RESUMO
Heat stress limits plant growth, development, and crop yield, but how plant cells precisely sense and transduce heat stress signals remains elusive. Here, we identified a conserved heat stress response mechanism to elucidate how heat stress signal is transmitted from the cytoplasm into the nucleus for epigenetic modifiers. We demonstrate that HISTONE DEACETYLASE 9 (HDA9) transduces heat signals from the cytoplasm to the nucleus to play a positive regulatory role in heat responses in Arabidopsis. Heat specifically induces HDA9 accumulation in the nucleus. Under heat stress, the phosphatase PP2AB'ß directly interacts with and dephosphorylates HDA9 to protect HDA9 from 26S proteasome-mediated degradation, leading to the translocation of nonphosphorylated HDA9 to the nucleus. This heat-induced enrichment of HDA9 in the nucleus depends on the nucleoporin HOS1. In the nucleus, HDA9 binds and deacetylates the target genes related to signaling transduction and plant development to repress gene expression in a transcription factor YIN YANG 1-dependent and -independent manner, resulting in rebalance of plant development and heat response. Therefore, we uncover an HDA9-mediated positive regulatory module in the heat shock signal transduction pathway. More important, this cytoplasm-to-nucleus translocation of HDA9 in response to heat stress is conserved in wheat and rice, which confers the mechanism significant implication potential for crop breeding to cope with global climate warming.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas , Células Vegetais/metabolismo , Melhoramento Vegetal , Arabidopsis/metabolismo , Histona Desacetilases/genética , Histona Desacetilases/metabolismoRESUMO
Although much is known about plant responses to heat shock (HS), how plants sense high temperature and the primary HS signal transduction pathway leading to HS-regulated gene expression are still poorly understood. To identify primary transcription factors that mediate HS-regulated gene expression and their target genes, RNA sequencing was performed to detect genes whose expression is rapidly altered by HS in Arabidopsis (Arabidopsis thaliana). The results showed several genes were induced after only 5 min of HS treatment, suggesting that HS signaling occurs very rapidly. Analysis of the cis-elements in the promoters of genes upregulated by 10 min of HS treatment identified HEAT SHOCK FACTOR A1s (HSFA1s) and circadian clock proteins REVEILLE4 (RVE4) and RVE8 as essential transcription factors that independently mediate early HS-induced gene expression. Using hsfa1a/b/d/e and rve4/8 mutants, we identified subsets of HSFA1s- or RVE4/8-dependent early HS-induced genes and showed RVE4/8 regulate plant thermotolerance partially by regulating the expression of downstream transcription factors ETHYLENE RESPONSIVE FACTOR53 (ERF53) and ERF54, specifically around noon. These findings reveal a potential transcriptional regulatory hierarchy governing the first wave of HS-induced gene expression. They also provided important insight into the mechanism by which the circadian clock gates thermotolerance and prepares plants for exposure to high temperatures during the day.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Relógios Circadianos/genética , Proteínas de Ligação a DNA/metabolismo , Fatores de Transcrição de Choque Térmico/metabolismo , Resposta ao Choque Térmico/genética , Fatores de Transcrição/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Relógios Circadianos/fisiologia , Proteínas de Ligação a DNA/genética , Regulação da Expressão Gênica de Plantas/genética , Ontologia Genética , Fatores de Transcrição de Choque Térmico/genética , Resposta ao Choque Térmico/fisiologia , Temperatura Alta , Regiões Promotoras Genéticas , Ligação Proteica/genética , RNA-Seq , Transdução de Sinais/genética , Estresse Fisiológico/genética , Termotolerância/genética , Termotolerância/fisiologia , Fatores de Transcrição/genética , Ativação Transcricional/genética , Ativação Transcricional/fisiologiaRESUMO
The receptor-like kinase SIT1 acts as a sensor in rice (Oryza sativa) roots, relaying salt stress signals via elevated kinase activity to enhance salt sensitivity. Here, we demonstrate that Protein Phosphatase 2A (PP2A) regulatory subunit B'κ constrains SIT1 activity under salt stress. B'κ-PP2A deactivates SIT1 directly by dephosphorylating the kinase at Thr515/516, a salt-induced phosphorylation site in the activation loop that is essential for SIT1 activity. B'κ overexpression suppresses the salt sensitivity of rice plants expressing high levels of SIT1, thereby contributing to salt tolerance. B'κ functions in a SIT1 kinase-dependent manner. During early salt stress, activated SIT1 phosphorylates B'κ; this not only enhances its binding with SIT1, it also promotes B'κ protein accumulation via Ser502 phosphorylation. Consequently, by blocking SIT1 phosphorylation, B'κ inhibits and fine-tunes SIT1 activity to balance plant growth and stress adaptation.
