RESUMEN
Pea is a grain legume crop with a high potential to accelerate the food transition due to its high seed protein content and relatively well-balanced amino acid composition. The critical role of external sulfur (S) supply in determining seed yield and seed quality in pea makes it essential to understand the impact of whole plant S management on the trade-off between these two traits. Here, we investigated the physiological relevance of vacuolar sulfate remobilization by targeting PsSULTR4, the only pea sulfate transporter showing substantial similarity to the vacuolar sulfate exporter AtSULTR4;1. Five mutations in PsSULTR4 were identified by TILLING (Targeting Induced Local Lesions IN Genomes), two of which, a loss of function (W78*) and a missense (E568K), significantly decreased seed yield under S deprivation. We demonstrate that PsSULTR4 triggers S distribution from source tissues, especially lower leaves, to reproductive organs to maintain seed yield under S deficiency. Under sufficient S supply, sultr4 seeds display lower levels of the S-rich storage protein PA1 at maturity. They also overaccumulate sulfate in the endosperm at the onset of seed filling. These findings uncover a role of PsSULTR4 in the remobilization of vacuolar sulfate during embryo development, allowing the efficient synthesis of S-rich proteins. Our study uncovers that PsSULTR4 functions (i) in source tissues to remobilize stored vacuolar sulfate for seed production under low S availability and (ii) in developing seeds well supplied with S to fine-tune sulfate remobilization from the endosperm as a critical control point for storage activities in the embryo.
RESUMEN
The afila (af) mutation causes the replacement of leaflets by a branched mass of tendrils in the compound leaves of pea - Pisum sativum L. This mutation was first described in 1953, and several reports of spontaneous af mutations and induced mutants with a similar phenotype exist. Despite widespread introgression into breeding material, the nature of af and the origin of the alleles used remain unknown. Here, we combine comparative genomics with reverse genetic approaches to elucidate the genetic determinants of af. We also investigate haplotype diversity using a set of AfAf and afaf cultivars and breeding lines and molecular markers linked to seven consecutive genes. Our results show that deletion of two tandemly arranged genes encoding Q-type Cys(2)His(2) zinc finger transcription factors, PsPALM1a and PsPALM1b, is responsible for the af phenotype in pea. Eight haplotypes were identified in the af-harbouring genomic region on chromosome 2. These haplotypes differ in the size of the deletion, covering more or less genes. Diversity at the af locus is valuable for crop improvement and sheds light on the history of pea breeding for improved standing ability. The results will be used to understand the function of PsPALM1a/b and to transfer the knowledge for innovation in related crops.
Asunto(s)
Haplotipos , Fenotipo , Pisum sativum , Fitomejoramiento , Pisum sativum/genética , Haplotipos/genética , Genes de Plantas , Proteínas de Plantas/genética , Mutación/genética , Hojas de la Planta/genética , Cruzamiento , Factores de Transcripción/genética , Variación GenéticaRESUMEN
Improving and stabilizing the quality of seed proteins are of growing interest in the current food and agroecological transitions. Sulfur is a key determinant of this quality since it is essential for the synthesis of sulfur-rich proteins in seeds. A lack of sulfur provokes drastic changes in seed protein composition, negatively impacting the nutritional and functional properties of proteins, and leading in some cases to diseases or health problems in humans. Sulfur also plays a crucial role in stress tolerance through the synthesis of antioxidant or protective molecules. In the context of climate change, questions arise regarding the trade-off between seed yield and seed quality with respect to sulfur availability and use by crops that represent important sources of proteins for human nutrition. Here, we review recent work obtained in legumes, cereals, as well as in Arabidopsis, that present major advances on: (i) the interaction between sulfur nutrition and environmental or nutritional stresses with regard to seed yield and protein composition; (ii) metabolic pathways that merit to be targeted to mitigate negative impacts of environmental stresses on seed protein quality; and (iii) the importance of sulfur homeostasis for the regulation of seed protein composition and its interplay with seed redox homeostasis.
