RESUMO
Gravity controls directional growth of plants, and the classical starch-statolith hypothesis proposed more than a century ago postulates that amyloplast sedimentation in specialized cells initiates gravity sensing, but the molecular mechanism remains uncharacterized. The LAZY proteins are known as key regulators of gravitropism, and lazy mutants show striking gravitropic defects. Here, we report that gravistimulation by reorientation triggers mitogen-activated protein kinase (MAPK) signaling-mediated phosphorylation of Arabidopsis LAZY proteins basally polarized in root columella cells. Phosphorylation of LAZY increases its interaction with several translocons at the outer envelope membrane of chloroplasts (TOC) proteins on the surface of amyloplasts, facilitating enrichment of LAZY proteins on amyloplasts. Amyloplast sedimentation subsequently guides LAZY to relocate to the new lower side of the plasma membrane in columella cells, where LAZY induces asymmetrical auxin distribution and root differential growth. Together, this study provides a molecular interpretation for the starch-statolith hypothesis: the organelle-movement-triggered molecular polarity formation.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Plastídeos , Arabidopsis/fisiologia , Proteínas de Arabidopsis/metabolismo , Sensação Gravitacional , Raízes de Plantas/metabolismo , Plastídeos/metabolismo , Amido/metabolismo , Proteínas de Membrana/metabolismoRESUMO
Upon stress, eukaryotes typically reprogram their translatome through GCN2-mediated phosphorylation of the eukaryotic translation initiation factor, eIF2α, to inhibit general translation initiation while selectively translating essential stress regulators. Unexpectedly, in plants, pattern-triggered immunity (PTI) and response to other environmental stresses occur independently of the GCN2/eIF2α pathway. Here, we show that while PTI induces mRNA decapping to inhibit general translation, defense mRNAs with a purine-rich element ("R-motif") are selectively translated using R-motif as an internal ribosome entry site (IRES). R-motif-dependent translation is executed by poly(A)-binding proteins (PABPs) through preferential association with the PTI-activating eIFiso4G over the repressive eIF4G. Phosphorylation by PTI regulators mitogen-activated protein kinase 3 and 6 (MPK3/6) inhibits eIF4G's activity while enhancing PABP binding to the R-motif and promoting eIFiso4G-mediated defense mRNA translation, establishing a link between PTI signaling and protein synthesis. Given its prevalence in both plants and animals, the PABP/R-motif translation initiation module may have a broader role in reprogramming the stress translatome.
Assuntos
Fator de Iniciação Eucariótico 4G , Proteínas de Ligação a Poli(A) , Animais , Fator de Iniciação Eucariótico 4G/genética , Fator de Iniciação Eucariótico 4G/metabolismo , Fatores de Iniciação em Eucariotos/metabolismo , Proteínas de Ligação a Poli(A)/metabolismo , Biossíntese de Proteínas , Purinas , RNA Mensageiro/metabolismoRESUMO
Lateral roots (LRs) increase root surface area and allow plants greater access to soil water and nutrients. LR formation is tightly regulated by the phytohormone auxin. Whereas the transcription factor ETHYLENE-RESPONSIVE ELEMENT BINDING FACTOR13 (ERF13) prevents LR emergence in Arabidopsis (Arabidopsis thaliana), auxin activates MITOGEN-ACTIVATED PROTEIN KINASE14 (MPK14), which leads to ERF13 degradation and ultimately promotes LR emergence. In this study, we discovered interactions between ERF13 and the E3 ubiquitin ligases MOS4-ASSOCIATED COMPLEX 3A (MAC3A) and MAC3B. As MAC3A and MAC3B gradually accumulate in the LR primordium, ERF13 levels gradually decrease. We demonstrate that MAC3A and MAC3B ubiquitinate ERF13, leading to its degradation and accelerating the transition of LR primordia from stage IV to stage V. Auxin enhances the MAC3A and MAC3B interaction with ERF13 by facilitating MPK14-mediated ERF13 phosphorylation. In summary, this study reveals the molecular mechanism by which auxin eliminates the inhibitory factor ERF13 through the MPK14-MAC3A and MAC3B signaling module, thus promoting LR emergence.
