Calcium-dependent Protein Kinases Gene Research Articles

Jute CDPK genes and their role in stress tolerance and fiber development: A genome-wide bioinformatic investigation of Chorchorus capsularis and C. olitorius

Calcium-dependent protein kinases (CDPKs) are plant-specific calcium sensors, which regulate plant growth and development and induce protective responses to environmental stresses. To date, no information on jute CDPKs is available and their uses in the genetic improvement program is underutilized. In this study, we exploited recently available genome sequence information of two jute species: Corchorus olitorius and C. capsularis to identify, characterise, and functional analysis. Using bioinformatics tools, 16 and 18 CDPK genes with serine/threonine protein kinase along with EF-hand calcium-binding domain were identified from the genomes of C. olitorius and C. capsularis, respectively. Different chromosomes harboured different CDPK genes. Four distinct clades of CDPK genes were identified by phylogenetic analysis. Protein of each clade showed higher conservation in length, structure, and presence of motif. The most prominent transcriptional responses were detected in saline and drought conditions where more than half of the CDPKs were upregulated, suggesting their important roles in abiotic stress responses of jute. Specifically, CoCDPK6, 11, 12, and their corresponding homologs CcCDPK18, 10, 8 were highly expressed in saline and drought conditions. On the other hand, CoCDPK6, 7, 11, 12, and their homologs CcCDPK18, 17, 10, 8 were highly expressed in fiber cell indicating their involvement in fiber cell development. Our results provide an important foundation for further functional dissection of CDPKs of C. olitorius and C. capsularis for salinity and drought stress adaptation and fiber quality improvement.

Improving plant drought tolerance and growth under water limitation through combinatorial engineering of signalling networks.

Agriculture is by far the biggest water consumer on our planet, accounting for 70 per cent of all freshwater withdrawals. Climate change and a growing world population increase pressure on agriculture to use water more efficiently (‘more crop per drop’). Water‐use efficiency (WUE) and drought tolerance of crops are complex traits that are determined by many physiological processes whose interplay is not well understood. Here, we describe a combinatorial engineering approach to optimize signalling networks involved in the control of stress tolerance. Screening a large population of combinatorially transformed plant lines, we identified a combination of calcium‐dependent protein kinase genes that confers enhanced drought stress tolerance and improved growth under water‐limiting conditions. Targeted introduction of this gene combination into plants increased plant survival under drought and enhanced growth under water‐limited conditions. Our work provides an efficient strategy for engineering complex signalling networks to improve plant performance under adverse environmental conditions, which does not depend on prior understanding of network function.

Genome-wide identification, and characterization of the CDPK gene family reveal their involvement in abiotic stress response in Fragaria x ananassa

Calcium-dependent protein kinases (CDPKs) are encoded by a large gene family and play important roles against biotic and abiotic stresses and in plant growth and development. To date, little is known about the CDPK genes in strawberry (Fragaria x ananassa). In this study, analysis of Fragaria x ananassa CDPK gene family was performed, including gene structures, phylogeny, interactome and expression profiles. Nine new CDPK genes in Fragaria x ananassa were identified based on RNA-seq data. These identified strawberry FaCDPK genes were classified into four main groups, based on the phylogenetic analysis and structural features. FaCDPK genes were differentially expressed during fruit development and ripening, as well as in response to abiotic stress (salt and drought), and hormone (abscisic acid) treatment. In addition, the interaction network analysis pointed out proteins involved in the ABA-dependent response to plant stress via Ca2+ signaling, especially RBOHs. To our knowledge, this is the first report on CDPK families in Fragaria x ananassa, and it will provide valuable information for development of biofortified fruits and stress tolerant plants.

Molecular cloning and expression of AmCDPK from mangrove Avicennia marina under elevated temperature.