Asunto(s)
Arabidopsis , Semillas , Humanos , Semillas/metabolismo , Grano Comestible/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Arabidopsis/metabolismo , Azufre/metabolismo , Estrés FisiológicoRESUMEN
Medicago truncatula NODULE ROOT1 (MtNOOT1) and Pisum sativum COCHLEATA1 (PsCOCH1) are orthologous genes belonging to the NOOT-BOP-COCH-LIKE (NBCL) gene family which encodes key transcriptional co-regulators of plant development. In Mtnoot1 and Pscoch1 mutants, the development of stipules, flowers, and symbiotic nodules is altered. MtNOOT2 and PsCOCH2 represent the single paralogues of MtNOOT1 and PsCOCH1, respectively. In M. truncatula, MtNOOT1 and MtNOOT2 are both required for the establishment and maintenance of symbiotic nodule identity. In legumes, the role of NBCL2 in above-ground development is not known. To better understand the roles of NBCL genes in legumes, we used M. truncatula and P. sativum nbcl mutants, isolated a knockout mutant for the PsCOCH2 locus and generated Pscoch1coch2 double mutants in P. sativum. Our work shows that single Mtnoot2 and Pscoch2 mutants develop wild-type stipules, flowers, and symbiotic nodules. However, the number of flowers was increased and the pods and seeds were smaller compared to the wild type. Furthermore, in comparison to the corresponding nbcl1 single mutants, both the M. truncatula and P. sativum nbcl double mutants show a drastic alteration in stipule, inflorescence, flower, and nodule development. Remarkably, in both M. truncatula and P. sativum nbcl double mutants, stipules are transformed into a range of aberrant leaf-like structures.
Asunto(s)
Medicago truncatula , Nódulos de las Raíces de las Plantas , Nódulos de las Raíces de las Plantas/metabolismo , Regulación de la Expresión Génica de las Plantas , Pisum sativum/genética , Medicago truncatula/metabolismo , Simbiosis/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Fijación del Nitrógeno/genética , MutaciónRESUMEN
As the frequency of extreme environmental events is expected to increase with climate change, identifying candidate genes for stabilizing the protein composition of legume seeds or optimizing this in a given environment is increasingly important. To elucidate the genetic determinants of seed protein plasticity, major seed proteins from 200 ecotypes of Medicago truncatula grown in four contrasting environments were quantified after one-dimensional electrophoresis. The plasticity index of these proteins was recorded for each genotype as the slope of Finlay and Wilkinson's regression and then used for genome-wide association studies (GWASs), enabling the identification of candidate genes for determining this plasticity. This list was enriched in genes related to transcription, DNA repair and signal transduction, with many of them being stress responsive. Other over-represented genes were related to sulfur and aspartate family pathways leading to the synthesis of the nutritionally essential amino acids methionine and lysine. By placing these genes in metabolic pathways, and using a M. truncatula mutant impaired in regenerating methionine from S-methylmethionine, we discovered that methionine recycling pathways are major contributors to globulin composition establishment and plasticity. These data provide a unique resource of genes that can be targeted to mitigate negative impacts of environmental stresses on seed protein composition.
Asunto(s)
Medicago truncatula/genética , Proteínas de Almacenamiento de Semillas/metabolismo , Estudio de Asociación del Genoma Completo , Genotipo , Globulinas/genética , Globulinas/metabolismo , Medicago truncatula/fisiología , Metionina/metabolismo , Mutación , Fenotipo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Proteínas de Almacenamiento de Semillas/genética , Semillas/genética , Semillas/fisiología , Estrés Fisiológico , Vitamina U/metabolismoRESUMEN
The use of pulses as ingredients for the production of food products rich in plant proteins is increasing. However, protein fractions prepared from pea or other pulses contain significant amounts of saponins, glycosylated triterpenes that can impart an undesirable bitter taste when used as an ingredient in foodstuffs. In this article, we describe the identification and characterization of a gene involved in saponin biosynthesis during pea seed development, by screening mutants obtained from two Pisum sativum TILLING (Targeting Induced Local Lesions IN Genomes) populations in two different genetic backgrounds. The mutations studied are located in a gene designated PsBAS1 (ß-amyrin synthase1), which is highly expressed in maturing pea seeds and which encodes a protein previously shown to correspond to an active ß-amyrin synthase. The first allele is a nonsense mutation, while the second mutation is located in a splice site and gives rise to a mis-spliced transcript encoding a truncated, nonfunctional protein. The homozygous mutant seeds accumulated virtually no saponin without affecting the seed nutritional or physiological quality. Interestingly, BAS1 appears to control saponin accumulation in all other tissues of the plant examined. These lines represent a first step in the development of pea varieties lacking bitterness off-flavors in their seeds. Our work also shows that TILLING populations in different genetic backgrounds represent valuable genetic resources for both crop improvement and functional genomics.