RESUMO
When stressed, cells need to adapt their proteome to maintain protein homeostasis. This requires increased proteasome assembly. Increased proteasome assembly is dependent on increased production of proteasome assembly chaperones. In Saccharomyces cerevisiae, inhibition of the growth-promoting kinase complex TORC1 causes increased proteasome assembly chaperone translation, including that of Adc17. This is dependent upon activation of the mitogen-activated protein kinase (MAPK) Mpk1 and relocalisation of assembly chaperone mRNA to patches of dense actin. We show here that TORC1 inhibition alters cell wall properties to induce these changes by activating the cell wall integrity pathway through the Wsc1, Wsc3 and Wsc4 sensor proteins. We demonstrate that, in isolation, these signals are insufficient to drive protein expression. We identify that the TORC1-activated S6 kinase Sch9 must be inhibited as well. This work expands our knowledge on the signalling pathways that regulate proteasome assembly chaperone production.
Assuntos
Chaperonas Moleculares , Complexo de Endopeptidases do Proteassoma , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae , Transdução de Sinais , Parede Celular/metabolismo , Regulação Fúngica da Expressão Gênica , Chaperonas Moleculares/metabolismo , Chaperonas Moleculares/genética , Complexo de Endopeptidases do Proteassoma/metabolismo , Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genéticaRESUMO
The plant hormone auxin plays a key role to maintain root stem cell identity which is essential for root development. However, the molecular mechanism by which auxin regulates root distal stem cell (DSC) identity is not well understood. In this study, we revealed that the cell cycle factor DPa is a vital regulator in the maintenance of root DSC identity through multiple auxin signaling cascades. On the one hand, auxin positively regulates the transcription of DPa via AUXIN RESPONSE FACTOR 7 and ARF19. On the other hand, auxin enhances the protein stability of DPa through MITOGEN-ACTIVATED PROTEIN KINASE 3 (MPK3)/MPK6-mediated phosphorylation. Consistently, mutation of the identified three threonine residues (Thr10, Thr25, and Thr227) of DPa to nonphosphorylated form alanine (DPa3A) highly decreased the phosphorylation level of DPa, which decreased its protein stability and affected the maintenance of root DSC identity. Taken together, this study provides insight into the molecular mechanism of how auxin regulates root distal stem cell identity through the dual regulations of DPa at both transcriptional and posttranslational levels.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Divisão Celular , Regulação da Expressão Gênica de Plantas , Ácidos Indolacéticos/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Raízes de Plantas/metabolismo , Células-Tronco/metabolismoRESUMO
Drought stress inhibits plant growth and agricultural production. Improving plant instantaneous water use efficiency (iWUE), which is strictly regulated by stomata, is an effective way to cope with drought stress. However, the mechanisms of iWUE regulation are poorly understood. Through genetic screening for suppressors of mpk12-4, an Arabidopsis (Arabidopsis thaliana) mutant with a major iWUE quantitative trait locus gene MITOGEN-ACTIVATED PROTEIN KINASE12 deleted, we identified HIGH LEAF TEMPERATURE1 (HT1). Genetic interaction and physiological analyses showed that MPK12 controls iWUE through multiple modules in a high CO2-induced stomatal closing pathway that regulate SLOW ANION CHANNEL-ASSOCIATED1 (SLAC1) activity. HT1 acts downstream of MPK12, whereas OPEN STOMATA1 (OST1) and GUARD CELL HYDROGEN PEROXIDE-RESISTANT1 (GHR1) function downstream of HT1 by activating SLAC1 in iWUE. Photosynthetic-CO2 response curves and biomass analyses under different water-supply conditions showed that HT1 dysfunction improved iWUE and also increased plant growth capacity, and products of HT1 putative orthologs from Brassica (Brassica napus) and rice (Oryza sativa) exhibited functions similar to that of Arabidopsis HT1 in iWUE and the CO2-signaling pathway. Our study revealed the mechanism of MPK12-mediated iWUE regulation in Arabidopsis and provided insight into the internal relationship between iWUE and CO2 signaling in guard cells and a potential target for improving crop iWUE and drought tolerance.
