RESUMEN
Arabinogalactan proteins (AGPs) are a plant-specific family of extracellular proteoglycans characterized by large and complex galactose-rich polysaccharide chains. Functional elucidation of AGPs, however, has been hindered by the high degree of redundancy of AGP genes. To uncover as yet unexplored roles of AGPs in Arabidopsis, a mutant of Hyp O-galactosyltransferase (HPGT), a critical enzyme that catalyzes the common initial step of Hyp-linked arabinogalactan chain biosynthesis, was used. Here we show, using the hpgt1,2,3 triple mutant, that a reduction in functional AGPs leads to a stomatal patterning defect in which two or more stomata are clustered together. This defect is attributed to increased and dysregulated symplastic transport following changes in plasmodesmata structure, such that highly permeable complex branched plasmodesmata with cavities in branching parts increased in the mutant. We also found that the hpgt1,2,3 mutation causes a reduction of cellulose in the cell wall and accumulation of pectin, which controls cell wall porosity. Our results highlight the importance of AGPs in the correct biogenesis of plasmodesmata, possibly acting through the regulation of cell wall properties surrounding the plasmodesmata.
Asunto(s)
Arabidopsis , Plasmodesmos , Plasmodesmos/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Arabidopsis/metabolismo , Mucoproteínas/genética , Pared Celular/metabolismoRESUMEN
Legumes can survive in nitrogen-deficient environments by forming root-nodule symbioses with rhizobial bacteria; however, forming nodules consumes energy, and nodule numbers must thus be strictly controlled. Previous studies identified major negative regulators of nodulation in Lotus japonicus, including the small peptides CLAVATA3/ESR (CLE)-RELATED-ROOT SIGNAL1 (CLE-RS1), CLE-RS2, and CLE-RS3, and their putative major receptor HYPERNODULATION AND ABERRANT ROOT FORMATION1 (HAR1). CLE-RS2 is known to be expressed in rhizobia-inoculated roots, and is predicted to be post-translationally arabinosylated, a modification essential for its activity. Moreover, all three CLE-RSs suppress nodulation in a HAR1-dependent manner. Here, we identified PLENTY as a gene responsible for the previously isolated hypernodulation mutant plenty. PLENTY encoded a hydroxyproline O-arabinosyltransferase orthologous to ROOT DETERMINED NODULATION1 in Medicago truncatula. PLENTY was localized to the Golgi, and an in vitro analysis of the recombinant protein demonstrated its arabinosylation activity, indicating that CLE-RS1/2/3 may be substrates for PLENTY. The constitutive expression experiments showed that CLE-RS3 was the major candidate substrate for PLENTY, suggesting the substrate preference of PLENTY for individual CLE-RS peptides. Furthermore, a genetic analysis of the plenty har1 double mutant indicated the existence of another PLENTY-dependent and HAR1-independent pathway negatively regulating nodulation.
