拟南芥AtMEKK1的结构特征和信号转导功能

促分裂原活化蛋白激酶（MAPK）级联途径是真核生物中高度保守的信号通路。MAPK级联途径由MAPKs、MAPKKs和MAPKKKs组成,通过MAPKKK→MAPKK→MAPK的逐级磷酸化传递细胞信号。AtMEKK1是拟南芥MAPKKK家族中的一员,是目前研究较为详细的MAPKKK。本文就AtMEKK1的结构特征、生理功能、信号转导中的＂交谈＂及其复杂性进行综述,旨在探讨植物MAPKKK的信号转导作用。     

RNA-Seq Analysis Reveals MAPKKK Family Members Related to Drought Tolerance in Maize

The mitogen-activated protein kinase (MAPK) cascade is an evolutionarily conserved signal transduction pathway that is involved in plant development and stress responses. As the first component of this phosphorelay cascade, mitogen-activated protein kinase kinase kinases (MAPKKKs) act as adaptors linking upstream signaling steps to the core MAPK cascade to promote the appropriate cellular responses; however, the functions of MAPKKKs in maize are unclear. Here, we identified 71 MAPKKK genes, of which 14 were novel, based on a computational analysis of the maize (Zea mays L.) genome. Using an RNA-seq analysis in the leaf, stem and root of maize under well-watered and drought-stress conditions, we identified 5,866 differentially expressed genes (DEGs), including 8 MAPKKK genes responsive to drought stress. Many of the DEGs were enriched in processes such as drought stress, abiotic stimulus, oxidation-reduction, and metabolic processes. The other way round, DEGs involved in processes such as oxidation, photosynthesis, and starch, proline, ethylene, and salicylic acid metabolism were clearly co-expressed with the MAPKKK genes. Furthermore, a quantitative real-time PCR (qRT-PCR) analysis was performed to assess the relative expression levels of MAPKKKs. Correlation analysis revealed that there was a significant correlation between expression levels of two MAPKKKs and relative biomass responsive to drought in 8 inbred lines. Our results indicate that MAPKKKs may have important regulatory functions in drought tolerance in maize.     

Genome-Wide Analysis of Mitogen-Activated Protein Kinase Gene Family in Maize

Mitogen-activated protein kinase (MAPK) cascades are universal signal transduction modules in plants. As the last component of the MAPK cascade (MAPKKK–MAPKK–MAPK), MAPK plays important roles in linking upstream kinases and downstream substrates. The MAPK proteins belong to a complex gene family in plants, with 20 MAPK genes in the Arabidopsis genome, 17 in the rice genome, and 21 in the poplar genome. Although the maize genome sequencing has been completed, no comprehensive study has been reported thus far for the MAPK gene family in maize. In this study, we identified 19 MAPK genes in maize. These ZmMPK genes belong to four groups (A–D) found in other plants. The phylogeny, chromosomal location, gene structure, and the functional relevancy of ZmMPK genes were analyzed. Moreover, we discuss the evolutionary divergence of MAPK genes in maize. Furthermore, we analyzed the expression profiles of ZmMPKs using the public microarray data and performed expression analyses in maize seedlings and adult plants. The data obtained from our study contribute to a better understanding of the complexity of MAPKs in plants and provide a useful reference for further functional analysis of MAPK genes in maize.     

植物MAPK信号转导组分的细胞定位与选择性剪接

促分裂原活化蛋白激酶(MAPK)级联途径在真核生物中是高度保守的,由MAPKs,MAPKKs,MAPKKKs组成,通过MAPKKK→MAPKK→MAPK逐级磷酸化传递细胞信号.已有大量研究表明,MAPK在植物响应生物与非生物胁迫,以及植物激素和细胞周期的信号转导中起重要作用.在植物响应各种逆境过程中激活的MAPK基因,细胞内的定位发生动态变化.选择性剪接是真核生物中调节基因表达的重要模式,能够影响蛋白的结合特性、胞内定位、酶的活性、蛋白的稳定性和翻译后的修饰.MAPK基因的选择性剪接能产生不同的转录异型并具有不同的亚细胞定位.本文综述这方面的研究进展.     