Considered as an essential calcium sensor, the calcium-dependent protein kinase (CDPK) family plays a critical part in terrestrial plants' responses to both biotic and abiotic stresses. In the study, Avicennia marina was proved to have better heat tolerance than other species. A CDPK gene was cloned from mangrove species A. marina using RACE-PCR and designated as AmCDPK. By predicting and analyzing its properties, structures and expression patterns, we found that the amino acid sequence, containing a kinase domain and four EF-hand Ca2+-binding sites, shared high identity with Handroanthus impetiginosus and Sesamum indicum. Quantitative real-time PCR data analysis suggested that AmCDPK demonstrated significant up-regulation under heat stress. It is likely that AmCDPK is a versatile gene involved in various stresses, including dehydration, cold, light, defense and ABA stress responses by analyzing cis-elements. It is the first time that CDPKs from mangroves have been cloned and our results brought evidence to the effect of AmCDPK on heat stress, which is particularly important under the background of global warming.

Genome-wide investigation of calcium-dependent protein kinase gene family in pineapple: evolution and expression profiles during development and stress

BackgroundCalcium-dependent protein kinase (CPK) is one of the main Ca2+ combined protein kinase that play significant roles in plant growth, development and response to multiple stresses. Despite an important member of the stress responsive gene family, little is known about the evolutionary history and expression patterns of CPK genes in pineapple.ResultsHerein, we identified and characterized 17 AcoCPK genes from pineapple genome, which were unevenly distributed across eight chromosomes. Based on the gene structure and phylogenetic tree analyses, AcoCPKs were divided into four groups with conserved domain. Synteny analysis identified 7 segmental duplication events of AcoCPKs and 5 syntenic blocks of CPK genes between pineapple and Arabidopsis, and 8 between pineapple and rice. Expression pattern of different tissues and development stages suggested that several genes are involved in the functional development of plants. Different expression levels under various abiotic stresses also indicated that the CPK family underwent functional divergence during long-term evolution. AcoCPK1, AcoCPK3 and AcoCPK6, which were repressed by the abiotic stresses, were shown to be function in regulating pathogen resistance.Conclusions17 AcoCPK genes from pineapple genome were identified. Our analyses provide an important foundation for understanding the potential roles of AcoCPKs in regulating pineapple response to biotic and abiotic stresses

Genome-wide identification and expression analysis of the citrus calcium-dependent protein kinase (CDPK) genes in response to arbuscular mycorrhizal fungi colonization and drought

The calcium-dependent protein kinases (CDPKs, also termed CPKs) are a large, diverse protein family that play significant roles in biotic and abiotic stress responses in plants. However, CDPKs in citrus are relatively unknown, as are their responses to drought stress (abiotic stress) and arbuscular mycorrhizal fungi (AMF) colonization. In this study, the citrus CDPKs were characterized by analyzing the presence of EF-hand and Pkinase domains and searching the citrus genome. Twenty-nine Citrus sinensis CDPKs (CsCDPKs) were identified on at least nine chromosomes in the citrus genome. Three clusters were found on chromosome 4 with three or two CsCDPKs, and 17 CsCDPKs showed syntenic relationships with Arabidopsis thaliana CPKs (AtCPKs). Twenty-nine CsCDPKs were expressed in the roots, and the regulation of the CsCDPKs was not always consistent with the cis-elements in their promoters. CsCDPK1, 2, 5, 6, 7, 8, 9, 13, 17, 18, 19, 21, 24, 25, 26, 27 and 29 were upregulated by AMF colonization, and CsCDPK3, 7 and 28 were downregulated. Drought-repressed expression of CsCDPK10, 11, 12, 15 and 16 induced CsCDPK20 and 22. Furthermore, AMF colonization and drought stress exerted synergistic effects; the expression levels of CsCDPK20 and CsCDPK22 were repressed in W_AMF treatment (well-watered with AMF) but induced in D_AMF treatment (drought stress with AMF). Together, these results provide a basis for further functional studies of the CsCDPK proteins and their responses to AMF and drought stress.