Assuntos
Oryza/metabolismo , Proteínas de Plantas/metabolismo , Proteínas Quinases/metabolismo , Proteína Fosfatase 2/metabolismo , Estresse Salino/fisiologia , Adaptação Fisiológica , Regulação da Expressão Gênica de Plantas , Oryza/genética , Oryza/crescimento & desenvolvimento , Fosforilação , Proteínas de Plantas/genética , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas , Estresse Salino/genética , Tolerância ao Sal/genética , Tolerância ao Sal/fisiologia , Estresse FisiológicoRESUMO
ROOT MERISTEM GROWTH FACTOR (RGF) 1 is an important peptide hormone that regulates root growth. Upon binding to its receptor, RGFR1, RGF1 regulates the expression of two transcription factors, PLETHORA 1 and 2 (PLT1/2), to influence root meristem development. Here, we show that the ubiquitin-specific proteases UBP12 and UBP13 are positive regulators of root meristem development and that UBP13 interacts directly with RGF1 receptor (RGFR1) and its close homolog RGFR2. The ubp12,13 double-mutant root is completely insensitive to exogenous applied RGF1. Consistent with this result, RGF1-induced ubiquitination and turnover of RGFR1 protein were accelerated in ubp12,13-mutant plants but were delayed in transgenic plants overexpressing UBP13 Genetic analysis showed that PLT2 or RGFR1 overexpression partially rescued the short-root phenotype and the reduced cortical root meristem cell number in ubp12,13 plants. Together, our results demonstrate that UBP12/13 are regulators of the RGF1-RGFR1-PLT1/2 signaling pathway and that UBP12/13 can counteract RGF1-induced RGFR1 ubiquitination, stabilize RGFR1, and maintain root cell sensitivity to RGF1.
Assuntos
Proteínas de Arabidopsis/metabolismo , Endopeptidases/metabolismo , Meristema/fisiologia , Peptídeos/metabolismo , Raízes de Plantas/fisiologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Arabidopsis/fisiologia , Membrana Celular/metabolismo , Regulação da Expressão Gênica de Plantas , Ligantes , Hormônios Peptídicos/metabolismo , Fenótipo , Fosforilação , Plantas Geneticamente Modificadas/fisiologia , Mapeamento de Interação de Proteínas , Transdução de Sinais , Proteases Específicas de Ubiquitina/metabolismo , UbiquitinaçãoRESUMO
BACKGROUND: As global warming becomes increasingly severe, it is urgent that we enhance the heat tolerance of crops. We previously reported that Arabidopsis thaliana PHOSPHOINOSITIDE-SPECIFIC PHOSPHOLIPASE C9 (AtPLC9) promotes heat tolerance. RESULTS: In this study, we ectopically expressed AtPLC9 in rice to examine its potential to improve heat tolerance in this important crop. Whereas AtPLC9 did not improve rice tolerance to salt, drought or cold, transgenic rice did exhibit greater heat tolerance than the wild type. High-throughput RNA-seq revealed extensive and dynamic transcriptome reprofiling in transgenic plants after heat stress. Moreover, the expression of some transcription factors and calcium ion-related genes showed specific upregulation in transgenic rice after heat stress, which might contribute to the enhanced heat tolerance. CONCLUSIONS: This study provides preliminary guidance for using AtPLC9 to improve heat tolerance in cereal crops and, more broadly, highlights that heterologous transformation can assist with molecular breeding.