Asunto(s)
Transferasas Intramoleculares/metabolismo , Pisum sativum/metabolismo , Proteínas de Plantas/metabolismo , Saponinas/metabolismo , Regulación de la Expresión Génica de las Plantas , Transferasas Intramoleculares/genética , Mutación con Pérdida de Función , Pisum sativum/genética , Proteínas de Plantas/genética , Saponinas/química , Saponinas/genética , Semillas/genética , Semillas/crecimiento & desarrollo , Semillas/metabolismo , Análisis Espacio-TemporalRESUMEN
Nitrogen-fixing root nodulation in legumes challenged with nitrogen-limiting conditions requires infection of the root hairs by soil symbiotic bacteria, collectively referred to as rhizobia, and the initiation of cell divisions in the root cortex. Cytokinin hormones are critical for early nodulation to coordinate root nodule organogenesis and the progression of bacterial infections. Cytokinin signaling involves regulation of the expression of cytokinin primary response genes by type-B response regulator (RRB) transcription factors. RNA interference or mutation of MtRRB3, the RRB-encoding gene most strongly expressed in Medicago truncatula roots and nodules, significantly decreased the number of nodules formed, indicating a function of this RRB in nodulation initiation. Fewer infection events were also observed in rrb3 mutant roots associated with a reduced Nod factor induction of the Early Nodulin 11 (MtENOD11) infection marker, and of the cytokinin-regulated Nodulation Signaling Pathway 2 (Mt NSP2) gene. Rhizobial infections correlate with an expansion of the nuclear area, suggesting the activation of endoreduplication cycles linked to the cytokinin-regulated Cell Cycle Switch 52A (Mt CCS52A) gene. Although no significant difference in nucleus size and endoreduplication were detected in rhizobia-infected rrb3 mutant roots, expression of the MtCCS52A endoreduplication marker was reduced. As the MtRRB3 expression pattern overlaps with those of MtNSP2 and MtCCS52A in roots and nodule primordia, chromatin immunoprecipitation-quantitative PCR and protoplast trans-activation assays were used to show that MtRRB3 can interact with and trans-activate MtNSP2 and MtCCS52A promoters. Overall, we highlight that the MtRRB3 cytokinin signaling transcription factor coordinates the expression of key early nodulation genes.
Asunto(s)
Citocininas/metabolismo , Nodulación de la Raíz de la Planta , Transducción de Señal , Factores de Transcripción/metabolismo , Tamaño del Núcleo Celular , Endorreduplicación , Regulación de la Expresión Génica de las Plantas , Genes de Plantas , Medicago truncatula/genética , Medicago truncatula/microbiología , Fenotipo , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Nodulación de la Raíz de la Planta/genética , Regiones Promotoras Genéticas , Unión Proteica , Dominios Proteicos , Sinorhizobium meliloti/fisiología , Activación Transcripcional/genéticaRESUMEN
Pea is a legume crop producing protein-rich seeds and is increasingly in demand for human consumption and animal feed. The aim of this study was to explore the proteome of developing pea seeds at three key stages covering embryogenesis, the transition to seed-filling, and the beginning of storage-protein synthesis, and to investigate how the proteome was influenced by S deficiency and water stress, applied either separately or combined. Of the 3184 proteins quantified by shotgun proteomics, 2473 accumulated at particular stages, thus providing insights into the proteome dynamics at these stages. Differential analyses in response to the stresses and inference of a protein network using the whole proteomics dataset identified a cluster of antioxidant proteins (including a glutathione S-transferase, a methionine sulfoxide reductase, and a thioredoxin) possibly involved in maintaining redox homeostasis during early seed development and preventing cellular damage under stress conditions. Integration of the proteomics data with previously obtained transcriptomics data at the transition to seed-filling revealed the transcriptional events associated with the accumulation of the stress-regulated antioxidant proteins. This transcriptional defense response involves genes of sulfate homeostasis and assimilation, thus providing candidates for targeted studies aimed at dissecting the signaling cascade linking S metabolism to antioxidant processes in developing seeds.