Evolutionary Analysis of Calcium-Dependent Protein Kinase in Five Asteraceae Species.

Calcium-dependent protein kinase (CPK) is crucial in Ca2+ signal transduction, and is a large gene family in plants. In our previous work, we reported Hevea brasiliensis CPKs were important for natural rubber biosynthesis. However, this CPK gene family in other rubber producing plants has not been investigated. Here, we report the CPKs in five representative Asteraceae species, including three rubber-producing and two non-rubber species. A total of 34, 34, 40, 34 and 30 CPKs were identified from Taraxacum koksaghyz, Lactuca sativa, Helianthus annuus, Chrysanthemum nankingense and Cynara cardunculus, respectively. All CPKs were classified into four individual groups (group I to IV). In addition, 10 TkCPKs, 11 LsCPKs, 20 HaCPKs, 13 CnCPKs and 7 CcCPKs duplicated paralogs were identified. Further evolutionary analysis showed that, compared to other subfamilies, the group III had been expanded in the Asteraceae species, especially in the rubber-producing species. Meanwhile, the CPKs in group III from Asteraceae species tend to expand with low calcium binding capacity. This study provides a systematical evolutionary investigation of the CPKs in five representative Asteraceae species, suggesting that the sub-family specific expansion of CPKs might be related to natural rubber producing.

Differential expression of calcium-dependent protein kinase genes (CDPK1–14) in Rubia cordifolia callus cultures transformed with the rolB and rolC genes

Integration of Agrobacterium rhizogenes T-DNA region leads to multiple changes in the physiology and biochemistry of the transformed plant cells. We have previously shown that one of these possible effects may be related to the changes in the expression profile of calcium-dependent protein kinase (CDPK) genes. Since the key determinants of T-DNA are rol genes, in this work, we studied the individual action of the rolB and rolC genes on the expression of CDPK genes in transgenic cells of Rubia cordifolia calli. Rol genes caused a general increase in total R. cordifolia CDPK transcripts abundance. Expression of RcCDPK2, RcCDPK7, RcCDPK8, RcCDPK9 and RcCDPK11 was 1.3- to 3.5-fold higher in both rolB- and rolC-transformed cells than in non-transformed cells, whereas expression of RcCDPK10 was 1.9-fold lower. Expression of RcCDPK1, RcCDPK13 and RcCDPK14 was up-regulated by the rolB gene, and the transcriptional level of RcCDPK4 and RcCDPK5 was higher in the rolC-expressing cells. Our results indicate that complex but often divergent effects of rolB and rolC on transformed plant cells may be attributed to changes caused by these genes on particular CDPK isoforms.

Identification, Expression, and Interaction Network Analyses of the CDPK Gene Family Reveal Their Involvement in the Development, Ripening, and Abiotic Stress Response in Banana.

Calcium-dependent protein kinases (CDPKs) play vital roles in the regulation of plant growth, development, and tolerance to various abiotic stresses. However, little information is available for this gene family in banana. In this study, 44 CDPKs were identified in banana and were classified into four groups based on phylogenetic, gene structure, and conserved motif analyses. The majority of MaCDPKs generally exhibited similar expression patterns in the different tissues. Transcriptome analyses revealed that many CDPKs showed strong transcript accumulation at the early stages of fruit development and postharvest ripening in both varieties. Interaction network and co-expression analysis further identified some CDPKs-mediated network that was potentially active at the early stages of fruit development. Comparative expression analysis suggested that the high levels of CDPK expression in FJ might be related to its fast ripening characteristic. CDPK expression following the abiotic stress treatments indicated a significant transcriptional response to osmotic, cold, and salt treatment, as well as differential expression profiles, between BX and FJ. The findings of this study elucidate the transcriptional control of CDPKs in development, ripening, and the abiotic stress response in banana. Some tissue-specific, development/ripening-dependent, and abiotic stress-responsive candidate MaCDPK genes were identified for further genetic improvement of banana.