RESUMO
The fidelity of a signaling pathway depends on its tight regulation in space and time. Extracellular signal-regulated kinase (ERK) controls wide-ranging cellular processes to promote organismal development and tissue homeostasis. ERK activation depends on a reversible dual phosphorylation on the TEY motif in its active site by ERK kinase (MEK) and dephosphorylation by DUSPs (dual specificity phosphatases). LIP-1, a DUSP6/7 homolog, was proposed to function as an ERK (MPK-1) DUSP in the Caenorhabditis elegans germline primarily because of its phenotype, which morphologically mimics that of a RAS/let-60 gain-of-function mutant (i.e., small oocyte phenotype). Our investigations, however, reveal that loss of lip-1 does not lead to an increase in MPK-1 activity in vivo. Instead, we show that loss of lip-1 leads to 1) a decrease in MPK-1 phosphorylation, 2) lower MPK-1 substrate phosphorylation, 3) phenocopy of mpk-1 reduction-of-function (rather than gain-of-function) allele, and 4) a failure to rescue mpk-1-dependent germline or fertility defects. Moreover, using diverse genetic mutants, we show that the small oocyte phenotype does not correlate with increased ectopic MPK-1 activity and that ectopic increase in MPK-1 phosphorylation does not necessarily result in a small oocyte phenotype. Together, these data demonstrate that LIP-1 does not function as an MPK-1 DUSP in the C. elegans germline. Our results caution against overinterpretation of the mechanistic underpinnings of orthologous phenotypes, since they may be a result of independent mechanisms, and provide a framework for characterizing the distinct molecular targets through which LIP-1 may mediate its several germline functions.
Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/enzimologia , Proteínas de Ciclo Celular/metabolismo , Células Germinativas/enzimologia , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteínas Tirosina Fosfatases/metabolismo , Animais , Proteínas de Caenorhabditis elegans/genética , Proteínas de Ciclo Celular/genética , Proliferação de Células , Ativação Enzimática , Mutação/genética , Oócitos/citologia , Oócitos/metabolismo , Estágio Paquíteno , Fenótipo , Fosforilação , Proteínas Tirosina Fosfatases/genética , Especificidade por Substrato , Complexo Sinaptonêmico/metabolismo , TemperaturaRESUMO
Energy production and metabolism are intimately linked to ecological and environmental constraints across the tree of life. In plants, which depend on sunlight to produce energy, the link between primary metabolism and the environment is especially strong. By governing CO2 uptake for photosynthesis and transpiration, leaf pores, or stomata, couple energy metabolism to the environment and determine productivity and water-use efficiency (WUE). Although evolution is known to tune physiological traits to the local environment, we lack knowledge of the specific links between molecular and evolutionary mechanisms that shape this process in nature. Here, we investigate the evolution of stomatal conductance and WUE in an Arabidopsis population that colonized an island with a montane cloud scrubland ecosystem characterized by seasonal drought and fog-based precipitation. We find that stomatal conductance increases and WUE decreases in the colonizing population relative to its closest outgroup population from temperate North Africa. Genome-wide association mapping reveals a polygenic basis of trait variation, with a substantial contribution from a nonsynonymous single-nucleotide polymorphism in MAP KINASE 12 (MPK12 G53R), which explains 35% of the phenotypic variance in WUE in the island population. We reconstruct the spatially explicit evolutionary history of MPK12 53R on the island and find that this allele increased in frequency in the population due to positive selection as Arabidopsis expanded into the harsher regions of the island. Overall, these findings show how adaptation shaped quantitative eco-physiological traits in a new precipitation regime defined by low rainfall and high humidity.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/genética , Arabidopsis/metabolismo , Ecossistema , Estudo de Associação Genômica Ampla , Proteínas de Arabidopsis/genética , Folhas de Planta , Fotossíntese/genética , Água/metabolismo , Genômica , SecasRESUMO