Asunto(s)
Lotus/enzimología , Pentosiltransferasa/metabolismo , Nódulos de las Raíces de las Plantas/microbiología , Aparato de Golgi/enzimología , Lotus/genética , Lotus/microbiología , Mesorhizobium/fisiología , Pentosiltransferasa/genética , Fenotipo , SimbiosisRESUMEN
A peptide hormone, root meristem growth factor (RGF), regulates root meristem development through the PLETHORA (PLT) stem cell transcription factor pathway, but it remains to be uncovered how extracellular RGF signals are transduced to the nucleus. Here we identified, using a combination of a custom-made receptor kinase (RK) expression library and exhaustive photoaffinity labeling, three leucine-rich repeat RKs (LRR-RKs) that directly interact with RGF peptides in Arabidopsis These three LRR-RKs, which we named RGFR1, RGFR2, and RGFR3, are expressed in root tissues including the proximal meristem, the elongation zone, and the differentiation zone. The triple rgfr mutant was insensitive to externally applied RGF peptide and displayed a short root phenotype accompanied by a considerable decrease in meristematic cell number. In addition, PLT1 and PLT2 protein gradients, observed as a gradual gradient decreasing toward the elongation zone from the stem cell area in wild type, steeply declined at the root tip in the triple mutant. Because RGF peptides have been shown to create a diffusion-based concentration gradient extending from the stem cell area, our results strongly suggest that RGFRs mediate the transformation of an RGF peptide gradient into a PLT protein gradient in the proximal meristem, thereby acting as key regulators of root meristem development.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/embriología , Meristema/embriología , Péptidos/metabolismo , Raíces de Plantas/embriología , Factores de Transcripción/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Línea Celular , Regulación de la Expresión Génica de las Plantas , Ácidos Indolacéticos/metabolismo , Meristema/metabolismo , Péptidos/genética , Raíces de Plantas/metabolismo , Transducción de Señal/fisiología , Nicotiana/citología , Nicotiana/metabolismoRESUMEN
Domestication of crops based on artificial selection has contributed numerous beneficial traits for agriculture. Wild characteristics such as red pericarp and seed shattering were lost in both Asian (Oryza sativa) and African (Oryza glaberrima) cultivated rice species as a result of human selection on common genes. Awnedness, in contrast, is a trait that has been lost in both cultivated species due to selection on different sets of genes. In a previous report, we revealed that at least three loci regulate awn development in rice; however, the molecular mechanism underlying awnlessness remains unknown. Here we isolate and characterize a previously unidentified EPIDERMAL PATTERNING FACTOR-LIKE (EPFL) family member named REGULATOR OF AWN ELONGATION 2 (RAE2) and identify one of its requisite processing enzymes, SUBTILISIN-LIKE PROTEASE 1 (SLP1). The RAE2 precursor is specifically cleaved by SLP1 in the rice spikelet, where the mature RAE2 peptide subsequently induces awn elongation. Analysis of RAE2 sequence diversity identified a highly variable GC-rich region harboring multiple independent mutations underlying protein-length variation that disrupt the function of the RAE2 protein and condition the awnless phenotype in Asian rice. Cultivated African rice, on the other hand, retained the functional RAE2 allele despite its awnless phenotype. Our findings illuminate the molecular function of RAE2 in awn development and shed light on the independent domestication histories of Asian and African cultivated rice.
Asunto(s)
Productos Agrícolas/crecimiento & desarrollo , Oryza/crecimiento & desarrollo , Proteínas de Plantas/fisiología , Alelos , Modelos Moleculares , Oryza/genética , Proteínas de Plantas/genéticaRESUMEN
The identification of hormones and their receptors in multicellular organisms is one of the most exciting research areas and has lead to breakthroughs in understanding how their growth and development are regulated. In particular, peptide hormones offer advantages as cell-to-cell signals in that they can be synthesized rapidly and have the greatest diversity in their structure and function. Peptides often undergo post-translational modifications and proteolytic processing to generate small oligopeptide hormones. In plants, such small post-translationally modified peptides constitute the largest group of peptide hormones. We initially explored this type of peptide hormone using bioassay-guided fractionation and later by in silico gene screening coupled with biochemical peptide detection, which led to the identification of four types of novel peptide hormones in plants. We also identified specific receptors for these peptides and transferases required for their post-translational modification. This review summarizes how we discovered these peptide hormone-receptor pairs and post-translational modification enzymes, and how these molecules function in plant growth, development and environmental adaptation.