Plant Homologues of Components of MAPK (Mitogen-Activated Protein Kinase) Signal Pathways in Yeast and Animal Cells

As they respond to numerous extracellular and intracellularstimuli, plants develop various morphological features and thecapacity for a large variety of physiological processes duringtheir growth. If we are to understand the molecular basis ofsuch developments, we must elucidate the way in which signalsgenerated by such stimuli can be transduced into plant cellsand transmitted by cellular components to induce the appropriateterminal events. In yeast and animal systems, signal pathwaysthat are known collectively as MAPK (mitogen-activated proteinkinase) cascades have been shown to play a central role in thetransmission of various signals. The components of these pathwaysinclude the MAPK family, the activator kinases of the MAPK family(the MAPKK family) and the activator kinases of the MAPKK family(the MAPKKK family). The members of each respective family arestructurally conserved and signals are transmitted by similarphosphotransfer reactions at corresponding steps that are mediatedby a specific member of each family in turn. Both cDNAs andgenes that encode putative homologues of these components haverecently been isolated from plant sources. Some of them havebeen shown to be related not only structurally but also functionallyto members of the MAPK cascades of other organisms. These findingssuggest that plants have signal pathways that are analogousto the MAPK cascades in yeast and animal cells but it remainsto be proven that plant homologues do in fact constitute kinasecascades. Given the presence of so many homologues of MAPKsand MAPKKKs in a single plant species, namely, Arabidopsis thaliana,we can be fairly confident that the putative MAPK cascades areinvolved in various physiological processes in plants. (Received March 28, 1995; )     

Identification of three MAPKKKs forming a linear signaling pathway leading to programmed cell death in

ABSTRACT: BACKGROUND: The mitogen-activated protein kinase (MAPK) cascade is an evolutionarily ancient mechanism of signal transduction found in eukaryotic cells. In plants, MAPK cascades are associated with responses to various abiotic and biotic stresses such as plant pathogens. MAPK cascades function through sequential phosphorylation: MAPK kinase kinases (MAPKKKs) phosphorylate MAPK kinases (MAPKKs), and phosphorylated MAPKKs phosphorylate MAPKs. Of these three types of kinase, the MAPKKKs exhibit the most divergence in the plant genome. Their great diversity is assumed to allow MAPKKKs to regulate many specific signaling pathways in plants despite the relatively limited number of MAPKKs and MAPKs. Although some plant MAPKKKs, including the MAPKKKalpha of Nicotiana benthamiana (NbMAPKKKalpha), are known to play crucial roles in plant defense responses, the functional relationship among MAPKKK genes is poorly understood. Here, we performed a comparative functional analysis of MAPKKKs to investigate the signaling pathway leading to the defense response. RESULTS: We cloned three novel MAPKKK genes from N. benthamiana: NbMAPKKKbeta, NbMAPKKKgamma, and NbMAPKKKepsilon2. Transient overexpression of full-length NbMAPKKKbeta or NbMAPKKKgamma or their kinase domains in N. benthamiana leaves induced hypersensitive response (HR)-like cell death associated with hydrogen peroxide production. This activity was dependent on the kinase activity of the overexpressed MAPKKK. In addition, virus-induced silencing of NbMAPKKKbeta or NbMAPKKKgamma expression significantly suppressed the induction of programmed cell death (PCD) by viral infection. Furthermore, in epistasis analysis of the functional relationships among NbMAPKKKbeta, NbMAPKKKgamma, and NbMAPKKKalpha (previously shown to be involved in plant defense responses) conducted by combining transient overexpression analysis and virus-induced gene silencing, silencing of NbMAPKKKalpha suppressed cell death induced by the overexpression of the NbMAPKKKbeta kinase domain or of NbMAPKKKgamma, but silencing of NbMAPKKKbeta failed to suppress cell death induced by the overexpression of NbMAPKKKalpha or NbMAPKKKgamma. Silencing of NbMAPKKKgamma suppressed cell death induced by the NbMAPKKKbeta kinase domain but not that induced by NbMAPKKKalpha. CONCLUSIONS: These results demonstrate that in addition to NbMAPKKKalpha, NbMAPKKKbeta and NbMAPKKKgamma also function as positive regulators of PCD. Furthermore, these three MAPKKKs form a linear signaling pathway leading to PCD; this pathway proceeds from NbMAPKKKbeta to NbMAPKKKgamma to NbMAPKKKalpha.     

真核生物的MAPK级联信号传递途径

MAPK级联途径在真核生物细胞的信号传递过程中起着重要的作用.MAPK级联途径由MAPK、MAPKK和MAPKKK三类酶蛋白组成.这三类蛋白质的结构非常保守,通过磷酸化作用传递各种信号.在酵母和动、植物细胞中已经发现了一系列的MAPK级联途径成员,使真核生物的信号传递途径逐渐得到阐明.     