The phytohormone auxin controls plant growth and development via TIR1-dependent protein degradation of canonical AUX/IAA proteins, which normally repress the activity of auxin response transcription factors (ARFs). IAA33 is a non-canonical AUX/IAA protein lacking a TIR1-binding domain, and its role in auxin signaling and plant development is not well understood. Here, we show that IAA33 maintains root distal stem cell identity and negatively regulates auxin signaling by interacting with ARF10 and ARF16. IAA33 competes with the canonical AUX/IAA repressor IAA5 for binding to ARF10/16 to protect them from IAA5-mediated inhibition. In contrast to auxin-dependent degradation of canonical AUX/IAA proteins, auxin stabilizes IAA33 protein via MITOGEN-ACTIVATED PROTEIN KINASE 14 (MPK14) and does not affect IAA33 gene expression. Taken together, this study provides insight into the molecular functions of non-canonical AUX/IAA proteins in auxin signaling transduction.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Proteínas de Ligação a DNA/metabolismo , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Ácidos Indolacéticos/farmacologia , Proteínas Nucleares/metabolismo , Plantas Geneticamente Modificadas/metabolismo , Arabidopsis/efeitos dos fármacos , Arabidopsis/crescimento & desenvolvimento , Proteínas de Arabidopsis/genética , Proteínas de Ligação a DNA/genética , Proteínas Nucleares/genética , Fosforilação , Reguladores de Crescimento de Plantas/farmacologia , Plantas Geneticamente Modificadas/efeitos dos fármacos , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Proteólise , Transdução de SinaisRESUMO
Plants have evolved a sophisticated immunity system for specific detection of pathogens and rapid induction of measured defences. Over- or constitutive activation of defences would negatively affect plant growth and development. Hence, the plant immune system is under tight positive and negative regulation. MAP kinase phosphatase1 (MKP1) has been identified as a negative regulator of plant immunity in model plant Arabidopsis. However, the molecular mechanisms by which MKP1 regulates immune signalling in wheat (Triticum aestivum) are poorly understood. In this study, we investigated the role of TaMKP1 in wheat defence against two devastating fungal pathogens and determined its subcellular localization. We demonstrated that knock-down of TaMKP1 by CRISPR/Cas9 in wheat resulted in enhanced resistance to rust caused by Puccinia striiformis f. sp. tritici (Pst) and powdery mildew caused by Blumeria graminis f. sp. tritici (Bgt), indicating that TaMKP1 negatively regulates disease resistance in wheat. Unexpectedly, while Tamkp1 mutant plants showed increased resistance to the two tested fungal pathogens they also had higher yield compared with wild-type control plants without infection. Our results suggested that TaMKP1 interacts directly with dephosphorylated and activated TaMPK3/4/6, and TaMPK4 interacts directly with TaPAL. Taken together, we demonstrated TaMKP1 exert negative modulating roles in the activation of TaMPK3/4/6, which are required for MAPK-mediated defence signalling. This facilitates our understanding of the important roles of MAP kinase phosphatases and MAPK cascades in plant immunity and production, and provides germplasm resources for breeding for high resistance and high yield.