Asunto(s)
Hormonas Peptídicas/química , Proteínas de Plantas/química , Plantas/química , Receptores de Péptidos/química , Comunicación Celular , Simulación por Computador , Enzimas/química , Desarrollo de la Planta , Proteínas de Plantas/metabolismo , Plantas/metabolismo , Procesamiento Proteico-Postraduccional , Transducción de SeñalRESUMEN
The CLAVATA signaling pathway is a key component of the network that controls stem cell renewal and differentiation in Arabidopsis thaliana. CLAVATA3 (CLV3) is a post-translationally arabinosylated secreted peptide signal that regulates WUSHEL (WUS) transcription to affect the balance of stem cell differentiation and proliferation in the shoot apical meristem (SAM). Known membrane-localized receptors involved in the perception of CLV3 signaling include CLV1, the CLV2/CORYNE (CRN) complex and RPK2. The CLV3 peptide can directly bind to CLV1; however, it is unclear whether the CLV3 peptide directly binds to CLV2 or RPK2. In this study, we re-evaluated the direct interaction between CLV3 and its receptors by photoaffinity labeling with photoactivatable arabinosylated CLV3. We showed that CLV2 and RPK2 exhibited no direct binding to the CLV3 peptide. Further analysis showed that the receptor kinase BAM1 directly binds the CLV3 peptide. A loss-of-function clv1 bam1 double mutant exhibited a large number of stem cells that accumulated in the SAM and was insensitive to exogenous treatment with the arabinosylated CLV3 peptide. WUS gene transcripts were up-regulated, and the region of WUS expression was enlarged at the SAM in the clv1 bam1 double mutant. These results indicate that CLV1 and BAM1 are direct receptors that are sufficient to affect the regulatory network controlling stem cell number in the SAM. In contrast, the CLV2/CRN complex and RPK2 are not involved in direct ligand interactions but may act as co-receptors.
Asunto(s)
Arabidopsis/metabolismo , Meristema/metabolismo , Brotes de la Planta/metabolismo , Transducción de Señal , Proteínas de Arabidopsis/metabolismo , Regulación de la Expresión Génica de las Plantas , Unión ProteicaRESUMEN
Arabinogalactan proteins (AGPs) are plant-specific extracellular glycoproteins implicated in a variety of processes during growth and development. AGP biosynthesis involves O-galactosylation of hydroxyproline (Hyp) residues followed by a stepwise elongation of the complex sugar chains. However, functionally dominant Hyp O-galactosyltransferases, such that their disruption produces phenocopies of AGP-deficient mutants, remain to be identified. Here, we purified and identified three potent Hyp O-galactosyltransferases, HPGT1, HPGT2 and HPGT3, from Arabidopsis microsomal fractions. Loss-of-function analysis indicated that approximately 90% of the endogenous Hyp O-galactosylation activity is attributable to these three enzymes. AGP14 expressed in the triple mutant migrated much faster on SDS-PAGE than when expressed in wild-type, confirming a considerable decrease in levels of glycosylation of AGPs in the mutant. Loss-of-function mutant plants exhibited a pleiotropic phenotype of longer lateral roots, longer root hairs, radial expansion of the cells in the root tip, small leaves, shorter inflorescence stems, reduced fertility and shorter siliques. Our findings provide genetic evidence that Hyp-linked arabinogalactan polysaccharide chains are critical for AGP function and clues to how arabinogalactan moieties of AGPs contribute to cell-to-cell communication during plant growth and development.
Asunto(s)
Arabidopsis/enzimología , Galactosiltransferasas/metabolismo , Hidroxiprolina/metabolismo , Secuencia de Aminoácidos , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Galactanos/metabolismo , Galactosiltransferasas/genética , Genes Reporteros , Glicosilación , Datos de Secuencia Molecular , Mucoproteínas/genética , Mucoproteínas/metabolismo , Mutación , Fenotipo , Filogenia , Hojas de la Planta/enzimología , Hojas de la Planta/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raíces de Plantas/enzimología , Raíces de Plantas/genética , Plantones/enzimología , Plantones/genética , Alineación de SecuenciaRESUMEN
There is a growing awareness that secreted pemediate organ-to-organ communication in higher plants. Xylem sap peptidomics is an effective but challenging approach for identifying long-distance mobile peptides. In this study we developed a simple, gel-free purification system that combines o-chlorophenol extraction with HPLC separation. Using this system, we successfully identified seven oligopeptides from soybean xylem sap exudate that had one or more post-transcriptional modifications: glycosylation, sulfation and/or hydroxylation. RNA sequencing and quantitative PCR analyses showed that the peptide-encoding genes are expressed in multiple tissues. We further analyzed the long-distance translocation of four of the seven peptides using gene-encoding peptides with single amino acid substitutions, and identified these four peptides as potential root-to-shoot mobile oligopeptides. Promoter-GUS analysis showed that all four peptide-encoding genes were expressed in the inner tissues of the root endodermis. Moreover, we found that some of these peptide-encoding genes responded to biotic and/or abiotic factors. These results indicate that our purification system provides a comprehensive approach for effectively identifying endogenous small peptides and reinforce the concept that higher plants employ various peptides in root-to-shoot signaling.