Assuntos
Grão Comestível/genética , Resposta ao Choque Térmico/genética , Resposta ao Choque Térmico/fisiologia , Oryza/genética , Estresse Fisiológico/genética , Termotolerância/genética , Termotolerância/fisiologia , Arabidopsis , Grão Comestível/fisiologia , Regulação da Expressão Gênica de Plantas , Técnicas de Transferência de Genes , Genes de Plantas , Oryza/fisiologia , Proteínas de Plantas/genética , Plantas Geneticamente ModificadasRESUMO
Cell-to-cell communication precisely controls the creation of new organs during reproductive growth. However, the sensor molecules that mediate developmental signals in monocot plants are poorly understood. Here, we report that DWARF AND RUNTISH SPIKELET1 (DRUS1) and DRUS2, two closely related receptor-like kinases (RLKs), redundantly control reproductive growth and development in rice (Oryza sativa). A drus1-1 drus2 double knockout mutant, but not either single mutant, showed extreme dwarfism and barren inflorescences that harbored sterile spikelets. The gibberellin pathway was not impaired in this mutant. A phenotypic comparison of mutants expressing different amounts of DRUS1 and 2 revealed that reproductive growth requires a threshold level of DRUS1/2 proteins. DRUS1 and 2 maintain cell viability by repressing protease-mediated cell degradation and likely by affecting sugar utilization or conversion. In the later stages of anther development, survival of the endothecium requires DRUS1/2, which may stimulate expression of the UDP-glucose pyrophosphorylase gene UGP2 and starch biosynthesis in pollen. Unlike their Arabidopsis thaliana ortholog FERONIA, DRUS1 and 2 mediate a fundamental signaling process that is essential for cell survival and represents a novel biological function for the CrRLK1L RLK subfamily.
Assuntos
Metabolismo dos Carboidratos/genética , Oryza/genética , Proteínas de Plantas/genética , Receptores Proteína Tirosina Quinases/genética , Sequência de Aminoácidos , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Morte Celular/genética , Flores/enzimologia , Flores/genética , Flores/ultraestrutura , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica de Plantas , Immunoblotting , Hibridização In Situ , Microscopia Confocal , Microscopia Eletrônica , Oryza/enzimologia , Fosfotransferases/genética , Fosfotransferases/metabolismo , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Interferência de RNA , Receptores Proteína Tirosina Quinases/metabolismo , Reprodução/genética , Homologia de Sequência de Aminoácidos , Amido/metabolismoRESUMO
To understand the early heat shock (HS)-regulated cellular responses that influence the tolerance of rice plant to high environmental temperatures, two-dimensional difference gel electrophoresis (2D-DIGE) is performed to explore the early HS-regulated proteome. Multiple proteins that show abundance changes after 1 and 5 min of HS treatment are identified. Of the early HS-regulated proteins identified, the abundance of a ubiquitin-specific protease, OsUBP21, and its Arabidopsis homolog, AtUBP13, is found to be upregulated by 5 min of HS treatment. Further, knocking the expression of OsUBP21 or AtUBP13 down or out increases the tolerance of rice and Arabidopsis plants to HS stress, suggesting that the function of these ubiquitin-specific proteases in regulating plant HS responses is conserved between monocots and dicots. 2D-DIGE showed a group of proteins are differentially regulated in wild-type and ubp21 mutant after 30 min of HS treatment. Among these proteins, 11 are found to interact directly with OsUBP21; thus, they may be targets of OsUBP21. Future analyses of the roles of these OsUBP21-interacting proteins in plant HS responses will help reveal the protein ubiquitination/deubiquitination-regulated cellular responses induced by HS in rice.
Assuntos
Resposta ao Choque Térmico , Oryza/metabolismo , Proteínas de Plantas/metabolismo , Proteases Específicas de Ubiquitina/metabolismo , Regulação da Expressão Gênica de Plantas , Oryza/química , Oryza/genética , Proteínas de Plantas/análise , Proteínas de Plantas/genética , Mapeamento de Interação de Proteínas/métodos , Proteômica/métodos , Eletroforese em Gel Diferencial Bidimensional/métodos , Proteases Específicas de Ubiquitina/análise , Proteases Específicas de Ubiquitina/genéticaRESUMO
In this work, a room-temperature atmospheric pressure direct-current plasma has been deployed for the one-step synthesis of gold nanoparticle/carboxyl group-functionalized carbon nanotube (AuNP/CNT-COOH) nanohybrids in aqueous solution for the first time. Uniformly distributed AuNPs are formed on the surface of CNT-COOH, without the use of reducing agents or surfactants. The size of the AuNP can be tuned by changing the gold salt precursor concentration. UV-vis, ζ-potential, and X-ray photoelectron spectroscopy suggest that carboxyl surface functional groups on CNTs served as nucleation and growth sites for AuNPs and the multiple potential reaction pathways induced by the plasma chemistry have been elucidated in detail. The nanohybrids exhibit significantly enhanced Raman scattering and photothermal conversion efficiency that are essential for potential multimodal cancer treatment applications.