Assuntos
Sistemas CRISPR-Cas , Resistência à Doença , Doenças das Plantas , Imunidade Vegetal , Triticum , Triticum/genética , Triticum/microbiologia , Triticum/imunologia , Doenças das Plantas/microbiologia , Doenças das Plantas/imunologia , Doenças das Plantas/genética , Imunidade Vegetal/genética , Resistência à Doença/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Ascomicetos/fisiologia , Mutagênese , Fosfatase 1 de Especificidade Dupla/genética , Fosfatase 1 de Especificidade Dupla/metabolismo , Fosfatases da Proteína Quinase Ativada por Mitógeno/genética , Fosfatases da Proteína Quinase Ativada por Mitógeno/metabolismo , Puccinia/fisiologia , Plantas Geneticamente ModificadasRESUMO
The plant hormone ethylene plays a critical role in fruit defense against Botrytis cinerea attack, but the underlying mechanisms remain poorly understood. Here, we showed that ethylene response factor SlERF.C1 acts as a key regulator to trigger the ethylene-mediated defense against B. cinerea in tomato fruits without compromising ripening. Knockout of SlERF.C1 increased fruit susceptibility to B. cinerea with no effect on ripening process, while overexpression enhanced resistance. RNA-Seq, transactivation assays, EMSA and ChIP-qPCR results indicated that SlERF.C1 activated the transcription of PR genes by binding to their promoters. Moreover, SlERF.C1 interacted with the mitogen-activated protein kinase SlMPK8 which allowed SlMPK8 to phosphorylate SlERF.C1 at the Ser174 residue and increases its transcriptional activity. Knocking out of SlMPK8 increased fruit susceptibility to B. cinerea, whereas overexpression enhanced resistance without affecting ripening. Furthermore, genetic crosses between SlMPK8-KO and SlERF.C1-OE lines reduced the resistance to B. cinerea attack in SlERF.C1-OE fruits. In addition, B. cinerea infection induced ethylene production which in turn triggered SlMPK8 transcription and enhanced the phosphorylation of SlERF.C1. Overall, our findings reveal the regulatory mechanism of the 'Ethylene-MPK8-ERF.C1-PR' module in resistance against B. cinerea and provide new insight into the manipulation of gray mold disease in fruits.
Assuntos
Frutas , Solanum lycopersicum , Frutas/metabolismo , Solanum lycopersicum/genética , Etilenos/metabolismo , Botrytis/fisiologia , Doenças das Plantas/genética , Resistência à Doença/genética , Regulação da Expressão Gênica de PlantasRESUMO
Heterotrimeric G proteins play key roles in cellular processes. Although phenotypic analyses of Arabidopsis Gß (AGB1) mutants have implicated G proteins in abscisic acid (ABA) signaling, the AGB1-mediated modules involved in ABA responses remain unclear. We found that a partial AGB1 protein was localized to the nucleus where it interacted with ABA-activated VirE2-interacting protein 1 (VIP1) and mitogen-activated protein kinase 3 (MPK3). AGB1 acts as an upstream negative regulator of VIP1 activity by initiating responses to ABA and drought stress, and VIP1 regulates the ABA signaling pathway in an MPK3-dependent manner in Arabidopsis. AGB1 outcompeted VIP1 for interaction with the C-terminus of MPK3, and prevented phosphorylation of VIP1 by MPK3. Importantly, ABA treatment reduced AGB1 expression in the wild type, but increased in vip1 and mpk3 mutants. VIP1 associates with ABA response elements present in the AGB1 promoter, forming a negative feedback regulatory loop. Thus, our study defines a new mechanism for fine-tuning ABA signaling through the interplay between AGB1 and MPK3-VIP1. Furthermore, it suggests a common G protein mechanism to receive and transduce signals from the external environment.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Subunidades beta da Proteína de Ligação ao GTP , Ácido Abscísico/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas , Subunidades beta da Proteína de Ligação ao GTP/genética , Subunidades beta da Proteína de Ligação ao GTP/metabolismo , FosforilaçãoRESUMO
Optimal stomatal regulation is important for plant adaptation to changing environmental conditions and for maintaining crop yield. The guard-cell signal GABA is produced from glutamate by Glutamate Decarboxylase (GAD) during a reaction that generates carbon dioxide (CO2) as a by-product. Here, we investigated a putative connection between GABA signalling and the more clearly defined CO2 signalling pathway in guard cells. The GABA-deficient mutant lines gad2-1, gad2-2 and gad1/2/4/5 were examined for stomatal sensitivity to various CO2 concentrations. Our findings show a phenotypical discrepancy between the allelic mutant lines gad2-1 and gad2-2 - a weakened CO2 response in gad2-1 (GABI_474_E05) in contrast to a wild-type response in gad2-2 (SALK_028819) and gad1/2/4/5. Through transcriptomic and genomic investigation, we traced the response of gad2-1 to a deletion of full-length Mitogen-activated protein kinase 12 (MPK12) in the GABI-KAT line, thereafter as renamed gad2-1*. Guard cell-specific complementation of MPK12 restored the gad2-1* CO2 phenotype, which confirms the proposed importance of MPK12 to CO2 sensitivity. Additionally, we found that stomatal opening under low atmospheric CO2 occurs independently of the GABA-modulated opening-channel ALMT9. Our results confirm that GABA has a role in modulating the rate of stomatal opening and closing - but not in response to CO2 per se.
