Calcium-dependent Protein Kinases Gene Research Articles

Genome-wide identification and stress response analysis of BcaCPK gene family in amphidiploid Brassica carinata.

Calcium-dependent protein kinases (CPKs) are crucial for recognizing and transmitting Ca2+ signals in plant cells, playing a vital role in growth, development, and stress response. This study aimed to identify and detect the potential roles of the CPK gene family in the amphidiploid Brassica carinata (BBCC, 2n = 34) using bioinformatics methods. Based on the published genomic information of B. carinata, a total of 123 CPK genes were identified, comprising 70 CPK genes on the B subgenome and 53 on the C subgenome. To further investigate the homologous evolutionary relationship between B. carinata and other plants, the phylogenetic tree was constructed using CPKs in B. carinata and Arabidopsis thaliana. The phylogenetic analysis classified 123 family members into four subfamilies, where gene members within the same subfamily exhibited similar conserved motifs. Each BcaCPK member possesses a core protein kinase domain and four EF-hand domains. Most of the BcaCPK genes contain 5 to 8 introns, and these 123 BcaCPK genes are unevenly distributed across 17 chromosomes. Among these BcaCPK genes, 120 replicated gene pairs were found, whereas only 8 genes were tandem duplication, suggesting that dispersed duplication mainly drove the family amplification. The results of the Ka/Ks analysis indicated that the CPK gene family of B. carinata was primarily underwent purification selection in evolutionary selection. The promoter region of most BcaCPK genes contained various stress-related cis-acting elements. qRT-PCR analysis of 12 selected CPK genes conducted under cadmium and salt stress at various points revealed distinct expression patterns among different family members in response to different stresses. Specifically, the expression levels of BcaCPK2.B01a, BcaCPK16.B02b, and BcaCPK26.B02 were down-regulated under both stresses, whereas the expression levels of other members were significantly up-regulated under at least one stress. This study systematically identified the BcaCPK gene family in B. carinata, which contributes to a better understanding the CPK genes in this species. The findings also serve as a reference for analyzing stress responses, particularly in relation to cadmium and salt stress in B. carinata.

The maize ZmCPK39-ZmKnox2 module regulates plant height

AbstractPlant height is an important agronomic trait that affects high-density tolerance and lodging resistance. However, the regulators and their underlying molecular mechanisms controlling plant height in maize remain understudied. Here, we report that knockout mutants of the calcium-dependent protein kinase gene ZmCPK39 (ZmCPK39-KO) exhibit dramatically reduced plant height, characterized by shorter internodes and a slight decrease in node numbers. Furthermore, we identified a ZmCPK39-interacting protein, the knotted-related homeobox (ZmKnox2), and observed that plant height was also significantly reduced in a mutator transposon-inserted mutant of ZmKnox2 (ZmKnox2-Mu). Combined analysis of transcriptomic and metabonomic data indicates that multiple phytohormone signaling and photosynthesis pathways are disrupted in both ZmCPK39-KO and ZmKnox2-Mu mutants. Taken together, these results provide new insights into the function of ZmCPK39 and identify potential targets for breeding lodging-resistant and high-density tolerant maize cultivars.

Identification of the CDPK gene family in patchouli and functional analysis in response to continuous cropping stress

To further reveal the molecular mechanisms underlying the formation of continuous cropping disorders in patchouli, this study analyzed the function of calcium dependent protein kinase (CDPK) genes at the molecular level in patchouli continuous cropping disorders. The findings unveiled the presence of 45 PcCDPK genes within the complete patchouli genome dataset. These genes exhibited a range of molecular weights from 50.78 to 78.96 kDa and aliphatic index values spanning from 74.42 to 88.49, and are shown to be hydrophilic proteins. The evolution of 45 PcCDPK members was divided into 4 subfamilies, with a total of 65 pairs of collinear genes. Each PcCDPK contains a STKc-CAMK domain and four EF-hand structures exhibiting a certain degree of conservatism during evolution. Transcriptome data further supported the significance of PcCDPK25 and PcCDPK38 genes, showing substantial upregulation, which was corroborated by qRT-PCR results. The 1629 bp and 1716 bp CDS sequences were obtained by cloning the PcCDPK25 and PcCDPK38 genes, respectively, and subcellular localization showed that both proteins were localized on the plasma membrane. This comprehensive study provides molecular-level confirmation of the pivotal roles played by CDPK genes in the emergence of continuous cropping challenges in patchouli plants, establishing a crucial foundation for a deeper comprehension of the molecular mechanisms underpinning these obstacles.