Asunto(s)
Glycine max/metabolismo , Péptidos/genética , Péptidos/metabolismo , Proteínas de Plantas/aislamiento & purificación , Xilema/metabolismo , Secuencia de Aminoácidos , Fraccionamiento Químico/métodos , Clorofenoles/química , Cromatografía Líquida de Alta Presión , Regulación de la Expresión Génica de las Plantas , Datos de Secuencia Molecular , Péptidos/análisis , Proteínas de Plantas/metabolismo , Raíces de Plantas/genética , Raíces de Plantas/metabolismo , Brotes de la Planta/metabolismo , Plantas Modificadas Genéticamente , Reacción en Cadena de la Polimerasa/métodos , Transporte de Proteínas , Procesamiento Postranscripcional del ARN , Análisis de Secuencia de ARNRESUMEN
Small peptides act as local signals during plant development, but few studies have examined their interaction with phytohormone signaling. Here, we show that application of gibberellin (GA) to Arabidopsis shoots induces substantial accumulation of transcripts encoded by CLE6, a member of the CLAVATA/ESR-RELATED (CLE) gene family, in the root stele, followed by promotion of organ growth by CLE6 in GA-deficient plants. The long-distance effect of GA4 was demonstrated by the observation that its application to the shoot apex of the GA-deficient mutant ga3ox1/ga3ox2 rescued the short-root phenotype. Microarray analysis was used to identify root-expressed genes that respond to systemic application of GA, and CLE6 was selected for further analysis. CLE6 was highly expressed in roots at the young seedling stage, and CLE6 promoter activity was strong in hypocotyls and roots, especially in root stele cells at branch points. Application of CLE6 peptide had no obvious effect on the growth and development of GA-deficient mutant plants. Nonetheless, the fact that ectopic over-expression of CLE6 in the GA-deficient mutant promoted root growth and branching, petiole elongation, bolting rate and stem length showed that CLE6 expression partially compensates for the GA deficiency. Reciprocal grafting of GA-deficient mutant plants to 35S::CLE6 transformants complemented the shoot phenotype associated with GA deficiency, demonstrating the systemic effect of CLE6 from root to shoot. These data suggest that root-expressed CLE6 is systemically involved in shoot growth under GA action in Arabidopsis.
Asunto(s)
Proteínas de Arabidopsis/fisiología , Arabidopsis/crecimiento & desarrollo , Giberelinas/metabolismo , Péptidos y Proteínas de Señalización Intercelular/fisiología , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Regulación de la Expresión Génica de las Plantas , Giberelinas/farmacología , Péptidos y Proteínas de Señalización Intercelular/genética , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Reguladores del Crecimiento de las Plantas/metabolismo , Reguladores del Crecimiento de las Plantas/farmacología , Brotes de la Planta/genética , Brotes de la Planta/crecimiento & desarrollo , Brotes de la Planta/metabolismo , ARN Mensajero/metabolismoRESUMEN
To increase our understanding of the adaptation for copper (Cu) deficiency, Arabidopsis mutants with apparent alterations under Cu deficiency were identified. In this report, a novel mutant, tpst-2, was found to be more sensitive than wild-type (Col-0) plants to Cu deficiency during root elongation. The positional cloning of tpst-2 revealed that this gene encodes a tyrosylprotein sulfotransferase (TPST). Moreover, the ethylene production of tpst-2 mutant was higher than that of Col-0 under Cu deficiency, and adding the ethylene response inhibitor AgNO3 partially rescued defects in root elongation. Interestingly, peptide hormone phytosulfokine (PSK) treatment also repressed the ethylene production of tpst-2 mutant plants. Our results revealed that TPST suppressed ethylene production through the action of PSK.