RESUMO
Stomatal movements allow the uptake of CO2 for photosynthesis and water loss through transpiration, therefore play a crucial role in determining water use efficiency. Both red and blue lights induce stomatal opening, and the stomatal apertures under light are finetuned by both positive and negative regulators in guard cells. However, the molecular mechanisms for precisely adjusting stomatal apertures under light have not been completely understood. Here, we provided evidence supporting that Arabidopsis thaliana mitogen-activated protein kinase 11 (MPK11) plays a negative role in red light-induced stomatal opening. First, MPK11 was found to be highly expressed in guard cells, and MPK11-GFP signals were detected in both nuclear and cytoplasm of guard cells. The transcript levels of MPK11 in guard cells were upregulated by white light, and the stomata of mpk11 opened wider than that of wild type under white light. Consistent with the larger stomatal aperture, mpk11 mutant exhibited higher stomatal conductance and CO2 assimilation rate under white light. The transcript levels of the genes responsible for osmolytes increases were higher in guard cells of mpk11 than that of wild type, which may contribute to the larger stomatal aperture of mpk11 under white light. Furthermore, MPK11 transcript levels in guard cells were upregulated by red light, and mpk11 mutant showed a larger stomatal aperture under red light. Taken together, these results demonstrate that red light-upregulated MPK11 plays a negative role in stomatal opening, which finetuning the stomatal opening apertures and preventing excessive water loss by transpiration under light.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Proteína Quinase 11 Ativada por Mitógeno/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Dióxido de Carbono/metabolismo , Estômatos de Plantas/metabolismo , Luz , Água/metabolismoRESUMO
Light is known to regulate anthocyanin pigment biosynthesis in plants on several levels, but the significance of protein phosphorylation in light-induced anthocyanin accumulation needs further investigation. In this study, we investigated the dynamics of the apple fruit phosphoproteome in response to light, using high-performance liquid chromatography-tandem mass spectrometry analysis. Among the differentially phosphorylated proteins, the bZIP (basic leucine zipper) transcription factor, HY5, which has been identified as an anthocyanin regulator, was rapidly activated by light treatment of the fruit. We hypothesized that phosphorylated MdHY5 may play a role in light-induced anthocyanin accumulation of apple fruit. Protein interaction and phosphorylation assays showed that mitogen-activated protein kinase MdMPK6 directly interacted with, and activated, MdHY5 via phosphorylation under light conditions, thereby increasing its stability. Consistent with this finding, the suppression of the mitogen-activated protein kinase genes MdMPK6 or MdHY5 resulted in an inhibition of anthocyanin accumulation, and further showed that light-induced anthocyanin accumulation is dependent on MdMPK6 kinase activity, and is required for maximum MdHY5 activity. Under light conditions, active MdMPK6 phosphorylated MdHY5 leading to accumulation of phospho-MdHY5, which enhanced the binding of MdHY5 to its target anthocyanin related genes in fruit. Our findings reveal an MdMPK6-MdHY5 phosphorylation pathway in light-induced anthocyanin accumulation, providing new insights into the regulation of light-induced anthocyanin biosynthesis in apple fruit at both the transcriptional and post-translational levels.