Expression analysis of calcium-dependent protein kinases (CDPKs) superfamily genes in Medicago lupulina in response to high calcium, carbonate and drought

The aim of this study is to identify calcium dependent protein kinases (CDPKs) superfamily genes (including calcium dependent protein kinase (CDPK) genes, CDPK-related kinase (CRK) genes, and phosphoenolpyruvate carboxylase kinase-related kinase (PEPRK) genes) and to analyze their expression in response to high calcium, bicarbonate, and water deficit in Medicago lupulina. Transcriptome sequencing databases and bioinformatics were used to identify CDPK superfamily genes. qRT-PCR analyses were used to study the expression of CDPK superfamily genes in response to high calcium, bicarbonate, water deficit, and their combined treatments. Ten MlCDPKs, four MlCRKs, and two MlPEPRKs genes were identified from transcriptome sequencing databases. The identified MlCDPK superfamily genes could be divided into six subgroups including CDPK I, CDPK II, CDPK III, CDPK IV, CRK, and PEPRK according to evolutionary relationships. Analysis of protein structure indicated that MlCDPK proteins contained a protein kinase domain and EF-hand domain. However, PEPRKs contained only a protein kinase domain while CRKs contained also degenerative EF-hands. Most of the MlCDPK superfamily genes were differentially expressed in different organs. According to their expression levels, MlCDPK superfamily genes such as MlCDPK1, MlCDPK9, and MlCDPK10 were more sensitive in response to individual treatments of high calcium, carbonate or PEG rather than their combination of treatments. The results indicated that MlCDPK superfamily genes may play an important role in response to those stresses. These results provide important insight into the role of CDPKs in M. lupulina, especially in the tolerance to Karst habitats, where high calcium and carbonate and drought conditions are the dominant ecological limiting factors.

Calcium-Dependent Protein Kinase Genes in Glycyrrhiza Uralensis Appear to be Involved in Promoting the Biosynthesis of Glycyrrhizic Acid and Flavonoids under Salt Stress.

As calcium signal sensors, calcium-dependent protein kinases (CPKs) play vital roles in stimulating the production of secondary metabolites to participate in plant development and response to environmental stress. However, investigations of the Glycyrrhiza uralensis CPK family genes and their multiple functions are rarely reported. In this study, a total of 23 GuCPK genes in G. uralensis were identified, and their phylogenetic relationships, evolutionary characteristics, gene structure, motif distribution, and promoter cis-acting elements were analyzed. Ten GuCPKs showed root-specific preferential expressions, and GuCPKs indicated different expression patterns under treatments of CaCl2 and NaCl. In addition, under 2.5 mM of CaCl2 and 30 mM of NaCl treatments, the diverse, induced expression of GuCPKs and significant accumulations of glycyrrhizic acid and flavonoids suggested the possible important function of GuCPKs in regulating the production of glycyrrhizic acid and flavonoids. Our results provide a genome-wide characterization of CPK family genes in G. uralensis, and serve as a foundation for understanding the potential function and regulatory mechanism of GuCPKs in promoting the biosynthesis of glycyrrhizic acid and flavonoids under salt stress.

The Role of Calcium-Dependent Protein Kinase Genes VaCPK1 and VaCPK26 in the Response of Vitis amurensis (in vitro) and Arabidopsis thaliana (in vivo) to Abiotic Stresses