Genome-Wide Identification and Characterization of CDPK Gene Family in Cultivated Peanut (Arachis hypogaea L.) Reveal Their Potential Roles in Response to Ca Deficiency.

This study identified 45 calcium-dependent protein kinase (CDPK) genes in cultivated peanut (Arachis hypogaea L.), which are integral in plant growth, development, and stress responses. These genes, classified into four subgroups based on phylogenetic relationships, are unevenly distributed across all twenty peanut chromosomes. The analysis of the genetic structure of AhCDPKs revealed significant similarity within subgroups, with their expansion primarily driven by whole-genome duplications. The upstream promoter sequences of AhCDPK genes contained 46 cis-acting regulatory elements, associated with various plant responses. Additionally, 13 microRNAs were identified that target 21 AhCDPK genes, suggesting potential post-transcriptional regulation. AhCDPK proteins interacted with respiratory burst oxidase homologs, suggesting their involvement in redox signaling. Gene ontology and KEGG enrichment analyses affirmed AhCDPK genes' roles in calcium ion binding, protein kinase activity, and environmental adaptation. RNA-seq data revealed diverse expression patterns under different stress conditions. Importantly, 26 AhCDPK genes were significantly induced when exposed to Ca deficiency during the pod stage. During the seedling stage, four AhCDPKs (AhCDPK2/-25/-28/-45) in roots peaked after three hours, suggesting early signaling roles in pod Ca nutrition. These findings provide insights into the roles of CDPK genes in plant development and stress responses, offering potential candidates for predicting calcium levels in peanut seeds.

Genome-Wide Identification of the CDPK Gene Family and Their Involvement in Taproot Cracking in Radish.

Taproot cracking, a severe and common physiological disorder, markedly reduces radish yield and commercial value. Calcium-dependent protein kinase (CDPK) plays a pivotal role in various plant developmental processes; however, its function in radish taproot cracking remains largely unknown. Here, 37 RsCDPK gene members were identified from the long-read radish genome "QZ-16". Phylogenetic analysis revealed that the CDPK members in radish, tomato, and Arabidopsis were clustered into four groups. Additionally, synteny analysis identified 13 segmental duplication events in the RsCDPK genes. Analysis of paraffin-embedded sections showed that the density and arrangement of fleshy taproot cortex cells are important factors that affect radish cracking. Transcriptome sequencing of the fleshy taproot cortex revealed 5755 differentially expressed genes (DEGs) (3252 upregulated and 2503 downregulated) between non-cracking radish "HongYun" and cracking radish "505". These DEGs were significantly enriched in plant hormone signal transduction, phenylpropanoid biosynthesis, and plant-pathogen interaction KEGG pathways. Furthermore, when comparing the 37 RsCDPK gene family members and RNA-seq DEGs, we identified six RsCDPK genes related to taproot cracking in radish. Soybean hairy root transformation experiments showed that RsCDPK21 significantly and positively regulates root length development. These findings provide valuable insights into the relationship between radish taproot cracking and RsCDPK gene function.

Identification of Dioscorea opposite Thunb. CDPK gene family reveals that DoCDPK20 is related to heat resistance.