Asunto(s)
Pisum sativum , Proteómica , Antioxidantes , Deshidratación , Regulación de la Expresión Génica de las Plantas , Pisum sativum/genética , Pisum sativum/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Semillas/metabolismo , Azufre/metabolismoRESUMEN
Plant -specific lysin-motif receptor-like kinases (LysM-RLKs) are implicated in the perception of N-acetyl glucosamine-containing compounds, some of which are important signal molecules in plant-microbe interactions. Among these, both lipo-chitooligosaccharides (LCOs) and chitooligosaccharides (COs) are proposed as arbuscular mycorrhizal (AM) fungal symbiotic signals. COs can also activate plant defence, although there are scarce data about CO production by pathogens, especially nonfungal pathogens. We tested Medicago truncatula mutants in the LysM-RLK MtLYK9 for their abilities to interact with the AM fungus Rhizophagus irregularis and the oomycete pathogen Aphanomyces euteiches. This prompted us to analyse whether A. euteiches can produce COs. Compared with wild-type plants, Mtlyk9 mutants had fewer infection events and were less colonised by the AM fungus. By contrast, Mtlyk9 mutants were more heavily infected by A. euteiches and showed more disease symptoms. Aphanomyces euteiches was also shown to produce short COs, mainly CO II, but also CO III and CO IV, and traces of CO V, both ex planta and in planta. MtLYK9 thus has a dual role in plant immunity and the AM symbiosis, which raises questions about the functioning and the ancestral origins of such a receptor protein.
Asunto(s)
Glomeromycota/fisiología , Medicago truncatula/microbiología , Micorrizas/fisiología , Inmunidad de la Planta , Proteínas de Plantas/metabolismo , Simbiosis , Secuencia de Aminoácidos , Aphanomyces/fisiología , Quitina/análogos & derivados , Quitina/biosíntesis , Quitosano , Regulación de la Expresión Génica de las Plantas , Medicago truncatula/genética , Mutación/genética , Oligosacáridos , Proteínas de Plantas/química , Proteínas de Plantas/genéticaRESUMEN
MAIN CONCLUSION: The LysM receptor-like kinase K1 is involved in regulation of pea-rhizobial symbiosis development. The ability of the crop legume Pisum sativum L. to perceive the Nod factor rhizobial signals may depend on several receptors that differ in ligand structure specificity. Identification of pea mutants defective in two types of LysM receptor-like kinases (LysM-RLKs), SYM10 and SYM37, featuring different phenotypic manifestations and impaired at various stages of symbiosis development, corresponds well to this assumption. There is evidence that one of the receptor proteins involved in symbiosis initiation, SYM10, has an inactive kinase domain. This implies the presence of an additional component in the receptor complex, together with SYM10, that remains unknown. Here, we describe a new LysM-RLK, K1, which may serve as an additional component of the receptor complex in pea. To verify the function of K1 in symbiosis, several P. sativum non-nodulating mutants in the k1 gene were identified using the TILLING approach. Phenotyping revealed the blocking of symbiosis development at an appropriately early stage, strongly suggesting the importance of LysM-RLK K1 for symbiosis initiation. Moreover, the analysis of pea mutants with weaker phenotypes provides evidence for the additional role of K1 in infection thread distribution in the cortex and rhizobia penetration. The interaction between K1 and SYM10 was detected using transient leaf expression in Nicotiana benthamiana and in the yeast two-hybrid system. Since the possibility of SYM10/SYM37 complex formation was also shown, we tested whether the SYM37 and K1 receptors are functionally interchangeable using a complementation test. The interaction between K1 and other receptors is discussed.
Asunto(s)
Pisum sativum/enzimología , Proteínas de Plantas/fisiología , Proteínas Quinasas/fisiología , Rhizobium leguminosarum/fisiología , Simbiosis , Western Blotting , Ingeniería Genética/métodos , Pisum sativum/microbiología , Pisum sativum/fisiología , Hojas de la Planta/enzimología , Hojas de la Planta/metabolismo , Plantas Modificadas Genéticamente , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Nicotiana/genética , Técnicas del Sistema de Dos HíbridosRESUMEN
Improving nutritional seed quality is an important challenge in grain legume breeding. However, the genes controlling the differential accumulation of globulins, which are major contributors to seed nutritional value in legumes, remain largely unknown. We combined a search for protein quantity loci with genome-wide association studies on the abundance of 7S and 11S globulins in seeds of the model legume species Medicago truncatula. Identified genomic regions and genes carrying polymorphisms linked to globulin variations were then cross-compared with pea (Pisum sativum), leading to the identification of candidate genes for the regulation of globulin abundance in this crop. Key candidates identified include genes involved in transcription, chromatin remodeling, post-translational modifications, transport and targeting of proteins to storage vacuoles. Inference of a gene coexpression network of 12 candidate transcription factors and globulin genes revealed the transcription factor ABA-insensitive 5 (ABI5) as a highly connected hub. Characterization of loss-of-function abi5 mutants in pea uncovered a role for ABI5 in controlling the relative abundance of vicilin, a sulfur-poor 7S globulin, in pea seeds. This demonstrates the feasibility of using genome-wide association studies in M. truncatula to reveal genes that can be modulated to improve seed nutritional value.