RESUMO
This is the first study on the deployment of direct current atmospheric pressure microplasma technique for the single step synthesis of gold nanoparticle/graphene oxide (AuNP/GO) nanocomposites. The nanocomposites were characterized using ultraviolet-visible spectroscopy (UV-vis), x-ray diffraction and x-ray photoelectron spectroscopy and their formation mechanisms have been discussed in detail. Our AuNP/GO nanocomposites are highly biocompatible and have demonstrated surface enhanced Raman scattering (SERS) properties as compared to pure AuNPs and pure GO. Their potential as SERS substrate has been further demonstrated using probe molecules (methylene blue) at different concentrations.
RESUMO
To understand the early signaling steps that regulate cold responses in rice, two-dimensional difference gel electrophoresis (2-D DIGE)(1)was used to study early cold-regulated proteins in rice seedlings. Using mass spectrometry, 32 spots, which represent 26 unique proteins that showed an altered expression level within 5 min of cold treatment were identified. Among these proteins, Western blot analyses confirmed that the cellular phospholipase D α1 (OsPLDα1) protein level was increased as early as 1 min after cold treatment. Genetic studies showed that reducing the expression ofOsPLDα1makes rice plants more sensitive to chilling stress as well as cold acclimation increased freezing tolerance. Correspondingly, cold-regulated proteomic changes and the expression of the cold-responsive C repeat/dehydration-responsive element binding 1 (OsDREB1) family of transcription factors were inhibited in thepldα1mutant. We also found that the expression ofOsPLDα1is directly regulated by OsDREB1A. This transcriptional regulation ofOsPLDα1could provide positive feedback regulation of the cold signal transduction pathway in rice. OsPLDα1 hydrolyzes phosphatidylcholine to produce the signal molecule phosphatidic acid (PA). By lipid-overlay assay, we demonstrated that the rice cold signaling proteins, MAP kinase 6 (OsMPK6) and OsSIZ1, bind directly to PA. Taken together, our results suggest that OsPLDα1 plays a key role in transducing cold signaling in rice by producing PA and regulatingOsDREB1s' expression by OsMPK6, OsSIZ1, and possibly other PA-binding proteins.
Assuntos
Aclimatação , Oryza/crescimento & desenvolvimento , Fosfolipase D/metabolismo , Eletroforese em Gel Diferencial Bidimensional/métodos , Temperatura Baixa , Regulação da Expressão Gênica de Plantas , Oryza/genética , Oryza/metabolismo , Ácidos Fosfatídicos/metabolismo , Fosfolipase D/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Proteômica/métodos , Transdução de SinaisRESUMO
High salinity causes growth inhibition and shoot bleaching in plants that do not tolerate high salt (glycophytes), including most crops. The molecules affected directly by salt and linking the extracellular stimulus to intracellular responses remain largely unknown. Here, we demonstrate that rice (Oryza sativa) Salt Intolerance 1 (SIT1), a lectin receptor-like kinase expressed mainly in root epidermal cells, mediates salt sensitivity. NaCl rapidly activates SIT1, and in the presence of salt, as SIT1 kinase activity increased, plant survival decreased. Rice MPK3 and MPK6 function as the downstream effectors of SIT1. SIT1 phosphorylates MPK3 and 6, and their activation by salt requires SIT1. SIT1 mediates ethylene production and salt-induced ethylene signaling. SIT1 promotes accumulation of reactive oxygen species (ROS), leading to growth inhibition and plant death under salt stress, which occurred in an MPK3/6- and ethylene signaling-dependent manner in Arabidopsis thaliana. Our findings demonstrate the existence of a SIT1-MPK3/6 cascade that mediates salt sensitivity by affecting ROS and ethylene homeostasis and signaling. These results provide important information for engineering salt-tolerant crops.