Temperature affects the growth and yield of yam (Dioscorea opposite Thunb.), and calcium-dependent protein kinases (CDPKs) play an important role in the plant stress response. However, there has been a lack of system analyses of yam's CDPK gene family. In this study, 29 CDPK transcriptome sequences with complete open reading frames (ORFs) were identified from yam RNA sequencing data. The sequences were classified into four groups (I-VI) using phylogenetic analysis. Two DoCDPK genes were randomly selected from each group and the gene patterns of yam leaves were determined using quantitative real-time PCR (qRT-PCR) under high and low temperature stress in order to show their unique functions in mediating specific responses. Among them, DoCDPK20 was significantly induced in high temperatures. The pPZP221-DoCDPK20 was transformed into tobacco leaves using an agrobacterium-mediated method. Under high temperature stress, DoCDPK20 overexpression reduced photosynthesis and improved heat tolerance in transgenic tobacco. Our research offers meaningful perspectives into CDPK genes and new avenues for the genetic engineering and molecular breeding of yam.

Proteome-Level Investigation of Vitis amurensis Calli Transformed with a Constitutively Active, Ca2+-Independent Form of the Arabidopsis AtCPK1 Gene.

Calcium-dependent protein kinases (CDPKs) are one of the main Ca2+ decoders in plants. Among them, Arabidopsis thaliana AtCPK1 is one of the most studied CDPK genes as a positive regulator of plant responses to biotic and abiotic stress. The mutated form of AtCPK1, in which the autoinhibitory domain is inactivated (AtCPK1-Ca), provides constitutive kinase activity by mimicking a stress-induced increase in the Ca2+ flux. In the present study, we performed a proteomic analysis of Vitis amurensis calli overexpressing the AtCPK1-Ca form using untransformed calli as a control. In our previous studies, we have shown that the overexpression of this mutant form leads to the activation of secondary metabolism in plant cell cultures, including an increase in resveratrol biosynthesis in V. amurensis cell cultures. We analyzed upregulated and downregulated proteins in control and transgenic callus cultures using two-dimensional gel electrophoresis, and Matrix-assisted laser desorption and ionization time-of-flight mass spectrometry (MALDI-TOF). In calli transformed with AtCPK1-Ca, an increased amounts of pathogenesis-related proteins were found. A quantitative real-time PCR analysis confirmed this result.

Foliar Application of Chelated Sugar Alcohol Calcium Improves Photosynthesis and Tuber Quality under Drought Stress in Potatoes (Solanum tuberosum L.).

Drought stress is a major threat to sustainable crop production worldwide. Despite the positive role of calcium (Ca2+) in improving plant drought tolerance in different crops, little attention has been paid to its role in mitigating drought stress in potatoes. In the present study, we studied the effect of foliar chelated sugar alcohol calcium treatments on two potato cultivars with different drought responses applied 15 and 30 days after limiting soil moisture. The results showed that the foliar application of calcium treatments alleviated the SPAD chlorophyll loss of the drought-sensitive cultivar 'Atlantic' (Atl) and reduced the inhibition of photosynthetic parameters, leaf anatomy deformation, and MDA and H2O2 content of both cultivars under drought stress. The Ca2+ treatments changed the expression of several Calcium-Dependent Protein Kinase (StCDPK) genes involved in calcium sensing and signaling and significantly increased antioxidant enzyme activities, average tuber weight per plant, and tuber quality of both cultivars. We conclude that calcium spray treatments improved the drought tolerance of both potato cultivars and were especially effective for the drought-sensitive cultivar. The present work suggests that the foliar application of calcium is a promising strategy to improve commercial potato yields and the economic efficiency of potato production under drought stress conditions.

Whole-Genome Identification of Regulatory Function of CDPK Gene Families in Cold Stress Response for Prunus mume and Prunus mume var. Tortuosa.