Asunto(s)
Globulinas/metabolismo , Medicago truncatula/genética , Medicago truncatula/metabolismo , Semillas/metabolismo , Electroforesis en Gel Bidimensional , Regulación de la Expresión Génica de las Plantas , Redes Reguladoras de Genes , Estudio de Asociación del Genoma Completo , Globulinas/genética , Mutación , Pisum sativum/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Transporte de Proteínas , Proteómica/métodos , Proteínas de Almacenamiento de Semillas/genética , Proteínas de Almacenamiento de Semillas/metabolismo , Semillas/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
PUB1, an E3 ubiquitin ligase, which interacts with and is phosphorylated by the LYK3 symbiotic receptor kinase, negatively regulates rhizobial infection and nodulation during the nitrogen-fixing root nodule symbiosis in Medicago truncatula In this study, we show that PUB1 also interacts with and is phosphorylated by DOES NOT MAKE INFECTIONS 2, the key symbiotic receptor kinase of the common symbiosis signaling pathway, required for both the rhizobial and the arbuscular mycorrhizal (AM) endosymbioses. We also show here that PUB1 expression is activated during successive stages of root colonization by Rhizophagus irregularis that is compatible with its interaction with DOES NOT MAKE INFECTIONS 2. Through characterization of a mutant, pub1-1, affected by the E3 ubiquitin ligase activity of PUB1, we have shown that the ubiquitination activity of PUB1 is required to negatively modulate successive stages of infection and development of rhizobial and AM symbioses. In conclusion, PUB1 represents, to our knowledge, a novel common component of symbiotic signaling integrating signal perception through interaction with and phosphorylation by two key symbiotic receptor kinases, and downstream signaling via its ubiquitination activity to fine-tune both rhizobial and AM root endosymbioses.
Asunto(s)
Medicago truncatula/metabolismo , Medicago truncatula/microbiología , Micorrizas/fisiología , Proteínas de Plantas/metabolismo , Rhizobium/fisiología , Simbiosis , Ubiquitinación , Recuento de Colonia Microbiana , Glomeromycota/fisiología , Micorrizas/crecimiento & desarrollo , Fosforilación , Proteínas de Plantas/química , Dominios Proteicos , Saccharomyces cerevisiae/metabolismo , Ubiquitina-Proteína Ligasas/metabolismoRESUMEN
The endosperm plays a pivotal role in the integration between component tissues of molecular signals controlling seed development. It has been shown to participate in the regulation of embryo morphogenesis and ultimately seed size determination. However, the molecular mechanisms that modulate seed size are still poorly understood especially in legumes. DASH (DOF Acting in Seed embryogenesis and Hormone accumulation) is a DOF transcription factor (TF) expressed during embryogenesis in the chalazal endosperm of the Medicago truncatula seed. Phenotypic characterization of three independent dash mutant alleles revealed a role for this TF in the prevention of early seed abortion and the determination of final seed size. Strong loss-of-function alleles cause severe defects in endosperm development and lead to embryo growth arrest at the globular stage. Transcriptomic analysis of dash pods versus wild-type (WT) pods revealed major transcriptional changes and highlighted genes that are involved in auxin transport and perception as mainly under-expressed in dash mutant pods. Interestingly, the exogenous application of auxin alleviated the seed-lethal phenotype, whereas hormonal dosage revealed a much higher auxin content in dash pods compared with WT. Together these results suggested that auxin transport/signaling may be affected in the dash mutant and that aberrant auxin distribution may contribute to the defect in embryogenesis resulting in the final seed size phenotype.