Asunto(s)
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Cobre/farmacología , Etilenos/biosíntesis , Mutación/genética , Fitoquímicos/metabolismo , Arabidopsis/efectos de los fármacos , Proteínas de Arabidopsis/metabolismo , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Genes de Plantas , Homeostasis/efectos de los fármacos , Fenotipo , Raíces de Plantas/efectos de los fármacos , Raíces de Plantas/genética , Empalme del ARN/genética , Receptores de Superficie Celular/metabolismo , Plantones/efectos de los fármacos , Plantones/metabolismo , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Nitrato de Plata/farmacología , Sulfotransferasas/metabolismoRESUMEN
Hydroxyproline (Hyp) O-arabinosylation is a post-translational modification that is prominent in extracellular glycoproteins in plants. Hyp O-arabinosylation is generally found in these glycoproteins in the form of linear oligoarabinoside chains and has a key role in their function by contributing to conformational stability. However, Hyp O-arabinosyltransferase (HPAT), a key enzyme that catalyzes the transfer of the L-arabinose to the hydroxyl group of Hyp residues, has remained undiscovered. Here, we purified and identified Arabidopsis HPAT as a Golgi-localized transmembrane protein that is structurally similar to the glycosyltransferase GT8 family. Loss-of-function mutations in HPAT-encoding genes cause pleiotropic phenotypes that include enhanced hypocotyl elongation, defects in cell wall thickening, early flowering, early senescence and impaired pollen tube growth. Our results indicate essential roles of Hyp O-arabinosylation in both vegetative and reproductive growth in plants.
Asunto(s)
Arabidopsis/enzimología , Hidroxiprolina/metabolismo , Pentosiltransferasa/análisis , Pentosiltransferasa/metabolismo , Conformación MolecularRESUMEN
Plants initiate specific defense responses by recognizing conserved epitope peptides within the flagellin proteins derived from bacteria. Proteolytic cleavage of epitope peptides from flagellin by plant apoplastic proteases is thought to be crucial for the perception of the epitope by the plant receptor. However, the identity of the plant proteases involved in this process remains unknown. Here, we establish an efficient identification system for the target proteases in Arabidopsis apoplastic fluid; the method employs native two-dimensional electrophoresis followed by an in-gel proteolytic assay using a fluorescence-quenching peptide substrate. We designed a substrate to specifically detect proteolytic activity at the C-terminus of the flg22 epitope in flagellin and identified two plant subtilases, SBT5.2 and SBT1.7, as specific proteases responsible for the C-terminal cleavage of flg22. In the apoplastic fluid of Arabidopsis mutant plants deficient in these two proteases, we observe a decrease in the C-terminal cleavage of the flg22 domain from flagellin, leading to a decrease in the efficiency of flg22 epitope liberation. Consequently, defensive reactive oxygen species (ROS) production is delayed in sbt5.2 sbt1.7 double-mutant leaf disks compared to wild type following flagellin exposure.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Epítopos , Flagelina , Especies Reactivas de Oxígeno , Subtilisinas , Arabidopsis/inmunología , Arabidopsis/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/inmunología , Epítopos/inmunología , Epítopos/metabolismo , Flagelina/metabolismo , Flagelina/inmunología , Mutación , Proteolisis , Especies Reactivas de Oxígeno/metabolismo , Subtilisinas/metabolismo , Subtilisinas/genéticaRESUMEN
Unlike plants in the field, which experience significant temporal fluctuations in environmental conditions, plants in the laboratory are typically grown in controlled, stable environments. Therefore, signaling pathways evolved for survival in fluctuating environments often remain functionally latent in laboratory settings. Here, we show that TGA1 and TGA4 act as hub transcription factors through which the expression of genes involved in high-affinity nitrate uptake are regulated in response to shoot-derived phloem mobile polypeptides, CEP DOWNSTREAM 1 (CEPD1), CEPD2 and CEPD-like 2 (CEPDL2) as nitrogen (N) deficiency signals, and Glutaredoxin S1 (GrxS1) to GrxS8 as N sufficiency