RESUMO
To understand the early signaling steps in the response of plant cells to increased environmental temperature, 2-D difference gel electrophoresis was used to study the proteins in microsomes of Arabidopsis seedlings that are regulated early during heat stress. Using mass spectrometry, 19 microsomal proteins that showed an altered expression level within 5 min after heat treatment were identified. Among these proteins, annexin 1 (AtANN1) was one of those up-regulated rapidly after heat-shock treatment. Functional studies show loss-of-function mutants for AtANN1 and its close homolog AtANN2 were more sensitive to heat-shock treatment, whereas plants overexpressing AtANN1 showed more resistance to this treatment. Correspondingly, the heat-induced expression of heat-shock proteins and heat-shock factors is inhibited in ann1/ann2 double mutant, and the heat-activated increase in cytoplasmic calcium concentration ([Ca(2+)]cyt) is greatly impaired in the ann1 mutant and almost undetectable in ann1/ann2 double mutant. Taken together these results suggest that AtANN1 is important in regulating the heat-induced increase in [Ca(2+)]cyt and in the response of Arabidopsis seedlings to heat stress.
Assuntos
Anexinas/genética , Anexinas/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Cálcio/metabolismo , Proteômica/métodos , Plântula/metabolismo , Anexina A2/genética , Anexina A2/metabolismo , Citoplasma/metabolismo , Regulação da Expressão Gênica de Plantas , Resposta ao Choque Térmico , Temperatura Alta , Microssomos/metabolismo , Mutação , Regulação para CimaRESUMO
Cytokinin plays an important role in plant development and stress tolerance. Studies of Arabidopsis (Arabidopsis thaliana) have demonstrated that cytokinin acts through a two-component system that includes a histidine (His) kinase, a His phosphotransfer protein (HP), and a response regulator. Phylogenetic analyses have revealed the conservation of His kinases but lineage-specific expansion of HPs and response regulators in rice (Oryza sativa). However, whether the functions of rice HPs have diverged remains unknown. In this study, two rice authentic HPs (OsAHP1 and OsAHP2) were knocked down simultaneously via RNA interference (RNAi), and the transgenic OsAHP-RNAi plants exhibited phenotypes expected for a deficiency in cytokinin signaling, including dwarfism with reduced internode lengths, enhanced lateral root growth, early leaf senescence, and reduced tiller numbers and fertility under natural conditions. The OsAHP-RNAi seedlings were also hyposensitive to exogenous cytokinin. Furthermore, OsAHP-RNAi seedlings were hypersensitive to salt treatment but resistant to osmotic stress relative to wild-type plants. These results indicate that OsAHPs function as positive regulators of the cytokinin signaling pathway and play different roles in salt and drought tolerance in rice.
Assuntos
Citocininas/metabolismo , Oryza/metabolismo , Oryza/fisiologia , Transdução de Sinais , Estresse Fisiológico , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Citocininas/farmacologia , Citosol/efeitos dos fármacos , Citosol/metabolismo , Histidina , Oryza/genética , Oryza/crescimento & desenvolvimento , Pressão Osmótica/efeitos dos fármacos , Fenótipo , Fosforilação/efeitos dos fármacos , Desenvolvimento Vegetal/efeitos dos fármacos , Proteínas de Plantas , Raízes de Plantas/anatomia & histologia , Raízes de Plantas/efeitos dos fármacos , Plantas Geneticamente Modificadas , Interferência de RNA/efeitos dos fármacos , Plântula/efeitos dos fármacos , Plântula/fisiologia , Transdução de Sinais/efeitos dos fármacos , Cloreto de Sódio/farmacologia , Estresse Fisiológico/efeitos dos fármacosRESUMO
The heat stress response is an important adaptation, enabling plants to survive challenging environmental conditions. Our previous work demonstrated that Arabidopsis thaliana Phosphoinositide-Specific Phospholipase C Isoform 9 (AtPLC9) plays an important role in thermotolerance. During prolonged heat treatment, mutants of AtPLC3 showed decreased heat resistance. We observed no obvious phenotypic differences between plc3 mutants and wild type (WT) seedlings under normal growth conditions, but after heat shock, the plc3 seedlings displayed a decline in thermotolerance compared with WT, and also showed a 40-50% decrease in survival rate and chlorophyll contents. Expression of AtPLC3 in plc3 mutants rescued the heat-sensitive phenotype; the AtPLC3-overexpressing lines also exhibited much higher heat resistance than WT and vector-only controls. The double mutants of plc3 and plc9 displayed increased sensitivity to heat stress, compared with either single mutant. In transgenic lines containing a AtPLC3:GUS promoter fusion, GUS staining showed that AtPLC3 expresses in all tissues, except anthers and young root tips. Using the Ca(2+)-sensitive fluorescent probe Fluo-3/AM and aequorin reconstitution, we showed that plc3 mutants show a reduction in the heat-induced Ca(2+) increase. The expression of HSP genes (HSP18.2, HSP25.3, HSP70-1 and HSP83) was down-regulated in plc3 mutants and up-regulated in AtPLC3-overexpressing lines after heat shock. These results indicated that AtPLC3 also plays a role in thermotolerance in Arabidopsis, and that AtPLC3 and AtPLC9 function additionally to each other.