Signal convergence through the lenses of MAP kinases: paradigms of stress and hormone signaling in plants

Common mechanisms plants use to translate the external stimuli into cellular responses are the activation of mitogen-activated protein kinase (MAPK) cascade. These MAPK cascades are highly conserved in eukaryotes and consist of three subsequently acting protein kinases, MAP kinase kinase kinase (MAPKKK), MAP kinase kinase (MAPKK) and MAP kinase (MAPK) which are linked in various ways with upstream receptors and downstream targets. Plant MAPK cascades regulate numerous processes, including various environmental stresses, hormones, cell division and developmental processes. The number of MAPKKs in Arabidopsis and rice is almost half the number of MAPKs pointing important role of MAPKKs in integrating signals from several MAPKKKs and transducing signals to various MAPKs. The cross talks between different signal transduction pathways are concentrated at the level of MAPKK in the MAPK cascade. Here we discussed the insights into MAPKK mediated response to environmental stresses and in plant growth and development.     

植物丝裂原活化蛋白激酶级联信号转导通路研究进展

丝裂原活化蛋白激酶(MAPK)是酵母、动物和植物等真核生物中普遍存在和高度保守的一类信号转导通路,由MAPKKK、MAPKK和MAPK等3部分组成,在应对生物非生物胁迫、激素、细胞分裂调控及植物生长发育等过程中发挥重要作用。该文对近年来国内外有关MAPK级联通路的组成、在植株体内的生物学功能以及MAPK通路的失活进行了概述,旨在为今后MAPK通路介导的信号转导机制的研究提供参考依据。     

Soybean homologs of MPK4 negatively regulate defense responses and positively regulate growth and development

Mitogen-activated protein kinase (MAPK) cascades play important roles in disease resistance in model plant species such as Arabidopsis (Arabidopsis thaliana) and tobacco (Nicotiana tabacum). However, the importance of MAPK signaling pathways in the disease resistance of crops is still largely uninvestigated. To better understand the role of MAPK signaling pathways in disease resistance in soybean (Glycine max), 13, nine, and 10 genes encoding distinct MAPKs, MAPKKs, and MAPKKKs, respectively, were silenced using virus-induced gene silencing mediated by Bean pod mottle virus. Among the plants silenced for various MAPKs, MAPKKs, and MAPKKKs, those in which GmMAPK4 homologs (GmMPK4s) were silenced displayed strong phenotypes including stunted stature and spontaneous cell death on the leaves and stems, the characteristic hallmarks of activated defense responses. Microarray analysis showed that genes involved in defense responses, such as those in salicylic acid (SA) signaling pathways, were significantly up-regulated in GmMPK4-silenced plants, whereas genes involved in growth and development, such as those in auxin signaling pathways and in cell cycle and proliferation, were significantly down-regulated. As expected, SA and hydrogen peroxide accumulation was significantly increased in GmMPK4-silenced plants. Accordingly, GmMPK4-silenced plants were more resistant to downy mildew and Soybean mosaic virus compared with vector control plants. Using bimolecular fluorescence complementation analysis and in vitro kinase assays, we determined that GmMKK1 and GmMKK2 might function upstream of GmMPK4. Taken together, our results indicate that GmMPK4s negatively regulate SA accumulation and defense response but positively regulate plant growth and development, and their functions are conserved across plant species.     

The rice Raf-like MAPKKK OsILA1 confers broad-spectrum resistance to bacterial blight by suppressing the OsMAPKK4–OsMAPK6 cascade

Mitogen-activated protein kinase kinase kinase (MAPKKK) are the first components of MAPK cascades, which play pivotal roles in signaling during plant development and physiological processes. The genome of rice encodes 75 MAPKKKs, of which 43 are Raf-like MAPKKKs. The functions and action modes of most of the Raf-like MAPKKKs, whether they function as bona fide MAPKKKs and which are their downstream MAPKKs, are largely unknown. Here, we identified the osmapkkk43 mutant, which conferred broad-spectrum resistance to Xanthomonas oryzae pv. oryzae (Xoo), the destructive bacterial pathogen of rice. Oryza sativa (Os)MAPKKK43 encoding a Raf-like MAPKKK was previously known as Increased Leaf Angle 1 (OsILA1). Genetic analysis indicated that OsILA1 functioned as a negative regulator and acted upstream of the OsMAPKK4–OsMAPK6 cascade in rice–Xoo interactions. Unlike classical MAPKKKs, OsILA1 mainly phosphorylated the threonine 34 site at the N-terminal domain of OsMAPKK4, which possibly influenced the stability of OsMAPKK4. The N-terminal domain of OsILA1 is required for its homodimer formation and its full phosphorylation capacity. Taken together, our findings reveal that OsILA1 acts as a negative regulator of the OsMAPKK4–OsMAPK6 cascade and is involved in rice–Xoo interactions.     