Calcium-dependent protein kinases (CDPK or CPK) are involved in protecting plants from abiotic stresses, but the properties and functions of CDPK are still poorly understood. Analysis of the Amur grape Vitis amurensis Rupr. is of great interest, since it demonstrates high tolerance to adverse environmental conditions. It was demonstrated that exposure of V. amurensis to salt and, to a lesser extent, osmotic and cold stress resulted in a considerable increase in the VaCPK1 and VaCPK26 transcription level. The present study was focused on the analysis of the effect of constitutive expression of the recombinant VaCPK1 and VaCPK26 genes on the resistance of V. amurensis cell cultures and Arabidopsis thaliana plants to salt, osmotic, and temperature stresses. It was demonstrated that expression of recombinant VaCPK26, as well as to a lesser extent VaCPK1, led to a 1.2–1.7 times increase in biomass accumulation by V. amurensis cell cultures and to a 1.2–2.1 times increase in survival of A. thaliana plants under salt stress. In addition, VaCPK26-expressing lines of A. thaliana lines demonstrated a slight degree of drought tolerance and were characterized by increased expression of some stress-induced protective genes involved in the formation of salt and dehydration tolerance. The data obtained indicate that the VaCPK1 and VaCPK26 genes can act as positive regulators of plant response to salt stress.

Nitrogen Metabolism is Affected in the Nitrogen-Deficient Rice Mutant esl4 with a Calcium-Dependent Protein Kinase Gene Mutation.

Calcium-dependent protein kinases are involved in various biological processes, including hormone response, growth and development, abiotic stress response, disease resistance, and nitrogen metabolism. We identified a novel mutant of a calcium-dependent protein-kinase-encoding gene, esl4, by performing map cloning. The esl4 mutant was nitrogen deficient, and expression and enzyme activities of genes related to nitrogen metabolism were down-regulated. ESL4 was mainly expressed in the vascular bundles of roots, stems, leaves, and sheaths. The ESL4 protein was localized in the cell membranes. Enzyme activity and physiological index analyzes and analysis of the expression of nitrogen metabolism and senescence-related genes indicated that ESL4 was involved in nitrogen metabolism. ESL4 overexpression in transgenic homozygous T2 plants increased nitrogen-use efficiency, improving yields when little nitrogen was available. The seed-set rates, yields per plant, numbers of grains per plant, grain nitrogen content ratios, and total nitrogen content per plant were significantly or very significantly higher for two ESL4 overexpression lines than for the control plants. These results suggest that ESL4 may function upstream of nitrogen-metabolism genes. The results will allow ESL4 to be used to breed novel cultivars for growing in low-nitrogen conditions.

Genome-Wide Analysis of CDPK Family in Foxtail Millet and Determination of SiCDPK24 Functions in Drought Stress.

Plant calcium-dependent protein kinases (CDPKs) were reported to play important roles in plant resistance to abiotic stress. Foxtail millet cultivation “H138” was used for RNA-seq analysis. The data from drought-induced de novo transcriptomic sequences of foxtail millet showed that CDPKs were up- or down-regulated by drought to different degrees. In this study, 29 foxtail millet CDPKs were classified into four subgroups. These genes were unevenly distributed on nine foxtail millet chromosomes, and chromosomes 2, 3, and 9 contained the most SiCDPK members. Analysis of putative cis-acting elements showed that most foxtail millet CDPK genes contained the ABRE, LTR, HSE, MYB, MYC, DRE, CGTCA-motif, and TGACG-motif cis-acting elements, which could be activated by abiotic stresses. Real-time PCR analysis indicated that 29 SiCDPK genes experienced different degrees of induction under drought and ABA stresses. SiCDPK24 had the highest expression levels at 6 and 12 h of drought treatment and was chosen for further analysis. SiCDPK24 localized to the cell membrane and the nucleus of Arabidopsis mesophyll protoplasts. Western blot analysis showed that SiCDPK24 protein had autophosphorylation activity. Overexpression of SiCDPK24 in Arabidopsis enhanced drought resistance and improved the survival rate under drought stress. It also activated the expressions of nine stress-related genes, namely RD29A, RD29B, RD22, KIN1, COR15, COR47, LEA14, CBF3/DREB1A, and DREB2A. These genes are involved in resistance to abiotic stresses in Arabidopsis. These results indicate that foxtail millet CDPK genes play important roles in resisting drought stress.

A calcium-dependent protein kinase, ZmCPK32, specifically expressed in maize pollen to regulate pollen tube growth.

