrid canola breeding was applied for various agronomic traits based on genome-wide marker profiles. A total of 475 genetically diverse spring-type canola pollinator lines were genotyped at 24,403 single-copy, genome-wide SNP loci. In parallel, the 950 F1 testcross combinations between the pollinators and two representative testers were evaluated for a number of important agronomic traits including seedling emergence, days to flowering, lodging, oil yield and seed yield along with essential seed quality characters including seed oil content and seed glucosinolate content. A ridge-regression best linear unbiased prediction (RR-BLUP) model was applied in combination with 500 cross-validations for each trait to predict testcross performance, both across the whole population as well as within individual subpopulations or clusters, based solely on SNP profiles. Subpopulations were determined using multidimensional scaling and K-means clustering. Genomic prediction accuracy across the whole population was highest for seed oil content (0.81) followed by oil yield (0.75) and lowest for seedling emergence (0.29). For seed yieId, seed glucosinolate, lodging resistance and days to onset of flowering (DTF), prediction accuracies were 0.45, 0.61, 0.39 and 0.56, respectively. Prediction accuracies could be increased for some traits by treating subpopulations separately; a strategy which only led to moderate improvements for some traits with low heritability, like seedling emergence. No useful or consistent increase in accuracy was obtained by inclusion of a population substructure covariate in the model. Testcross performance prediction using genome-wide SNP markers shows considerable potential for pre-selection of promising hybrid combinations prior to resource-intensive field testing over multiple locations and years.

Dataset Information

Identification and analysis of MKK and MPK gene families in canola (Brassica napus L.).

Publications

Identification and analysis of MKK and MPK gene families in canola (Brassica napus L.).

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