Calcium-dependent protein kinases (CDPK) are known to mediate plant growth and development and respond to various environmental changes. Here, we performed whole-genome identification of CDPK families in cultivated and wild mei (Prunus mume). We identified 14 and 17 CDPK genes in P. mume and P. mume var. Tortuosa genomes, respectively. All 270 CPDK proteins were classified into four clade, displaying frequent homologies between these two genomes and those of other Rosaceae species. Exon/intron structure, motif and synteny blocks were conserved between P. mume and P. mume var. Tortuosa. The interaction network revealed all PmCDPK and PmvCDPK proteins is interacted with respiratory burst oxidase homologs (RBOHs) and mitogen-activated protein kinase (MAPK). RNA-seq data analysis of cold experiments show that cis-acting elements in the PmCDPK genes, especially PmCDPK14, are associated with cold hardiness. Our results provide and broad insights into CDPK gene families in mei and their role in modulating cold stress response in plants.

Identification, characterization, and expression analysis of PpCPK gene family in response to Alternaria-infected peach fruits collected from different geographical regions of China

Peach is a popular and economically important fruit tree highly susceptible to biotic and abiotic stresses, including diseases caused by fungi, bacteria and viruses, pests and insects that feed fruits and leaves. Alteranria is a deadly disease agent, a genus of fungi that can cause significant damage to plants and crops such as peaches. Here, we investigated the role of CPK genes against in Alternaria species. The calcium-dependent protein kinase (CPK) gene family plays a vital role in the biotic stress response of plants. In this study, we aimed to identify, characterize, and analyze the expression pattern of the PpCPK gene family in response to Alternaria-infected peach fruits collected from different geographical regions of China. Based on phylogenetic analysis, 17 PpCPK genes were identified and classified into seven groups. The physicochemical properties, gene structure, conserved domains, and motifs of the PpCPK genes were analyzed to understand their molecular characteristics further. The Ka/Ks ratio analysis revealed the evolutionary facets of duplication events of PpCPKs and predicted the divergence time between 23.5 and 43.6 million years ago. The promoter region analysis confirmed the presence of plant growth, light, stress, and hormone-related cis-elements in PpCPKs. Additionally, the expression pattern of PpCPKs was analyzed using real-time quantitative PCR in Alternaria-infected peach fruits from different regions of China. The results indicated a differential expression pattern of PpCPK genes responding to Alternaria infection, suggesting their potential role in defense against biotic stress. Overall, our study provides insights into the molecular mechanisms of PpCPK genes in response to biotic stress, which could aid in developing strategies for improving peach resistance to biotic stress.

Fine mapping and transcriptome sequencing reveal candidate genes conferring all-stage resistance to stripe rust on chromosome arm 1AL in Chinese wheat landrace AS1676

Stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), threatens wheat production worldwide, and resistant varieties tend to become susceptible after a period of cultivation owing to the variation of pathogen races. In this study, a new resistance gene against Pst race CYR34 was identified and predicted using the descendants of a cross between AS1676, a highly resistant Chinese landrace, and Avocet S, a susceptible cultivar. From a heterozygous plant from a F7 recombinant inbred line (RIL) population lacking the Yr18 gene, a near-isogenic line (NIL) population was developed to map the resistance gene. An all-stage resistance gene, YrAS1676, was identified on chromosome arm 1AL via bulked-segregant exome-capture sequencing. By analyzing a large NIL population consisting of 6537 plants, the gene was further mapped to the marker interval between KA1A_485.36 and KA1A_490.13, spanning 485.36–490.13 Mb on 1AL. A total of 66 annotated genes have been reported in this region. To characterize and predict the candidate gene(s), an RNA-seq was performed using NIL-R and NIL-S seedlings 3 days after CYR34 inoculation. Compared to NIL-S plants, NIL-R plants showed stronger immune reaction and higher expression levels of genes encoding pathogenesis-associated proteins. These differences may help to explain why NIL-R plants were more resistant to Pst race CYR34 than NIL-S plants. By combining fine-mapping and transcriptome sequencing, a calcium-dependent protein kinase gene was finally predicted as the potential candidate gene of YrAS1676. This gene contained a single-nucleotide polymorphism. The candidate gene was more highly expressed in NIL-R than in NIL-S plants. In field experiments with Pst challenge, the YrAS1676 genotype showed mitigation of disease damage and yield loss without adverse effects on tested agronomic traits. These results suggest that YrAS1676 has potential use in wheat stripe rust resistance breeding.