Calcium-dependent protein kinases (CPKs) play an essential role in the regulation of pollen tube growth. Although CPK genes have been identified in maize, and some have been functionally characterized, the molecular function of ZmCPKs associated with pollen tube development remains less well studied. Here, we report that a pollen-specific CPK, ZmCPK32, is involved in the regulation of pollen germination and tube extension. ZmCPK32 exhibited CPK activity and was localized on the plasma membrane and punctate internal membrane compartments via N-terminal acylation. In situ hybridization and real-time PCR revealed that ZmCPK32 transcripts accumulated in pollen and expression was dramatically upregulated during shedding. To elucidate the function of this gene, we transiently expressed a ZmCPK32-GFP fusion protein in tobacco pollen using microparticle bombardment. ZmCPK32 accumulation inhibited pollen germination and reduced pollen tube growth, but this effect was abolished when the kinase-inactive variant was expressed, indicating that kinase activity is critical for its regulatory function. In addition, the plasma membrane localization of ZmCPK32 is essential for regulating polar growth, as pollen expressing the cytosol-localized kinase displayed reduced tube length but germinated well. Moreover, the constitutively active form of ZmCPK32 enhanced the reduction in the germination rate, indicating that the specific activation of ZmCPK32 via calcium ions at the cortical growth point is essential for regulating appropriate germination. The results suggest that ZmCPK32 is functionally associated with pollen tube growth, and could represent a potential target for breeding male-sterile maize.

Calcium-dependent protein kinases in cotton: insights into early plant responses to salt stress

BackgroundSoil salinization is one of the major environmental constraints to plant growth and agricultural production worldwide. Signaling components involving calcium (Ca2+) and the downstream calcium-dependent protein kinases (CPKs) play key roles in the perception and transduction of stress signals. However, the study of CPKs in cotton and their functions in response to salt stress remain unexplored.ResultsA total of 98 predicted CPKs were identified from upland cotton (Gossypium hirsutum L. ‘TM-1’), and phylogenetic analyses classified them into four groups. Gene family distribution studies have revealed the substantial impacts of the genome duplication events to the total number of GhCPKs. Transcriptome analyses showed a wide distribution of CPKs’ expression among different organs. A total of 19 CPKs were selected for their rapid responses to salt stress at the transcriptional level, most of which were also incduced by the thylene-releasing chemical ethephon, suggesting a partal overlap of the salinity and ethylene responses. Silencing of 4 of the 19 CPKs (GhCPK8, GhCPK38, GhCPK54, and GhCPK55) severely compromised the basal cotton resistance to salt stress.ConclusionsOur genome-wide expression analysis of CPK genes from up-land cotton suggests that CPKs are involved in multiple developmental responses as well as the response to different abiotic stresses. A cluster of the cotton CPKs was shown to participate in the early signaling events in cotton responses to salt stress. Our results provide significant insights on functional analysis of CPKs in cotton, especially in the context of cotton adaptions to salt stress.

Bioinformatic Analysis of Calcium-dependent Protein Kinase Family of <i>Medicago truncatula</i>

Calcium plays an important biological role in plants, regulating cell wall formation, pollen tube development, biotic and abiotic stress signal transduction and so on. Calcium-dependent protein kinases (CDPKs) genes are able to perceive the change of intracellular Ca 2+ concentrations and convert it into specific phosphorylation events, initiating downstream signal transduction. We used bioinformatics data and analysis techniques to identify family members of the CDPKs gene in two varieties of Medicago , namely Medicago truncatula and Medicago lupulina L., and to analyze the structure, evolution and expression of these CDPKs genes, preliminarily predicted the functions of these genes. 25 CDPKs genes present in the genome of Medicago truncatula , which were divided into four evolutionary groups. The evolutionary relationship of CDPKs gene of Medicago truncatula was close to that of Arabidopsis thaliana but had a distant relation with Oryza sativa . Three CDPKs genes, Medtr1g041150, Medtr1g054865 and Medtr1g055255, clustered on chromosome 1 of Medicago truncatula , were highly similar and might be amplified from one gene. We analyzed the expression of CDPKs gene of Medicago truncatula in in different organs and under different treatments with existing gene expression data of Medicago truncatula , and found several genes related to the salt tolerance, root rot resistance and nodule growth of Medicago truncatula .