OMTK1, a novel MAPKKK, channels oxidative stress signaling through direct MAPK interaction

In common with other eukaryotes, plants utilize mitogen-activated protein kinase (MAPK) cascades to mediate responses to a wide variety of stimuli. In contrast to other eukaryotes, plants have an unusually large number of MAPK components, such as more than 20 MAPKs, 10 MAPK kinases (MAPKKs), and 60 MAPKK kinases (MAPKKKs) in Arabidopsis (MAPK Group (2002) Trends Plant Sci. 7, 301-308). Presently it is mostly unknown how MAPK signaling specificity is generated in plants. Here we have isolated OMTK1 (oxidative stress-activated MAP triple-kinase 1), a novel MAPKKK from alfalfa (Medicago sativa). In plant protoplasts, OMTK1 showed basal kinase activity and was found to induce cell death. Among a panel of hormones and stresses tested, only H(2)O(2) was found to activate OMTK1. Out of four MAPKs, OMTK1 specifically activated MMK3 resulting in an increased cell death rate. Pull-down analysis between recombinant proteins indicated that OMTK1 directly interacts with MMK3 and that OMTK1 and MMK3 are part of a protein complex in vivo. These results indicate that OMTK1 plays a MAPK scaffolding role and functions in activation of H(2)O(2) -induced cell death in plants.     

A docking site determining specificity of Pbs2 MAPKK for Ssk2/Ssk22 MAPKKKs in the yeast HOG pathway

Mitogen-activated protein kinase (MAPK) cascades are conserved signaling modules composed of three sequentially activated kinases (MAPKKK, MAPKK and MAPK). Because individual cells contain multiple MAPK cascades, mechanisms are required to ensure the fidelity of signal transmission. In yeast, external high osmolarity activates the HOG (high osmolarity glycerol) MAPK pathway, which consists of two upstream branches (SHO1 and SLN1) and common downstream elements including the Pbs2 MAPKK and the Hog1 MAPK. The Ssk2/Ssk22 MAPKKKs in the SLN1 branch, when activated, exclusively phosphorylate the Pbs2 MAPKK. We found that this was due to an Ssk2/Ssk22-specific docking site in the Pbs2 N-terminal region. The Pbs2 docking site constitutively bound the Ssk2/Ssk22 kinase domain. Docking site mutations drastically reduced the Pbs2-Ssk2/Ssk22 interaction and hampered Hog1 activation by the SLN1 branch. Fusion of the Pbs2 docking site to a different MAPKK, Ste7, allowed phosphorylation of Ste7 by Ssk2/Ssk22. Thus, the docking site contributes to both the efficiency and specificity of signaling. During these analyses, we also found a nuclear export signal and a possible nuclear localization signal in Pbs2.     

植物中的MAPK及其在信号传导中的作用

促分裂原活化蛋白激酶(MAPKs)是一类存在于真核生物中的丝氨酸/苏氨酸蛋白激酶。同动物和酵母中MAPKs类似,植物中的MAPK级联途径也是由MAPKs、MAPKKs、MAPKKKs三种类型的激酶组成。植物细胞内受体接受外界刺激信号,然后依次磷酸化激活MAPKKKs、MAPKKs和MAPKs,并影响相关基因表达。目前已经从植物中分离到一些MAPKs、MAPKKs和MAPKKKs,它们参与了植物激素、生物胁迫及非生物胁迫等过程的信号传导。介绍了植物响应外界环境胁迫过程中,不同机制和因子对MAPKs级联途径的调控。     

The Expression Pattern of the Gene for NPK1 Protein Kinase Related to Mitogen-Activated Protein Kinase Kinase Kinase (MAPKKK) in a Tobacco Plant: Correlation with Cell Proliferation

Mitogen-activated protein kinase (MAPK) cascades consist ofmembers of three families of protein kinases: the MAPK family,the MAPK kinase family, and the MAPK kinase kinase (MAPKKK)family. Some of these cascades have been shown to play centralroles in the transmission of signals that control various cellularprocesses including cell proliferation. Protein kinase NPK1is a structural and functional tobacco homologue of MAPKKK,but its physiological function is yet unknown. In the presentstudy, we have investigated sites of expression of the NPK1gene in a tobacco plant and developmental and physiologicalcontrols of this expression. After germination, expression ofNPK1 was first detected in tips of a radicle and cotyledons,then in shoot and root apical meristems, surrounding tissuesof the apical meristems, primordia of lateral roots, and youngdeveloping organs. No expression was, however, observed in matureorgans. Incubation of discs from mature leaves of tobacco withboth auxin and cytokinin induced NPK1 expression before thedivision of cells. It was also induced at early stages of thedevelopment of primordia of lateral roots and adventitious roots.Thus, NPK1 expression appears to be tightly correlated withcell division or division competence. Even when an inhibitorof DNA synthesis was added during the germination or the inductionof lateral roots by auxin, NPK1 expression was detected. Theseresults showed that the NPK1 expression precedes DNA replication.We propose that NPK1 participates in a process involving thedivision of plant cells. (Received January 26, 1998; Accepted April 9, 1998)