Genome-wide analysis of the CDPK gene family and their important roles response to cold stress in white clover

ABSTRACT Calcium-dependent protein kinases (CDPKs) are an important class of calcium-sensitive response proteins that play an important regulatory role in response to abiotic stresses. To date, little is known about the CDPK genes in white clover. White clover is a high-quality forage grass with high protein content, but it is susceptible to cold stress. Therefore, we performed a genome-wide analysis of the CDPK gene family in white clover and identified 50 members of the CDPK genes. Phylogenetic analysis using CDPKs from the model plant Arabidopsis divided the TrCDPK genes into four groups based on their sequence similarities. Motif analysis showed that TrCDPKs within the same group had similar motif compositions. Gene duplication analysis revealed the evolution and expansion of TrCDPK genes in white clover. Meanwhile, a genetic regulatory network (GRN) containing TrCDPK genes was reconstructed, and gene ontology (GO) annotation analysis of these functional genes showed that they contribute to signal transduction, cellular response to stimuli, and biological regulation, all of which are important processes in response to abiotic stresses. To determine the function of TrCDPK genes, we analyzed the RNA-seq dataset and found that most TrCDPK genes were highly up-regulated under cold stress, particularly in the early stages of cold stress. These results were validated by qRT-PCR experiments, implying that TrCDPK genes are involved in various gene regulatory pathways in response to cold stress. Our study may help to further investigate the function of TrCDPK genes and their role in response to cold stress, which is important for understanding the molecular mechanisms of cold tolerance in white clover and improving its cold tolerance.

Genome-wide Analysis of Drought-related Genes of “Alphonso” and “Tommy Atkins” Mango (Mangifera indica L.) Cultivars

Mango (Mangifera indica L.) is one of the most popular fruit crops grown and traded globally. Mango can withstand seasonal drought periods, especially during flowering. However, prolonged drought stress can cause the weakening of the plant and may encourage injuries and diseases caused by abiotic and biotic factors. With the availability of the mango genome, genomic studies related to drought adaptation of mango can now be extensively explored. In this study, genome- wide drought-related genes (DRGs) of mango were analyzed using the whole genome sequences (WGS) of “Alphonso” and “Tommy Atkins” cultivars (PRJNA487154 and PRJNA450143, respectively). A total of 261 and 257 DRGs were identified in the genome of “Alphonso” and “Tommy Atkins,” respectively, using BLASTP. Approximately 50% of these genes are involved in both molecular and physiological adaptations of mango to drought. Among the drought stress genes, the ABC transporter gene ABCG40 had the highest number of homologs in mango, followed by calcium-dependent protein kinase genes ZmCPK4, CPK21, and CDPK7, as well as the plasma membrane proton ATPase OST2. Gene ontology (GO) analysis of the DRGs revealed that protein binding, ATP binding, and mRNA binding are the most common molecular functions, whereas the predominant biological process of these DRGs is related to their response to water deprivation. Phylogenetic analysis showed that the drought-related proteins in “Alphonso” and “Tommy Atkins” are broadly clustered into seven and six major clades, respectively. This study, to date, offers the most comprehensive information on the genome-wide DRGs of the mango, which can strengthen marker-assisted breeding programs for drought tolerance in mango and other related fruit trees, as well as the future incorporation of favorable alleles toward improving the overall agronomic characteristics of this Philippine important fruit crop.

Comparative transcriptome analysis provides novel insights into molecular response of salt-tolerant and sensitive polyembryonic mango genotypes to salinity stress at seedling stage.