Comparative Analysis of CDPK Family in Maize, Arabidopsis, Rice, and Sorghum Revealed Potential Targets for Drought Tolerance Improvement.

Calcium dependent protein kinases (CDPKs) play significant role in regulation of plant growth and development in response to various stresses including drought. A set of 32 CDPK genes identified in maize were further used for searching of orthologs in the model plant Arabidopsis (72) and major food crops such as rice (78) and sorghum (91). We comprehensively studied the phylogenetic relationship, annotations, gene duplications, gene structure, divergence time, 3-D protein structures and tissue-specific drought induced expression of CDPK genes in all four species. Variation in intron frequency in the studied species was one of the reasons for the functional diversity of CDPK genes to various stress responses. Protein kinase and protein kinase C phosphorylation site domains were the most conserved motifs identified in all species. Four groups were identified from the sequence-based phylogenetic analysis, in which maize CDPKs were clustered in group III. Expression data showed that the CDPK genes were highly expressed in leaf of maize, rice, and sorghum whereas in Arabidopsis the maximum expression was observed in root. The expression assay showed 5, 6, 11, and 9 were the commonly and differentially expressed drought-related orthologous genes in maize, Arabidopsis, rice, and sorghum, respectively. 3-D protein structure were predicted for the nine genes (Arabidopsis: 2, maize: 2, rice: 3, and sorghum: 2) showing differential expression in at least three species. The predicted 3-D structures were further evaluated and validated by Ramachandran plot, ANOLEA, ProSA, and Verify-3D. The superimposed 3-D structure of drought-related orthologous proteins retained similar folding pattern owing to their conserved nature. Functional annotation revealed the involvement of CDPK genes in various pathways such as osmotic homeostasis, cell protection, and root growth. The interactions of CDPK genes in various pathways play crucial role in imparting drought tolerance through different ABA and MAPK signaling cascades. These selected candidate genes could be targeted in development of drought tolerant genotypes in maize, rice, and sorghum through appropriate breeding approaches. Our comparative experiments of CDPK genes could also be extended in the drought stress breeding programmes of the related species.

A Functional Analysis of OsCPK11, a Calcium-dependent Protein Kinase (CDPK) Gene in Rice

A Functional Analysis of OsCPK11, a Calcium-dependent Protein Kinase (CDPK) Gene in Rice

Calcium-dependent protein kinase 21 phosphorylates 14-3-3 proteins in response to ABA signaling and salt stress in rice

The calcium-dependent protein kinases (CDPKs) are a class of plant-specific kinase that directly bind Ca2+ and mediate the calcium-signaling pathways to play important physiological roles in growth and development. The rice genome contains 31 CDPK genes, one of which, OsCPK21, is known to modulate the abscisic acid (ABA) and salt stress responses in this crop; however, the molecular mechanisms underlying this regulation are largely unknown. In the present study, we performed yeast two-hybrid screening, glutathione S-transferase pull-down, co-immunoprecipitation, and bimolecular fluorescence complementation assays to confirm the interaction between OsCPK21 and one of its putative targets, Os14-3-3 (OsGF14e). We used an in vitro kinase assay and site-directed mutagenesis to verify that OsCPK21 phosphorylates OsGF14e at Tyr-138. We used real-time PCR to reveal that several ABA and salt inducible genes were more highly expressed in the OsCPK21-OE and OsGF14e WT-OE plants than in the mutant OsGF14e Y138A-OE and wild-type plants. These results suggest that OsCPK21 phosphorylates OsGF14e to facilitate the response to ABA and salt stress.