Genome-wide identification of mitogen-activated protein kinase (MAPK) cascade and expression profiling of CmMAPKs in melon (Cucumis melo L.).
ABSTRACT: Mitogen-activated protein kinase (MAPK) is a form of serine/threonine protein kinase that activated by extracellular stimulation acting through the MAPK cascade (MAPKKK-MAPKK-MAPK). The MAPK cascade gene family, an important family of protein kinases, plays a vital role in responding to various stresses and hormone signal transduction processes in plants. In this study, we identified 14 CmMAPKs, 6 CmMAPKKs and 64 CmMAPKKKs in melon genome. Based on structural characteristics and a comparison of phylogenetic relationships of MAPK gene families from Arabidopsis, cucumber and watermelon, CmMAPKs and CmMAPKKs were categorized into 4 groups, and CmMAPKKKs were categorized into 3 groups. Furthermore, chromosome location revealed an unevenly distribution on chromosomes of MAPK cascade genes in melon, respectively. Eventually, qRT-PCR analysis showed that all 14 CmMAPKs had different expression patterns under drought, salt, salicylic acid (SA), methyl jasmonate (MeJA), red light (RL), and Podosphaera xanthii (P. xanthii) treatments. Overall, the expression levels of CmMAPK3 and CmMAPK7 under different treatments were higher than those in control. Our study provides an important basis for future functional verification of MAPK genes in regulating responses to stress and signal substance in melon.
Project description:The calcium-dependent protein kinase (CDPK) is a ser/thr protein kinase that plays vital roles in plant growth, development, and responses to multiple stresses. Despite an important member of the stress responsive gene family, little is known about the evolutionary history and expression patterns of CDPK genes in melon. Herein, a total of 18 CDPK genes and 7 CDPK-related protein kinases (CRK) genes were identified in the melon genome via bioinformatic analysis, which were unevenly distributed across eleven chromosomes with an apparent exception for chromosome 3. Comparative syntenic analysis between Cucumis melo L. and Arabidopsis thaliana revealed that 13 CmCDPKs and 19 AtCPKs existed in 20 corresponding syntenic blocks. In addition, based on gene structure and phylogenetic analyses, all CmCDPKs were divided into four groups (CDPK I-IV) and CmCRKs clustered into one group (CRK I). Interestingly, group CDPK IV was clearly distinct from the other three CDPK groups, but clustered with CRK I on the phylogenetic tree, implying their origination from a common ancestor. Furthermore, CmCDPKand CmCRK genes were differentially expressed in response to various stimuli, such as biotic stress (Podosphaera xanthii), abiotic stress (salt and cold), and hormone (abscisic acid) treatment. To our knowledge, this is the first report on CDPK and CRK gene families in melon, which provides a basic foundation for functional characterizations of CmCDPK and CmCRK genes in the future.
Project description:The fungal kingdom displays an extraordinary diversity of lifestyles, developmental processes, and ecological niches. The MAPK (mitogen-activated protein kinase) cascade consists of interlinked MAPKKK, MAPKK, and MAPK, and collectively such cascades play pivotal roles in cellular regulation in fungi. However, the mechanism by which evolutionarily conserved MAPK cascades regulate diverse output responses in fungi remains unknown. Here we identified the full complement of MAPK cascade components from 231 fungal species encompassing 9 fungal phyla. Using the largest data set to date, we found that MAPK family members could have two ancestors, while MAPKK and MAPKKK family members could have only one ancestor. The current MAPK, MAPKK, and MAPKKK subfamilies resulted from duplications and subsequent subfunctionalization during the emergence of the fungal kingdom. However, the gene structure diversification and gene expansion and loss have resulted in significant diversity in fungal MAPK cascades, correlating with the evolution of fungal species and lifestyles. In particular, a distinct evolutionary trajectory of MAPK cascades was identified in single-celled fungi in the Saccharomycetes. All MAPK, MAPKK, and MAPKKK subfamilies expanded in the Saccharomycetes; genes encoding MAPK cascade components have a similar exon-intron structure in this class that differs from those in other fungi.