signals. CEPD1/2/CEPDL2 and GrxS1-S8 competitively bind to TGA1/4 in roots, with the former acting as transcription coactivators that enhance the uptake of nitrate, while the latter function as corepressor complexes together with TOPLESS (TPL), TPL-related 1 (TPR1) and TPR4 to limit nitrate uptake. Arabidopsis plants deficient in TGA1/4 maintain basal nitrate uptake and exhibit growth similar to wild-type plants in a stable N environment, but are impaired in regulation of nitrate acquisition in response to shoot N demand, leading to defective growth under fluctuating N environments where rhizosphere nitrate ions switch periodically between deficient and sufficient states. TGA1/4 are crucial transcription factors that enable plants to survive under fluctuating and challenging N environmental conditions.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Regulación de la Expresión Génica de las Plantas , Nitratos , Nitrógeno , Raíces de Plantas , Brotes de la Planta , Transducción de Señal , Factores de Transcripción , Arabidopsis/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Nitrógeno/metabolismo , Raíces de Plantas/metabolismo , Raíces de Plantas/crecimiento & desarrollo , Brotes de la Planta/metabolismo , Brotes de la Planta/genética , Nitratos/metabolismo , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Péptidos/metabolismo , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/genéticaRESUMEN
Legumes control root nodule symbiosis (RNS) in response to environmental nitrogen availability. Despite the recent understanding of the molecular basis of external nitrate-mediated control of RNS, it remains mostly elusive how plants regulate physiological processes depending on internal nitrogen status. In addition, iron (Fe) acts as an essential element that enables symbiotic nitrogen fixation; however, the mechanism of Fe accumulation in nodules is poorly understood. Here, we focus on the transcriptome in response to internal nitrogen status during RNS in Lotus japonicus and identify that IRON MAN (IMA) peptide genes are expressed during symbiotic nitrogen fixation. We show that LjIMA1 and LjIMA2 expressed in the shoot and root play systemic and local roles in concentrating internal Fe to the nodule. Furthermore, IMA peptides have conserved roles in regulating nitrogen homeostasis by adjusting nitrogen-Fe balance in L. japonicus and Arabidopsis thaliana. These findings indicate that IMA-mediated Fe provision plays an essential role in regulating nitrogen-related physiological processes.
Asunto(s)
Arabidopsis , Lotus , Humanos , Nódulos de las Raíces de las Plantas/metabolismo , Nitrógeno , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Lotus/metabolismo , Fijación del Nitrógeno/fisiología , Simbiosis/fisiología , Homeostasis , Arabidopsis/genética , Arabidopsis/metabolismo , Regulación de la Expresión Génica de las Plantas , Nodulación de la Raíz de la Planta/genéticaRESUMEN
Leucine-rich repeat receptor kinases (LRR-RKs) are the largest sub-family of transmembrane receptor kinases in plants. In several LRR-RKs, a loop-out region called an 'island domain', which intercepts the extracellular tandem LRRs at a position near the transmembrane domain, constitutes the ligand-binding pocket, but the absence of the island domain in numerous LRR-RKs raises questions about which domain recognizes the ligand in non-island domain LRR-RKs. Here, we used photoaffinity labeling followed by chemical and enzymatic digestion to show that BAM1, a CLV1/BAM-family LRR-RK whose extracellular domain comprises 22 consecutive LRRs, directly interacts with the small peptide ligand CLE9 at the LRR6-LRR8 region that is relatively distal from the transmembrane domain. Multiple sequence alignment and homology modeling revealed that the inner concave side of LRR6-LRR8 of CLV1/BAM-family LRR-RKs deviates slightly from the LRR consensus. In support of our findings, the clv1-4 mutant carries a missense mutation at the inner concave side of LRR6 of CLV1, and introduction of the corresponding mutation in BAM1 resulted in complete loss of ligand binding activity. Our results indicate that the ligand recognition mechanisms of plant LRR-RKs are more complex and diverse than anticipated.