Asunto(s)
Ácidos Indolacéticos/metabolismo , Medicago truncatula/metabolismo , Proteínas de Plantas/fisiología , Semillas/crecimiento & desarrollo , Factores de Transcripción/fisiología , Transporte Biológico/genética , Regulación de la Expresión Génica de las Plantas , Homeostasis , Medicago truncatula/embriología , Medicago truncatula/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Semillas/genética , Semillas/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
Among a set of genes in pea (Pisum sativum L.) that were induced under drought-stress growth conditions, one encoded a protein with significant similarity to a regulator of chlorophyll catabolism, SGR. This gene, SGRL, is distinct from SGR in genomic location, encoded carboxy-terminal motif, and expression through plant and seed development. Divergence of the two encoded proteins is associated with a loss of similarity in intron/exon gene structure. Transient expression of SGRL in leaves of Nicotiana benthamiana promoted the degradation of chlorophyll, in a manner that was distinct from that shown by SGR. Removal of a predicted transmembrane domain from SGRL reduced its activity in transient expression assays, although variants with and without this domain reduced SGR-induced chlorophyll degradation, indicating that the effects of the two proteins are not additive. The combined data suggest that the function of SGRL during growth and development is in chlorophyll re-cycling, and its mode of action is distinct from that of SGR. Studies of pea sgrL mutants revealed that plants had significantly lower stature and yield, a likely consequence of reduced photosynthetic efficiencies in mutant compared with control plants under conditions of high light intensity.
Asunto(s)
Clorofila/metabolismo , Pisum sativum/crecimiento & desarrollo , Pisum sativum/metabolismo , Proteínas de Plantas/metabolismo , Secuencia de Aminoácidos , Regulación del Desarrollo de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Genes de Plantas , Datos de Secuencia Molecular , Mutación , Pisum sativum/genética , Fotosíntesis/genética , Filogenia , Hojas de la Planta/metabolismo , Proteínas de Plantas/química , Proteínas de Plantas/genética , Plantas Modificadas Genéticamente , Estructura Terciaria de Proteína , Homología de Secuencia de Aminoácido , Nicotiana/genética , Nicotiana/metabolismoRESUMEN
Reductions in sulfur dioxide emissions and the use of sulfur-free mineral fertilizers are decreasing soil sulfur levels and threaten the adequate fertilization of most crops. To provide knowledge regarding legume adaptation to sulfur restriction, we subjected Medicago truncatula, a model legume species, to sulfur deficiency at various developmental stages, and compared the yield, nutrient allocation and seed traits. This comparative analysis revealed that sulfur deficiency at the mid-vegetative stage decreased yield and altered the allocation of nitrogen and carbon to seeds, leading to reduced levels of major oligosaccharides in mature seeds, whose germination was dramatically affected. In contrast, during the reproductive period, sulfur deficiency had little influence on yield and nutrient allocation, but the seeds germinated slowly and were characterized by low levels of a biotinylated protein, a putative indicator of germination vigor that has not been previously related to sulfur nutrition. Significantly, plants deprived of sulfur at an intermediary stage (flowering) adapted well by remobilizing nutrients from source organs to seeds, ensuring adequate quantities of carbon and nitrogen in seeds. This efficient remobilization of photosynthates may be explained by vacuolar sulfate efflux to maintain leaf metabolism throughout reproductive growth, as suggested by transcript and metabolite profiling. The seeds from these plants, deprived of sulfur at the floral transition, contained normal levels of major oligosaccharides but their germination was delayed, consistent with low levels of sucrose and the glycolytic enzymes required to restart seed metabolism during imbibition. Overall, our findings provide an integrative view of the legume response to sulfur deficiency.