Increased soil salinity in the recent years has adversely affected the productivity of mango globally. Extending the cultivation of mango in salt affected regions warrants the use of salinity tolerant/resistant rootstocks. However, the lack of sufficient genomic and transcriptomic information impedes comprehensive research at the molecular level. We employed RNA sequencing-based transcriptome analysis to gain insight into molecular response to salt stress by using two polyembryonic mango genotypes with contrasting response to salt stress viz., salt tolerant Turpentine and salt susceptible Mylepelian. RNA sequencing by Novaseq6000 resulted in a total of 2795088, 17535948, 7813704 and 5544894 clean reads in Mylepelian treated (MT), Mylepelian control (MC), Turpentine treated (TT) and Turpentine control (TC) respectively. In total, 7169 unigenes annotated against all the five public databases, including NR, NT, PFAM, KOG, Swissport, KEGG and GO. Further, maximum number of differentially expressed genes were found between MT and MC (2106) followed by MT vs TT (1158) and TT and TC (587). The differentially expressed genes under different treatment levels included transcription factors (bZIP, NAC, bHLH), genes involved in Calcium-dependent protein kinases (CDPKs), ABA biosynthesis, Photosynthesis etc. Expression of few of these genes was experimentally validated through quantitative real-time PCR (qRT-PCR) and contrasting expression pattern of Auxin Response Factor 2 (ARF2), Late Embryogenesis Abundant (LEA) and CDPK genes were observed between Turpentine and Mylepelian. The results of this study will be useful in understanding the molecular mechanism underlying salt tolerance in mango which can serve as valuable baseline information to generate new targets in mango breeding for salt tolerance.

Tinospora crispa Ethanolic Extract Downregulates Protein Kinase Genes Expression and Activity during Toxoplasma gondii Infection: A Prospective Drug Target for Lytic Cycle Inhibition

Infection with Toxoplasma gondii remains widespread among humans and animals as water, soil and food continue to serve as the major carriers of the sporulated oocyst. The infection is poorly controlled due to lack of a potent vaccine against the parasite, and the current medication presents severe side effects on the host, less efficacy on the parasite and is accompanied by the potential development of resistance by the parasite. The aim of this study was to evaluate the in vitro activities of ethanolic extract of Tinospora crispa (EETC) on protein kinases involved in the lytic cycle of T. gondii infection in Vero cell line. The EETC was obtained through the maceration of dried stem powder. Vero cells infected with the RH strain of T. gondii were used to evaluate the inhibitory effect of EETC against T. gondii calcium dependent protein kinase (CDPK) genes and microneme proteins (MIC) that are essential for host cell invasion and intracellular replication of the tachyzoite. Gene expression profiling of CDPK genes was determined through quantitative real-time PCR (qPCR) after 24 h of treatment. The expression of microneme protein was determined through western blot technique. The RT-qPCR revealed downregulation of most protein kinase (PK) genes after treatment with EETC. The expression of CDPK1, PKG, CDPK3, CDPK6, CDPK7 genes that participate in the lytic cycle of T. gondii infection was downregulated. The expression of the TgMIC1 and TgMIC2 proteins were observed to have decreased in both 4 and 24 h post-infection treatment models. This study showed that EETC contains promising drug candidates effective against T. gondii that can target the protein kinase genes involved in the lytic cycle of the parasite to prevent disease progression. HIGHLIGHTS This study explored the possible mechanism of action of phytochemicals on Toxoplasma gondii parasite-host cell invasion and intracellular replication The study evaluated the effects of ethanolic extract of Tinospora crispa Miers on protein kinase genes that are involved in host cell invasion The target protein kinase genes were downregulated which prevents the secretion of microneme proteins Inhibition of microneme secretion prevents host cell invasion and intracellular replication by Toxoplasma gondi GRAPHICAL ABSTRACT

Genome-wide identification of the CPK gene family in wheat (Triticum aestivum L.) and characterization of TaCPK40 associated with seed dormancy and germination