Project description:The mitogen-activated protein kinase (MAPK) cascade, which is a major signal transduction pathway widely distributed in eukaryotes, has an important function in plant development and stress responses. However, less information is known regarding the MAPKKK and MAPKK gene families in the important fruit crop banana. In this study, 10 MAPKK and 77 MAPKKK genes were identified in the banana genome, and were classified into 4 and 3 subfamilies respectively based on phylogenetic analysis. Majority of MAPKKK and MAPKK genes in the same subfamily shared similar gene structures and conserved motifs. The comprehensive transcriptome analysis indicated that MAPKKK-MAPKK genes is involved in tissue development, fruit development and ripening, and response to abiotic stress of drought, cold and salt in two banana genotypes. Interaction networks and co-expression assays demonstrated that MAPK signaling cascade mediated network participates in multiple stress signaling, which was strongly activated in Fen Jiao (FJ). The findings of this study advance understanding of the intricately transcriptional control of MAPKKK-MAPKK genes and provide robust candidate genes for further genetic improvement of banana.
Project description:In plants, lipoxygenases (LOXs) play a crucial role in biotic and abiotic stresses. In our previous study, five 13-LOX genes of oriental melon were regulated by abiotic stress but it is unclear whether the 9-LOX is involved in biotic and abiotic stresses. The promoter analysis revealed that CmLOX09 (type of 9-LOX) has hormone elements, signal substances, and stress elements. We analyzed the expression of CmLOX09 and its downstream genes-CmHPL and CmAOS-in the leaves of four-leaf stage seedlings of the oriental melon cultivar "Yumeiren" under wound, hormone, and signal substances. CmLOX09, CmHPL, and CmAOS were all induced by wounding. CmLOX09 was induced by auxin (indole acetic acid, IAA) and gibberellins (GA3); however, CmHPL and CmAOS showed differential responses to IAA and GA3. CmLOX09, CmHPL, and CmAOS were all induced by hydrogen peroxide (H2O2) and methyl jasmonate (MeJA), while being inhibited by abscisic acid (ABA) and salicylic acid (SA). CmLOX09, CmHPL, and CmAOS were all induced by the powdery mildew pathogen Podosphaera xanthii. The content of 2-hexynol and 2-hexenal in leaves after MeJA treatment was significantly higher than that in the control. After infection with P. xanthii, the diseased leaves of the oriental melon were divided into four levels-levels 1, 2, 3, and 4. The content of jasmonic acid (JA) in the leaves of levels 1 and 3 was significantly higher than that in the level 0 leaves. In summary, the results suggested that CmLOX09 might play a positive role in the response to MeJA through the hydroperoxide lyase (HPL) pathway to produce C6 alcohols and aldehydes, and in the response to P. xanthii through the allene oxide synthase (AOS) pathway to form JA.
Project description:The cucurbit powdery mildew elicited by Podosphaera xanthii is one of the most important limiting factors in cucurbit production. Our knowledge of the genetic and molecular bases underlying the physiological processes governing this disease is very limited. We used RNA-sequencing to identify differentially expressed genes in leaves of Cucumis melo upon inoculation with P. xanthii, using RNA samples obtained at different time points during the early stages of infection and their corresponding uninfected controls. In parallel, melon plants were phenotypically characterized using imaging techniques. We found a high number of differentially expressed genes (DEGs) in infected plants, which allowed for the identification of many plant processes that were dysregulated by the infection. Among those, genes involved in photosynthesis and related processes were found to be upregulated, whereas genes involved in secondary metabolism pathways, such as phenylpropanoid biosynthesis, were downregulated. These changes in gene expression could be functionally validated by chlorophyll fluorescence imaging and blue-green fluorescence imaging analyses, which corroborated the alterations in photosynthetic activity and the suppression of phenolic compound biosynthesis. The powdery mildew disease in melon is a consequence of a complex and multifaceted process that involves the dysregulation of many plant pathways such as primary and secondary metabolism.