Asunto(s)
Adaptación Fisiológica , Medicago truncatula/fisiología , Semillas/fisiología , Azufre/deficiencia , Transporte Biológico , Biomasa , Metabolismo de los Hidratos de Carbono , Carbono/metabolismo , Clorofila/metabolismo , Medicago truncatula/genética , Medicago truncatula/crecimiento & desarrollo , Modelos Biológicos , Nitrógeno/metabolismo , Oligosacáridos/metabolismo , Especificidad de Órganos , Oxidación-Reducción , Fenotipo , Hojas de la Planta/genética , Hojas de la Planta/crecimiento & desarrollo , Hojas de la Planta/fisiología , ARN Mensajero/genética , Rafinosa/metabolismo , Semillas/genética , Semillas/crecimiento & desarrollo , Sulfatos/metabolismo , Azufre/metabolismoRESUMEN
Seeds of several agriculturally important legumes are rich sources of the only halogenated plant hormone, 4-chloroindole-3-acetic acid. However, the biosynthesis of this auxin is poorly understood. Here, we show that in pea (Pisum sativum) seeds, 4-chloroindole-3-acetic acid is synthesized via the novel intermediate 4-chloroindole-3-pyruvic acid, which is produced from 4-chlorotryptophan by two aminotransferases, TRYPTOPHAN AMINOTRANSFERASE RELATED1 and TRYPTOPHAN AMINOTRANSFERASE RELATED2. We characterize a tar2 mutant, obtained by Targeting Induced Local Lesions in Genomes, the seeds of which contain dramatically reduced 4-chloroindole-3-acetic acid levels as they mature. We also show that the widespread auxin, indole-3-acetic acid, is synthesized by a parallel pathway in pea.
Asunto(s)
Ácidos Indolacéticos/metabolismo , Pisum sativum/metabolismo , Vías Biosintéticas , Cromatografía Líquida de Alta Presión , Regulación de la Expresión Génica de las Plantas , Ácidos Indolacéticos/química , Indoles/metabolismo , Marcaje Isotópico , Espectrometría de Masas , Mutación/genética , Pisum sativum/genética , Pisum sativum/crecimiento & desarrollo , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Estándares de Referencia , Semillas/genética , Semillas/crecimiento & desarrollo , EstereoisomerismoRESUMEN
The function of PsBRC1, the pea (Pisum sativum) homolog of the maize (Zea mays) TEOSINTE BRANCHED1 and the Arabidopsis (Arabidopsis thaliana) BRANCHED1 (AtBRC1) genes, was investigated. The pea Psbrc1 mutant displays an increased shoot-branching phenotype, is able to synthesize strigolactone (SL), and does not respond to SL application. The level of pleiotropy of the SL-deficient ramosus1 (rms1) mutant is higher than in the Psbrc1 mutant, rms1 exhibiting a relatively dwarf phenotype and more extensive branching at upper nodes. The PsBRC1 gene is mostly expressed in the axillary bud and is transcriptionally up-regulated by direct application of the synthetic SL GR24 and down-regulated by the cytokinin (CK) 6-benzylaminopurine. The results suggest that PsBRC1 may have a role in integrating SL and CK signals and that SLs act directly within the bud to regulate its outgrowth. However, the Psbrc1 mutant responds to 6-benzylaminopurine application and decapitation by increasing axillary bud length, implicating a PsBRC1-independent component of the CK response in sustained bud growth. In contrast to other SL-related mutants, the Psbrc1 mutation does not cause a decrease in the CK zeatin riboside in the xylem sap or a strong increase in RMS1 transcript levels, suggesting that the RMS2-dependent feedback is not activated in this mutant. Surprisingly, the double rms1 Psbrc1 mutant displays a strong increase in numbers of branches at cotyledonary nodes, whereas branching at upper nodes is not significantly higher than the branching in rms1. This phenotype indicates a localized regulation of branching at these nodes specific to pea.
Asunto(s)
Lactonas/metabolismo , Pisum sativum/crecimiento & desarrollo , Brotes de la Planta/crecimiento & desarrollo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Proteínas de Arabidopsis/metabolismo , Compuestos de Bencilo , Citocininas/genética , Citocininas/metabolismo , Regulación de la Expresión Génica de las Plantas , Isopenteniladenosina/análogos & derivados , Isopenteniladenosina/metabolismo , Cinetina/farmacología , Datos de Secuencia Molecular , Mutación , Pisum sativum/efectos de los fármacos , Pisum sativum/genética , Pisum sativum/metabolismo , Reguladores del Crecimiento de las Plantas/farmacología , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Brotes de la Planta/genética , Brotes de la Planta/metabolismo , Purinas , Transducción de Señal/genética , Regulación hacia Arriba , Xilema/genética , Xilema/metabolismoRESUMEN
Phytohormonal interactions are essential to regulate plant organogenesis. In response to the presence of signals from symbiotic bacteria, the Nod factors, legume roots generate a new organ: the nitrogen-fixing nodule. Analysis of mutants in the Medicago truncatula CRE1 cytokinin receptor and of the MtRR4 cytokinin primary response gene expression pattern revealed that cytokinin acts in initial cortical cell divisions and later in the transition between meristematic and differentiation zones of the mature nodule. MtCRE1 signaling is required for activation of the downstream nodulation-related transcription factors MtERN1, MtNSP2 and MtNIN, as well as to regulate expression and accumulation of PIN auxin efflux carriers. Whereas the MtCRE1 pathway is required to allow the inhibition of polar auxin transport in response to rhizobia, nodulation is still negatively regulated by the MtEIN2/SICKLE-dependent ethylene pathway in cre1 mutants. Hence, MtCRE1 signaling acts as a regulatory knob, integrating positive plant and bacterial cues to control legume nodule organogenesis.