Calcium-dependent protein kinases (CPKs), important sensors of calcium signals, play an essential role in plant growth, development, and stress responses. Although the CPK gene family has been characterized in many plants, the functions of the CPK gene family in wheat, including their relationship to seed dormancy and germination, remain unclear. In this study, we identified 84 TaCPK genes in wheat (TaCPK1-84). According to their phylogenetic relationship, they were divided into four groups (I-IV). TaCPK genes in the same group were found to have similar gene structures and motifs. Chromosomal localization indicated that TaCPK genes were unevenly distributed across 21 wheat chromosomes. TaCPK gene expansion occurred through segmental duplication events and underwent strong negative selection. A large number of cis-regulatory elements related to light response, phytohormone response, and abiotic stress response were identified in the upstream promoter sequences of TaCPK genes. TaCPK gene expression was found to be tissue- and growth-stage-diverse. Analysis of the expression patterns of several wheat varieties with contrasting seed dormancy and germination phenotypes resulted in the identification of 11 candidate genes (TaCPK38/-40/-43/-47/-50/-60/-67/-70/-75/-78/-80) which are likely associated with seed dormancy and germination. The ectopic expression of TaCPK40 in Arabidopsis promoted seed germination and reduced abscisic acid (ABA) sensitivity during germination, indicating that TaCPK40 negatively regulates seed dormancy and positively regulates seed germination. These findings advance our understanding of the multifaceted functions of CPK genes in seed dormancy and germination, and provide potential candidate genes for controlling wheat seed dormancy and germination.

Identification of CDPKs involved in TaNOX7 mediated ROS production in wheat.

As the critical sensors and decoders of calcium signal, calcium-dependent protein kinase (CDPK) has become the focus of current research, especially in plants. However, few resources are available on the properties and functions of CDPK gene family in Triticum aestivum (TaCDPK). Here, a total of 79 CDPK genes were identified in the wheat genome. These TaCDPKs could be classified into four subgroups on phylogenesis, while they may be classified into two subgroups based on their tissue and organ-spatiotemporal expression profiles or three subgroups according to their induced expression patterns. The analysis on the signal network relationships and interactions of TaCDPKs and NADPH (reduced nicotinamide adenine dinucleotide phosphate oxidases, NOXs), the key producers for reactive oxygen species (ROS), showed that there are complicated cross-talks between these two family proteins. Further experiments demonstrate that, two members of TaCDPKs, TaCDPK2/4, can interact with TaNOX7, an important member of wheat NOXs, and enhanced the TaNOX7-mediated ROS production. All the results suggest that TaCDPKs are highly expressed in wheat with distinct tissue or organ-specificity and stress-inducible diversity, and play vital roles in plant development and response to biotic and abiotic stresses by directly interacting with TaNOXs for ROS production.

Genome-wide survey of Calcium-Dependent Protein Kinases (CPKs) in five Brassica species and identification of CPKs induced by Plasmodiophora brassicae in B. rapa, B. oleracea, and B. napus.

Calcium-dependent protein kinase (CPK) is a class of Ser/Thr protein kinase that exists in plants and some protozoa, possessing Ca2+ sensing functions and kinase activity. To better reveal the roles that Brassica CPKs played during plant response to stresses, five Brassica species, namely Brassica rapa (B. rapa), Brassica nigra (B. nigra), Brassica oleracea (B. oleracea), Brassica juncea (B. juncea), and Brassica napus (B. napus) were selected and analyzed. In total, 51 BraCPK, 56 BniCPK, 56 BolCPK, 88 BjuCPK, and 107 BnaCPK genes were identified genome wide and phylogenetics, chromosomal mapping, collinearity, promoter analysis, and biological stress analysis were conducted. The results showed that a typical CPK gene was constituted by a long exon and tandem short exons. They were unevenly distributed on most chromosomes except chromosome A08 in B. napus and B. rapa, and almost all CPK genes were located on regions of high gene density as non-tandem form. The promoter regions of BraCPKs, BolCPKs, and BnaCPKs possessed at least three types of cis-elements, among which the abscisic acid responsive-related accounted for the largest proportion. In the phylogenetic tree, CPKs were clustered into four primary groups, among which group I contained the most CPK genes while group IV contained the fewest. Some clades, like AT5G23580.1(CPK12) and AT2G31500.1 (CPK24) contained much more gene members than others, indicating a possibility that gene expansion occurred during evolution. Furthermore, 4 BraCPKs, 14 BolCPKs, and 31 BnaCPKs involved in the Plasmodiophora brassicae (P. brassicae) defense response in resistant (R) or susceptible (S) materials were derived from online databases, leading to the discovery that some R-specific induced CPKs, such as BnaC02g08720D, BnaA03g03800D, and BolC04g018270.2J.m1 might be ideal candidate genes for P. brassicae resistant research. Overall, these results provide valuable information for research on the function and evolution of CDK genes.