Assuntos
Malus , Malus/genética , Malus/metabolismo , Frutas/genética , Frutas/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Antocianinas , Fosforilação , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/genética , Proteínas Quinases Ativadas por Mitógeno/genética , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Regulação da Expressão Gênica de Plantas/genéticaRESUMO
MAPKs are universal eukaryotic signaling factors whose functioning is assumed to depend on the recognition of a common docking motif (CD) by its activators, substrates, and inactivators. We studied the role of the CD domain of Arabidopsis MPK4 by performing interaction studies and determining the ligand-bound MPK4 crystal structure. We revealed that the CD domain of MPK4 is essential for interaction and activation by its upstream MAPKKs MKK1, MKK2, and MKK6. Cys181 in the CD site of MPK4 was shown to become sulfenylated in response to reactive oxygen species in vitro. To test the function of C181 in vivo, we generated wild-type (WT) MPK4-C181, nonsulfenylatable MPK4-C181S, and potentially sulfenylation mimicking MPK4-C181D lines in the mpk4 knockout background. We analyzed the phenotypes in growth, development, and stress responses, revealing that MPK4-C181S has WT activity and complements the mpk4 phenotype. By contrast, MPK4-C181D cannot be activated by upstream MAPKK and cannot complement the phenotypes of mpk4. Our findings show that the CD motif is essential and is required for activation by upstream MAPKK for MPK4 function. Furthermore, growth, development, or immunity functions require upstream activation of the MPK4 protein kinase.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Proteínas de Arabidopsis/metabolismo , Quinases de Proteína Quinase Ativadas por Mitógeno/genética , Sistema de Sinalização das MAP Quinases , Arabidopsis/metabolismo , Imunidade Vegetal/genéticaRESUMO
ROOT MERISTEM GROWTH FACTOR1 (RGF1) and its receptors RGF1 INSENSITIVEs (RGIs) regulate primary root meristem activity via a mitogen-activated protein kinase (MPK) signaling cascade in Arabidopsis. However, it is unknown how RGF1 regulates lateral root (LR) development. Here, we show that the RGF1-RGI1 peptide-receptor pair negatively regulates LR development via activation of PUCHI encoding AP2/EREBP. Exogenous RGF1 peptides inhibited LR development of the wild type. However, the rgi1 mutants were partially or fully insensitive to RGF1 during LR development, whereas four other rgi single mutants, namely rgi2, rgi3, rgi4, and rgi5, were sensitive to RGF1 in inhibiting LR formation. Consistent with this, the red fluorescent protein (RFP) signals driven by the RGF1 promoter were detected at stage I and the following stages, overlapping with RGI1 expression. PUCHI expression was significantly up-regulated by RGF1 but completely inhibited in rgi1. LR development of puchi1-1 was insensitive to RGF1. PUCHI expression driven by the RGI1 promoter reduced LR density in both the wild type and rgi1,2,3. Further, mpk6, but not mpk3, displayed significantly down-regulated PUCHI expression and insensitive LR development in response to RGF1. Collectively, these results suggest that the RGF1-RGI1 module negatively regulates LR development by activating PUCHI expression via MPK6.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas , Meristema , Proteínas Quinases Ativadas por Mitógeno/genética , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Peptídeos/metabolismo , Raízes de Plantas/metabolismo , Receptores de Peptídeos/metabolismo , Fatores de Transcrição/metabolismoRESUMO
Cytokinins are phytohormones that regulate plant development, growth, and responses to stress. In particular, cytokinin has been reported to negatively regulate plant adaptation to high salinity; however, the molecular mechanisms that counteract cytokinin signaling and enable salt tolerance are not fully understood. Here, we provide evidence that salt stress induces the degradation of the cytokinin signaling components Arabidopsis (Arabidopisis thaliana) response regulator 1 (ARR1), ARR10 and ARR12. Furthermore, the stress-activated mitogen-activated protein kinase 3 (MPK3) and MPK6 interact with and phosphorylate ARR1/10/12 to promote their degradation in response to salt stress. As expected, salt tolerance is decreased in the mpk3/6 double mutant, but enhanced upon ectopic MPK3/MPK6 activation in an MKK5DD line. Importantly, salt hypersensitivity phenotypes of the mpk3/6 line were significantly alleviated by mutation of ARR1/12. The above results indicate that MPK3/6 enhance salt tolerance in part via their negative regulation of ARR1/10/12 protein stability. Thus, our work reveals a new molecular mechanism underlying salt-induced stress adaptation and the inhibition of plant growth, via enhanced degradation of cytokinin signaling components.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas , Proteína Quinase 3 Ativada por Mitógeno , Tolerância ao Sal/genéticaRESUMO