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Fosfoinositídeo Fosfolipase C/metabolismo , Proteínas de Arabidopsis/genética , Sinalização do Cálcio , Clorofila/metabolismo , Regulação da Expressão Gênica de Plantas , Teste de Complementação Genética , Proteínas de Choque Térmico/genética , Resposta ao Choque Térmico , Isoenzimas/genética , Isoenzimas/metabolismo , Mutação , Fosfoinositídeo Fosfolipase C/genética , Plantas Geneticamente Modificadas , Plântula/genética , Plântula/fisiologiaRESUMO
Calcium ions (Ca(2+)) play a crucial role in many key physiological processes; thus, the maintenance of Ca(2+) homeostasis is of primary importance. Na(+)/Ca(2+) exchangers (NCXs) play an important role in Ca(2+) homeostasis in animal excitable cells. Bioinformatic analysis of the Arabidopsis genome suggested the existence of a putative NCX gene, Arabidopsis NCX-like (AtNCL), encoding a protein with an NCX-like structure and different from Ca(2+)/H(+) exchangers and Na(+)/H(+) exchangers previously identified in plant. AtNCL was identified to localize in the Arabidopsis cell membrane fraction, have the ability of binding Ca(2+), and possess NCX-like activity in a heterologous expression system of cultured mammalian CHO-K1 cells. AtNCL is broadly expressed in Arabidopsis, and abiotic stresses stimulated its transcript expression. Loss-of-function atncl mutants were less sensitive to salt stress than wild-type or AtNCL transgenic overexpression lines. In addition, the total calcium content in whole atncl mutant seedlings was higher than that in wild type by atomic absorption spectroscopy. The level of free Ca(2+) in the cytosol and Ca(2+) flux at the root tips of atncl mutant plants, as detected using transgenic aequorin and a scanning ion-selective electrode, required a longer recovery time following NaCl stress compared with that in wild type. All of these data suggest that AtNCL encodes a Na(+)/Ca(2+) exchanger-like protein that participates in the maintenance of Ca(2+) homeostasis in Arabidopsis. AtNCL may represent a new type of Ca(2+) transporter in higher plants.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Trocador de Sódio e Cálcio/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Cálcio/metabolismo , Regulação da Expressão Gênica de Plantas , Trocador de Sódio e Cálcio/genética , Estresse FisiológicoRESUMO
Intracellular calcium (Ca(2+)) increases rapidly after heat shock (HS) in the Ca(2+)/calmodulin (Ca(2+)/CaM) HS signal transduction pathway: a hypothesis proposed based on our previous findings. However, evidence for the increase in Ca(2+) after HS was obtained only through physiological and pharmacological experiments; thus, direct molecular genetic evidence is needed. The role of phosphoinositide-specific phospholipase C (PI-PLC) is poorly understood in the plant response to HS. In this work, atplc9 mutant plants displayed a serious thermosensitive phenotype compared with wild-type (WT) plants after HS. Complementation of atplc9 with AtPLC9 rescued both the basal and acquired thermotolerance phenotype of the WT plants. In addition, thermotolerance was even improved in overexpressed lines. The GUS staining of AtPLC9 promoter:GUS transgenic seedlings showed that AtPLC9 expression was ubiquitous. The fluorescence distribution of the fusion protein AtPLC9 promoter:AtPLC9:GFP revealed that the subcellular localization of AtPLC9 was restricted to the plasma membrane. The results of a PLC activity assay showed a reduction in the accumulation of inositol-1,4,5-trisphosphate (IP(3)) in atplc9 during HS and improved IP(3) generation in the overexpressed lines. Furthermore, the heat-induced increase in intracellular Ca(2+) was decreased in atplc9. Accumulation of the small HS proteins HSP18.2 and HSP25.3 was downregulated in atplc9 and upregulated in the overexpressed lines after HS. Together, these results provide molecular genetic evidence showing that AtPLC9 plays a role in thermotolerance in Arabidopsis.