ABA-Dependent Regulation of Calcium-Dependent Protein Kinase Gene GmCDPK5 in Cultivated and Wild Soybeans.

Calcium-dependent protein kinases (CDPKs) regulate plant development and stress responses. However, the interaction of these protein kinases with the abscisic acid (ABA) stress hormone signalling system has not been studied in detail. In Arabidopsis, AtCPK1 plays an important role in the acclimation of plants to environmental stresses. Phylogenetic and molecular analyses showed that, among 50 isoforms of Glycine max (L.) Merrill CDPKs, the GmCDPK27/GmCDPK48, GmCDPK5/GmCDPK24, and GmCDPK10/GmCDPK46 paralogous pairs were the isoforms most related to AtCDPK1. We investigated the expression of the corresponding six GmCDPKs genes during treatment with cold, heat, and salt stress. Wild soybean was the most resistant to stresses, and among the three cultivars studied (Sfera, Hodgson, and Hefeng25), Sfera was close to the wild type in terms of resistance. GmCDPK5 and GmCDPK10 were the most responsive to stress treatments, especially in wild soybean, compared with cultivars. Among the studied GmCDPK isoforms, only GmCDPK5 expression increased after treatment with abscisic acid (ABA) in a dose- and time-dependent manner. Targeted LC-MS/MS analysis of endogenous ABA levels showed that wild soybean and Sfera had nearly twice the ABA content of Hodgson and Hefeng25. An analysis of the expression of marker genes involved in ABA biosynthesis showed that GmNCED1-gene-encoding 9-cis-epoxycarotenoid dioxygenase 1 is induced to the greatest extent in wild soybean and Sfera under salt, cold, and heat exposure. Our data established a correlation between the induction of GmCDPK5 and ABA biosynthesis genes. GmCDPK5 is an interesting target for genetic and bioengineering purposes and can be used for genetic editing, overexpression, or as a marker gene in soybean varieties growing under unfavourable conditions.

A calcium-dependent protein kinase gene SpCPK33 from Solanum pennellii associated with increased cold tolerance in tomato

Calcium-dependent protein kinases (CDPKs, CPKs) represent a vital class of calcium sensors, which play a crucial role in plant growth, development and adaption to complex environmental stresses. Wild species tend to exhibit greater tolerance than cultivated species under environmental stress. Here, we isolated a calcium-dependent protein kinase gene SpCPK33 located primarily on the plasma membrane of abiotic-resistant species (Solanum pennellii LA0716). It was highly expressed in stems and leaves and was also induced by cold stress. Compared with WT plants, the overexpression of SpCPK33 in cultivated tomato (cv M82) enhanced its tolerance to cold stress. Transgenic lines demonstrated strong vitality under low temperature treatment. Moreover, the levels of malondialdehyde (MDA) and reactive oxygen species (ROS) were decreased in SpCPK33-overexpressing plants. The activities of antioxidant enzymes and the levels of osmotic regulatory substances were higher. The transcript levels of cold stress-related genes were up-regulated. In summary, the results indicate that SpCPK33-overexpressing transgenic plants experience less severe chilling injury under cold stress, and improved tomato cold tolerance by scavenging ROS accumulation and modulating the expression of stress-related genes.