Project description:The mitogen-activated protein kinase (MAPK) cascade consists of three types of reversibly phosphorylated kinases, namely, MAPK, MAPK kinase (MAPKK/MEK), and MAPK kinase kinase (MAPKKK/MEKK), playing important roles in plant growth, development, and defense response. The MAPK cascade genes have been investigated in detail in model plants, including Arabidopsis, rice, and tomato, but poorly characterized in cucumber (Cucumis sativus L.), a major popular vegetable in Cucurbitaceae crops, which is highly susceptible to environmental stress and pathogen attack.A genome-wide analysis revealed the presence of at least 14 MAPKs, 6 MAPKKs, and 59 MAPKKKs in the cucumber genome. Phylogenetic analyses classified all the CsMAPK and CsMAPKK genes into four groups, whereas the CsMAPKKK genes were grouped into the MEKK, RAF, and ZIK subfamilies. The expansion of these three gene families was mainly contributed by segmental duplication events. Furthermore, the ratios of non-synonymous substitution rates (Ka) and synonymous substitution rates (Ks) implied that the duplicated gene pairs had experienced strong purifying selection. Real-time PCR analysis demonstrated that some MAPK, MAPKK and MAPKKK genes are preferentially expressed in specific organs or tissues. Moreover, the expression levels of most of these genes significantly changed under heat, cold, drought, and Pseudoperonospora cubensis treatments. Exposure to abscisic acid and jasmonic acid markedly affected the expression levels of these genes, thereby implying that they may play important roles in the plant hormone network.A comprehensive genome-wide analysis of gene structure, chromosomal distribution, and evolutionary relationship of MAPK cascade genes in cucumber are present here. Further expression analysis revealed that these genes were involved in important signaling pathways for biotic and abiotic stress responses in cucumber, as well as the response to plant hormones. Our first systematic description of the MAPK, MAPKK, and MAPKKK families in cucumber will help to elucidate their biological roles in plant.
Project description:Mitogen-activated protein kinase (MAPK) cascade signal transduction modules play crucial roles in regulating many biological processes in plants. These cascades are composed of three classes of hierarchically organized protein kinases, MAPKKKs, MAPKKs and MAPKs. Here, we analyzed gene retention, phylogenetic, evolution and expression patterns of MAPK cascade genes in Brassica rapa. We further found that the MAPK branches, classes III and IV, appeared after the split of bryophytes and green algae after analyzing the MAPK cascade genes in 8 species, and their rapid expansion led to the great size of the families of MAPKs. In contrast, the ancestral class I subfamily of MAPKK gene families have been highly conserved from algae to angiosperms. For the MAPKKK family, the MEKK and Raf subfamily share a common evolutionary origin, and Raf plays a major role in the expansion of the MAPKKK gene family. The cis-elements and interaction network analyses showed the important function of MAPK cascade genes in development and stress responses in B. rapa. This study provides a solid foundation for molecular evolution analyses of MAPK cascade genes.
Project description:Mitogen-activated protein kinase (MAPK) cascades have important functions in plant growth, development, and response to various stresses. The MAPKK and MAPKKK gene families in tomato have never been systematically analyzed. In this study, we performed a genome-wide analysis of the MAPKK and MAPKKK gene families in tomato and identified 5 MAPKK genes and 89 MAPKKK genes. Phylogenetic analyses of the MAPKK and MAPKKK gene families showed that all the MAPKK genes formed four groups (groups A, B, C, and D), whereas all the MAPKKK genes were classified into three subfamilies, namely, MEKK, RAF, and ZIK. Evolutionary analysis showed that whole genome or chromosomal segment duplications were the main factors responsible for the expansion of the MAPKK and MAPKKK gene families in tomato. Quantitative real-time RT-PCR analysis showed that the majority of MAPKK and MAPKKK genes were expressed in all tested organs with considerable differences in transcript levels indicating that they might be constitutively expressed. However, the expression level of most of these genes changed significantly under heat, cold, drought, salt, and Pseudomonas syringae treatment. Furthermore, their expression levels exhibited significant changes in response to salicylic acid and indole-3-acetic acid treatment, implying that these genes might have important roles in the plant hormone network. Our comparative analysis of the MAPKK and MAPKKK families would improve our understanding of the evolution and functional characterization of MAPK cascades in tomato.