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Leucina/metabolismo , Mapeo de Interacción de Proteínas/métodos , Proteínas Serina-Treonina Quinasas/metabolismo , Secuencia de Aminoácidos , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Sitios de Unión , Secuencia de Consenso , Electroforesis en Gel de Poliacrilamida , Ligandos , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida , Mutación Missense , Fragmentos de Péptidos/metabolismo , Etiquetas de Fotoafinidad/metabolismo , Unión Proteica , Proteínas Serina-Treonina Quinasas/genética , Estructura Terciaria de Proteína , Proteínas Tirosina Quinasas Receptoras/genética , Proteínas Tirosina Quinasas Receptoras/metabolismo , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Nicotiana/genética , Nicotiana/metabolismoRESUMEN
Arabinosylation of hydroxyproline (Hyp) is a post-translational modification often found in secreted peptide signals in plants. The physiological importance of this modification was highlighted by the finding that CLAVATA3 (CLV3), a key peptide signal for regulating the fate of stem cells in the shoot apical meristem in Arabidopsis, contains three l-arabinose residues linked via linear ß-1,2-linkages. However, understanding the functions and properties of arabinosylated peptides has been hindered by difficulties in synthesizing the complex arabinose chain. Here we report the stereoselective total synthesis of ß-1,2-linked triarabinosylated CLV3 peptide ([Ara3]CLV3). Chemically synthesized [Ara3]CLV3 restricted stem cell activity more effectively than did unmodified CLV3 peptide. Comparison of mono-, di- and triarabinosylated CLV3 glycopeptides revealed that the biological activity increased progressively as the arabinose chain length increased. Thus, the arabinose chain length of CLV3 is important for its biological activity. Nuclear magnetic resonance spectroscopy and nuclear Overhauser effect-based structure calculations further revealed the structural impact of the arabinose chain on peptide conformation. The arabinose chain of [Ara3]CLV3 extends toward the C-terminal end of the peptide, and its non-reducing end is positioned proximal to the peptide backbone. Consequently, the arabinose chain causes distinct distortion in the C-terminal half of the peptide in a highly directional manner. The established synthetic route of [Ara3]CLV3 will greatly contribute to our understanding of the biology and biochemistry of arabinosylated peptide signals in plants.
Asunto(s)
Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/metabolismo , Arabidopsis/efectos de los fármacos , Modelos Moleculares , Procesamiento Proteico-Postraduccional , Arabidopsis/crecimiento & desarrollo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/síntesis química , Proteínas de Arabidopsis/farmacología , Arabinosa/química , Glicopéptidos/síntesis química , Glicopéptidos/química , Glicopéptidos/metabolismo , Glicopéptidos/farmacología , Hidroxiprolina/química , Espectroscopía de Resonancia Magnética , Meristema/efectos de los fármacos , Meristema/crecimiento & desarrollo , Meristema/metabolismo , Brotes de la Planta/efectos de los fármacos , Brotes de la Planta/crecimiento & desarrollo , Brotes de la Planta/metabolismo , Conformación Proteica , Plantones/efectos de los fármacos , Plantones/crecimiento & desarrollo , Plantones/metabolismo , Transducción de SeñalRESUMEN
Peptide signaling plays a major role in various aspects of plant growth and development, as has been shown in recent biochemical, genetic and bioinformatic studies. There are over a dozen secreted peptides recognized in plants known to regulate cellular functions. To become functional, these secreted peptide signals often undergo post-translational modifications, such as tyrosine sulfation, proline hydroxylation, and hydroxyproline arabinosylation, and successive proteolytic processing. These types of 'small post-translationally modified peptide signals' are one of the major groups of peptide signals found in plants. In parallel with the discovery of peptide signals, specific receptors for such peptide signals were identified as being membrane-localized leucine-rich repeat receptor kinases. This short review highlights the recent progress in research on small post-translationally modified peptide signals, including our own research.