Assuntos
Aclimatação/fisiologia , Arabidopsis/enzimologia , Cálcio/metabolismo , Resposta ao Choque Térmico/fisiologia , Fosfolipases/metabolismo , Arabidopsis/genética , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Cálcio/análise , Membrana Celular/enzimologia , Expressão Gênica/genética , Regulação da Expressão Gênica de Plantas/genética , Proteínas de Choque Térmico/metabolismo , Temperatura Alta , Inositol 1,4,5-Trifosfato/metabolismo , Mutagênese Insercional , Fenótipo , Fosfatidilinositóis/metabolismo , Fosfolipases/genética , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Regiões Promotoras Genéticas/genética , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Plântula/genética , Plântula/metabolismo , Plântula/fisiologia , Transdução de Sinais , Análise de SobrevidaRESUMO
An increased concentration of cytosolic calcium ions (Ca²âº) is an early response by plant cells to heat shock. However, the molecular mechanism underlying the heat-induced initial Ca²âº response in plants is unclear. In this study, we identified and characterized a heat-activated Ca²âº-permeable channel in the plasma membrane of Arabidopsis thaliana root protoplasts using reverse genetic analysis and the whole-cell patch-clamp technique. The results indicated that A. thaliana cyclic nucleotide-gated ion channel 6 (CNGC6) mediates heat-induced Ca²âº influx and facilitates expression of heat shock protein (HSP) genes and the acquisition of thermotolerance. GUS and GFP reporter assays showed that CNGC6 expression is ubiquitous in A. thaliana, and the protein is localized to the plasma membrane of cells. Furthermore, it was found that the level of cytosolic cAMP was increased by a mild heat shock, that CNGC6 was activated by cytosolic cAMP, and that exogenous cAMP promoted the expression of HSP genes. The results reveal the role of cAMP in transduction of heat shock signals in plants. The correlation of an increased level of cytosolic cAMP in a heat-shocked plant with activation of the Ca²âº channels and downstream expression of HSP genes sheds some light on how plants transduce a heat stimulus into a signal cascade that leads to a heat shock response.
Assuntos
Proteínas de Arabidopsis/fisiologia , Arabidopsis/genética , AMP Cíclico/metabolismo , Canais de Cátion Regulados por Nucleotídeos Cíclicos/fisiologia , Resposta ao Choque Térmico , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Cálcio/metabolismo , Canais de Cátion Regulados por Nucleotídeos Cíclicos/genética , Citosol/metabolismo , Regulação da Expressão Gênica de Plantas , Proteínas de Choque Térmico/fisiologia , Mutagênese Insercional , Técnicas de Patch-Clamp , Transdução de SinaisRESUMO
The palea and lemma are unique organs in grass plants that form a protective barrier around the floral organs and developing kernel. The interlocking of the palea and lemma is critical for maintaining fertility and seed yield in rice; however, the molecules that control the interlocking structure remain largely unknown. Here, we showed that when OsCR4 mRNA expression was knocked down in rice by RNA interference, the palea and lemma separated at later spikelet stages and gradually turned brown after heading, resulting in the severe interruption of pistil pollination and damage to the development of embryo and endosperm, with defects in aleurone. The irregular architecture of the palea and lemma was caused by tumour-like cell growth in the outer epidermis and wart-like cell masses in the inner epidermis. These abnormal cells showed discontinuous cuticles and uneven cell walls, leading to organ self-fusion that distorted the interlocking structures. Additionally, the faster leakage of chlorophyll, reduced silica content and elevated accumulation of anthocyanin in the palea and lemma indicated a lesion in the protective barrier, which also impaired seed quality. OsCR4 is an active receptor-like kinase associated with the membrane fraction. An analysis of promoter::GUS reporter plants showed that OsCR4 is specifically expressed in the epidermal cells of paleas and lemmas. Together, these results suggest that OsCR4 plays an essential role in maintaining the interlocking of the palea and lemma by promoting epidermal cell differentiation.