Project description:The major components of the mitogen-activated protein kinase (MAPK) cascades are MAPK, MAPK kinase (MAPKK), and MAPKK kinase (MAPKKK). Recent rapid progress in identifying members of MAPK cascades suggests that a number of such signaling pathways exist in cells. To date, however, how the specificity and efficiency of the MAPK cascades is maintained is poorly understood. Here, we have identified a novel mouse protein, termed Jun N-terminal protein kinase (JNK)/stress-activated protein kinase-associated protein 1 (JSAP1), by a yeast two-hybrid screen, using JNK3 MAPK as the bait. Of the mammalian MAPKs tested (JNK1, JNK2, JNK3, ERK2, and p38alpha), JSAP1 preferentially coprecipitated with the JNKs in cotransfected COS-7 cells. JNK3 showed a higher binding affinity for JSAP1, compared with JNK1 and JNK2. In similar cotransfection studies, JSAP1 also interacted with SEK1 MAPKK and MEKK1 MAPKKK, which are involved in the JNK cascades. The regions of JSAP1 that bound JNK, SEK1, and MEKK1 were distinct from one another. JNK and MEKK1 also bound JSAP1 in vitro, suggesting that these interactions are direct. In contrast, only the activated form of SEK1 associated with JSAP1 in cotransfected COS-7 cells. The unstimulated SEK1 bound to MEKK1; thus, SEK1 might indirectly associate with JSAP1 through MEKK1. Although JSAP1 coprecipitated with MEK1 MAPKK and Raf-1 MAPKKK, and not MKK6 or MKK7 MAPKK, in cotransfected COS-7 cells, MEK1 and Raf-1 do not interfere with the binding of SEK1 and MEKK1 to JSAP1, respectively. Overexpression of full-length JSAP1 in COS-7 cells led to a considerable enhancement of JNK3 activation, and modest enhancement of JNK1 and JNK2 activation, by the MEKK1-SEK1 pathway. Deletion of the JNK- or MEKK1-binding regions resulted in a significant reduction in the enhancement of the JNK3 activation in COS-7 cells. These results suggest that JSAP1 functions as a scaffold protein in the JNK3 cascade. We also discuss a scaffolding role for JSAP1 in the JNK1 and JNK2 cascades.
Project description:Mitogen-activated protein kinases (MAPKs) are activated through cascades or modules consisting of a MAPK, a MAPK kinase (MAPKK), and a MAPKK kinase (MAPKKK). Investigating the molecular basis of activation of the c-Jun N-terminal kinase (JNK) subgroup of MAPK by the MAPKKK MEKK2, we found that strong and specific JNK1 activation by MEKK2 was mediated by the MAPKK JNK kinase 2 (JNKK2) rather than by JNKK1 through formation of a tripartite complex consisting of MEKK2, JNKK2, and JNK1. No scaffold protein was required for the MEKK2-JNKK2-JNK1 tripartite-complex formation. Expression of JNK1, JNKK2, and MEKK2 significantly augmented the coprecipitation of, respectively, MEKK2-JNKK2, MEKK2-JNK1, and JNKK2-JNK1, indicating that the interaction of MEKK2, JNKK2, and JNK1 is synergistic. Finally, the JNK1 was activated more efficiently in the MEKK2-JNKK2-JNK1 complex than was the JNK1 excluded from the complex. Thus, formation of a signaling complex through synergistic interaction of a MAPKKK, a MAPKK, and a MAPK molecule like MEKK2-JNKK2-JNK1 is likely to be responsible for the efficient, specific flow of information via MAPK cascades.