Protein kinases of the MAPK cascade family (MAPKKK-MAPKK-MAPK) play an essential role in plant stress response and hormone signal transduction. However, their role in the cold hardiness of Prunus mume (Mei), a class of ornamental woody plant, remains unclear. In this study, we use bioinformatic approaches to assess and analyze two related protein kinase families, namely, MAP kinases (MPKs) and MAPK kinases (MKKs), in wild P. mume and its variety P. mume var. tortuosa. We identify 11 PmMPK and 7 PmMKK genes in the former species and 12 PmvMPK and 7 PmvMKK genes in the latter species, and we investigate whether and how these gene families contribute to cold stress responses. Members of the MPK and MKK gene families located on seven and four chromosomes of both species are free of tandem duplication. Four, three, and one segment duplication events are exhibited in PmMPK, PmvMPK, and PmMKK, respectively, suggesting that segment duplications play an essential role in the expansion and evolution of P. mume and its gene variety. Moreover, synteny analysis suggests that most MPK and MKK genes have similar origins and involved similar evolutionary processes in P. mume and its variety. A cis-acting regulatory element analysis shows that MPK and MKK genes may function in P. mume and its variety's development, modulating processes such as light response, anaerobic induction, and abscisic acid response as well as responses to a variety of stresses, such as low temperature and drought. Most PmMPKs and PmMKKs exhibited tissue-specifific expression patterns, as well as time-specific expression patterns that protect them through cold. In a low-temperature treatment experiment with the cold-tolerant cultivar P. mume 'Songchun' and the cold-sensitive cultivar 'Lve', we find that almost all PmMPK and PmMKK genes, especially PmMPK3/5/6/20 and PmMKK2/3/6, dramatically respond to cold stress as treatment duration increases. This study introduces the possibility that these family members contribute to P. mume's cold stress response. Further investigation is warranted to understand the mechanistic functions of MAPK and MAPKK proteins in P. mume development and response to cold stress.
Assuntos
Quinases de Proteína Quinase Ativadas por Mitógeno , Prunus , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Resposta ao Choque Frio/genética , Prunus/genética , Prunus/metabolismo , Genoma de Planta , Sequência de Aminoácidos , Alinhamento de Sequência , Plantas/metabolismo , Filogenia , Regulação da Expressão Gênica de PlantasRESUMO
The chemical diversity of sphingolipids in plants allows the assignment of specific roles to special molecular species. These roles include NaCl receptors for glycosylinositolphosphoceramides or second messengers for long-chain bases (LCBs), free or in their acylated forms. Such signaling function has been associated with plant immunity, with an apparent connection to mitogen-activated protein kinase 6 (MPK6) and reactive oxygen species (ROS). This work used in planta assays with mutants and fumonisin B1 (FB1) to generate varying levels of endogenous sphingolipids. This was complemented with in planta pathogenicity tests using virulent and avirulent Pseudomonas syringae strains. Our results indicate that the surge of specific free LCBs and ceramides induced by FB1 or an avirulent strain trigger a biphasic ROS production. The first transient phase is partially produced by NADPH oxidase, and the second is sustained and is related to programmed cell death. MPK6 acts downstream of LCB buildup and upstream of late ROS and is required to selectively inhibit the growth of the avirulent but not the virulent strain. Altogether, these results provide evidence that a LCB- MPK6- ROS signaling pathway contributes differentially to the two forms of immunity described in plants, upregulating the defense scheme of a non-compatible interaction.