Assuntos
Diferenciação Celular , Oryza/enzimologia , Epiderme Vegetal/citologia , Proteínas de Plantas/metabolismo , Proteínas Quinases/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Regulação da Expressão Gênica de Plantas , Técnicas de Silenciamento de Genes , Oryza/citologia , Oryza/genética , Epiderme Vegetal/crescimento & desenvolvimento , Infertilidade das Plantas , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/citologia , Plantas Geneticamente Modificadas/enzimologia , Plantas Geneticamente Modificadas/genética , Proteínas Quinases/genética , Interferência de RNARESUMO
The wall-associated kinase (WAK) gene family is a unique subfamily of receptor-like kinases (RLKs) in plants. WAK-RLKs play roles in cell expansion, pathogen resistance, and metal tolerance in Arabidopsis (Arabidopsis thaliana). Rice (Oryza sativa) has far more WAK-RLK genes than Arabidopsis, but the functions of rice WAK-RLKs are poorly understood. In this study, we found that one rice WAK-RLK gene, DEFECT IN EARLY EMBRYO SAC1 (OsDEES1), is involved in the regulation of early embryo sac development. OsDEES1 silencing by RNA interference caused a high rate of female sterility. Crossing experiments showed that female reproductive organs lacking OsDEES1 carried a functional defect. A detailed investigation of the ovaries from OsDEES1 RNA interference plants indicated that the knockdown of OsDEES1 expression did not affect megasporogenesis but that it disturbed female gametophyte formation, resulting in a degenerated embryo sac and defective seed formation. OsDEES1 exhibited a tissue-specific expression pattern in flowers and seedlings. In the ovary, OsDEES1 was expressed in the megagametophyte region and surrounding nucellus cells in the ovule near the micropylar region. OsDEES1 was found to be a membrane-localized protein with a unique sequence compared with other WAK-RLKs. These data indicate that OsDEES1 plays a role in rice sexual reproduction by regulating female gametophyte development. This study offers new insight into the functions of the WAK-RLK family.
Assuntos
Parede Celular/enzimologia , Oryza/enzimologia , Óvulo Vegetal/crescimento & desenvolvimento , Proteínas de Plantas/metabolismo , Proteínas Quinases/metabolismo , Sequência de Aminoácidos , Membrana Celular/genética , Membrana Celular/metabolismo , Sobrevivência Celular , Cruzamentos Genéticos , Regulação Enzimológica da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Técnicas de Silenciamento de Genes , Genes de Plantas , Imuno-Histoquímica , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Dados de Sequência Molecular , Oryza/embriologia , Oryza/genética , Óvulo Vegetal/enzimologia , Infertilidade das Plantas , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/embriologia , Plantas Geneticamente Modificadas/enzimologia , Plantas Geneticamente Modificadas/genética , Proteínas Quinases/genética , Interferência de RNARESUMO
High-temperature stress affects crop yields worldwide. Identifying thermotolerant crop varieties and understanding the basis for this thermotolerance would have important implications for agriculture, especially in the face of climate change. Rice (Oryza sativa) varieties have evolved protective strategies to acclimate to high temperature, with different thermotolerance levels. In this review, we examine the morphological and molecular effects of heat on rice in different growth stages and plant organs, including roots, stems, leaves and flowers. We also explore the molecular and morphological differences among thermotolerant rice lines. In addition, some strategies are proposed to screen new rice varieties for thermotolerance, which will contribute to the improvement of rice for agricultural production in the future.