Asunto(s)
Citocininas/metabolismo , Medicago truncatula/genética , Reguladores del Crecimiento de las Plantas/metabolismo , Proteínas de Plantas/metabolismo , Nodulación de la Raíz de la Planta/genética , Receptores de Superficie Celular/metabolismo , Simbiosis , Secuencia de Aminoácidos , Regulación de la Expresión Génica de las Plantas , Ácidos Indolacéticos/metabolismo , Medicago truncatula/metabolismo , Medicago truncatula/microbiología , Datos de Secuencia Molecular , Mutación , Proteínas de Plantas/genética , ARN de Planta/genética , Receptores de Superficie Celular/genética , Nódulos de las Raíces de las Plantas/crecimiento & desarrollo , Nódulos de las Raíces de las Plantas/microbiología , Transducción de Señal , Factores de Transcripción/metabolismo , Transformación GenéticaRESUMEN
KAI2 proteins are plant α/ß hydrolase receptors which perceive smoke-derived butenolide signals and endogenous, yet unidentified KAI2-ligands (KLs). The number of functional KAI2 receptors varies among species and KAI2 gene duplication and sub-functionalization likely plays an adaptative role by altering specificity towards different KLs. Legumes represent one of the largest families of flowering plants and contain many agronomic crops. Prior to their diversification, KAI2 underwent duplication resulting in KAI2A and KAI2B. Here we demonstrate that Pisum sativum KAI2A and KAI2B are active receptors and enzymes with divergent ligand stereoselectivity. KAI2B has a higher affinity for and hydrolyses a broader range of substrates including strigolactone-like stereoisomers. We determine the crystal structures of PsKAI2B in apo and butenolide-bound states. The biochemical, structural, and mass spectra analyses of KAI2s reveal a transient intermediate on the catalytic serine and a stable adduct on the catalytic histidine, confirming its role as a bona fide enzyme. Our work uncovers the stereoselectivity of ligand perception and catalysis by diverged KAI2 receptors and proposes adaptive sensitivity to KAR/KL and strigolactones by KAI2B.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Catálisis , Pisum sativum/genética , Pisum sativum/metabolismo , Percepción , Reguladores del Crecimiento de las Plantas/genéticaRESUMEN
Grain legumes are highly valuable plant species, as they produce seeds with high protein content. Increasing seed protein production and improving seed nutritional quality represent an agronomical challenge in order to promote plant protein consumption of a growing population. In this study, we used the genetic diversity, naturally present in Medicago truncatula, a model plant for legumes, to identify genes/loci regulating seed traits. Indeed, using sequencing data of 162 accessions from the Medicago HAPMAP collection, we performed genome-wide association study for 32 seed traits related to seed size and seed composition such as seed protein content/concentration, sulfur content/concentration. Using different GWAS and postGWAS methods, we identified 79 quantitative trait nucleotides (QTNs) as regulating seed size, 41 QTNs for seed composition related to nitrogen (i.e. storage protein) and sulfur (i.e. sulfur-containing amino acid) concentrations/contents. Furthermore, a strong positive correlation between seed size and protein content was revealed within the selected Medicago HAPMAP collection. In addition, several QTNs showed highly significant associations in different seed phenotypes for further functional validation studies, including one near an RNA-Binding Domain protein, which represents a valuable candidate as central regulator determining both seed size and composition. Finally, our findings in M. truncatula represent valuable resources to be exploitable in many legume crop species such as pea, common bean, and soybean due to its high synteny, which enable rapid transfer of these results into breeding programs and eventually help the improvement of legume grain production.