Asunto(s)
Péptidos/metabolismo , Proteínas de Plantas/metabolismo , Plantas/metabolismo , Procesamiento Proteico-Postraduccional , Señales de Clasificación de Proteína/fisiología , Secuencia de Aminoácidos , Distinciones y Premios , Japón , Biología Molecular , Datos de Secuencia Molecular , Transducción de Señal , Sociedades MédicasRESUMEN
Ultrasmall algae have attracted the attention of biologists investigating the basic mechanisms underlying living systems. Their potential as effective organisms for producing useful substances is also of interest in bioindustry. Although genomic information is indispensable for elucidating metabolism and promoting molecular breeding, many ultrasmall algae remain genetically uncharacterized. Here, we present the nuclear genome sequence of an ultrasmall green alga of freshwater habitats, Medakamo hakoo. Evolutionary analyses suggest that this species belongs to a new genus within the class Trebouxiophyceae. Sequencing analyses revealed that its genome, comprising 15.8 Mbp and 7629 genes, is among the smallest known genomes in the Viridiplantae. Its genome has relatively few genes associated with genetic information processing, basal transcription factors, and RNA transport. Comparative analyses revealed that 1263 orthogroups were shared among 15 ultrasmall algae from distinct phylogenetic lineages. The shared gene sets will enable identification of genes essential for algal metabolism and cellular functions.
Asunto(s)
Chlorophyta , Genoma , Filogenia , Chlorophyta/genética , Genómica , Agua DulceRESUMEN
Tyrosine sulfation is a posttranslational modification common in peptides and proteins synthesized by the secretory pathway in most eukaryotes. In plants, this modification is critical for the biological activities of a subset of peptide hormones such as PSK and PSY1. In animals, tyrosine sulfation is catalyzed by Golgi-localized type II transmembrane proteins called tyrosylprotein sulfotransferases (TPSTs). However, no orthologs of animal TPST genes have been found in plants, suggesting that plants have evolved plant-specific TPSTs structurally distinct from their animal counterparts. To investigate the mechanisms of tyrosine sulfation in plants, we purified TPST activity from microsomal fractions of Arabidopsis MM2d cells, and identified a 62-kDa protein that specifically interacts with the sulfation motif of PSY1 precursor peptide. This protein is a 500-aa type I transmembrane protein that shows no sequence similarity to animal TPSTs. A recombinant version of this protein expressed in yeast catalyzed tyrosine sulfation of both PSY1 and PSK precursor polypeptide in vitro, indicating that the newly identified protein is indeed an Arabidopsis (At)TPST. AtTPST is expressed throughout the plant body, and the highest levels of expression are in the root apical meristem. A loss-of-function mutant of AtTPST displayed a marked dwarf phenotype accompanied by stunted roots, pale green leaves, reduction in higher order veins, early senescence, and a reduced number of flowers and siliques. Our results indicate that plants and animals independently acquired tyrosine sulfation enzymes through convergent evolution.
Asunto(s)
Arabidopsis/enzimología , Sulfotransferasas/metabolismo , Secuencia de Aminoácidos , Arabidopsis/crecimiento & desarrollo , Cromatografía de Afinidad , Clonación Molecular , Secuencia Conservada , Regulación de la Expresión Génica de las Plantas , Humanos , Datos de Secuencia Molecular , Hormonas Peptídicas/metabolismo , Precursores de Proteínas/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Alineación de Secuencia , Fracciones Subcelulares/metabolismo , Especificidad por Sustrato , Sulfotransferasas/química , Sulfotransferasas/genética , Sulfotransferasas/aislamiento & purificaciónRESUMEN
Deciding whether to grow or to divert energy to stress responses is a major physiological trade-off for plants surviving in fluctuating environments. We show that three leucine-rich repeat receptor kinases (LRR-RKs) act as direct ligand-perceiving receptors for PLANT PEPTIDE CONTAINING SULFATED TYROSINE (PSY)-family peptides and mediate switching between two opposing pathways. By contrast to known LRR-RKs, which activate signaling upon ligand binding, PSY receptors (PSYRs) activate the expression of various genes encoding stress response transcription factors upon depletion of the ligands. Loss of PSYRs results in defects in plant tolerance to both biotic and abiotic stresses. This ligand-deprivation-dependent activation system potentially enables plants to exert tuned regulation of stress responses in the tissues proximal to metabolically dysfunctional damaged sites where ligand production is impaired.