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Allium crop breeding remains severely hindered due to the lack of high-quality reference genomes. Here we report high-quality chromosome-level genome assemblies for three key Allium crops (Welsh onion, garlic and onion), which are 11.17 Gb, 15.52 Gb and 15.78 Gb in size with the highest recorded contig N50 of 507.27 Mb, 109.82 Mb and 81.66 Mb, respectively. Beyond revealing the genome evolutionary process of Allium species, our pathogen infection experiments and comparative metabolomic and genomic analyses showed that genes encoding enzymes involved in the metabolic pathway of Allium-specific flavor compounds may have evolved from an ancient uncharacterized plant defense system widely existing in many plant lineages but extensively boosted in alliums. Using in situ hybridization and spatial RNA sequencing, we obtained an overview of cell-type categorization and gene expression changes associated with spongy mesophyll cell expansion during onion bulb formation, thus indicating the functional roles of bulb formation genes.
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Allium , Allium/genética , Fitomejoramiento , Cebollas/genética , Genoma , CromosomasRESUMEN
Taraxacum kok-saghyz has been identified as one of the most promising alternative rubber crops, with laticifer cells that produce high-quality rubber. To uncover the underlying molecular mechanisms regulating natural rubber biosynthesis under MeJA induction, a reference transcriptome was constructed from nine samples of T. kok-saghyz. MeJA treatment was applied for 0 h (control), 6 h, and 24 h. A total of 7452 differentially expressed genes (DEGs) were identified in response to MeJA stress, relative to the control. Functional enrichment showed that these DEGs were primarily related to hormone signaling, defensive responses, and secondary metabolism. Combined analysis of the DEGs induced by MeJA and high-expression genes in laticifer cells further identified seven DEGs related to natural rubber biosynthesis that were upregulated in latex tissue, suggesting that these candidate genes could prove valuable in studying the mechanism of MeJA-mediated natural rubber biosynthesis. In addition, 415 MeJA-responsive DEGs were from several transcription factor families associated with drought resistance. This study helps to elucidate the mechanism of natural rubber biosynthesis in T. kok-saghyz in response to MeJA stress and identifies key candidate MeJA-induced DEGs in laticifer tissue, as well as a candidate drought-response target gene, whose knowledge will promote the breeding of T. kok-saghyz in the aspect of rubber yields and quality, and drought tolerance.
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Goma , Taraxacum , Goma/metabolismo , Taraxacum/genética , Taraxacum/metabolismo , Resistencia a la Sequía , Fitomejoramiento , Perfilación de la Expresión Génica , Transcriptoma , Regulación de la Expresión Génica de las PlantasRESUMEN
Trichomes are epidermal outgrowths on plant shoots. Their roles in protecting plants against herbivores and in the biosynthesis of specialized metabolites have long been recognized. Recently, studies are increasingly showing that trichomes also play important roles in water absorption and metal detoxication, with these roles having important implications for ecology, the environment, and agriculture. However, these two functions of trichomes have been largely overlooked and much remains unknown. In this review, we show that the trichomes of 37 plant species belonging to 14 plant families are involved in water absorption, while the trichomes of 33 species from 13 families are capable of sequestering metals within their trichomes. The ability of trichomes to absorb water results from their decreased hydrophobicity compared to the remainder of the leaf surface as well as the presence of special structures for collecting and absorbing water. In contrast, the metal detoxication function of trichomes results not only from the good connection of their basal cells to the underlying vascular tissues, but also from the presence of metal-chelating ligands and transporters within the trichomes themselves. Knowledge gaps and critical future research questions regarding these two trichome functions are highlighted. This review improves our understanding on trichomes.
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Tricomas , Agua , Agua/metabolismo , Tricomas/metabolismo , Metales/metabolismo , Hojas de la Planta/metabolismo , PlantasRESUMEN
Taraxacum kok-saghyz Rodin (Tk) is a promising alternative rubber-producing grass. However, low biomass and rubber-producing capability limit its commercial application. As a carbon source transporter in plants, sugar will eventually be exported transporters (SWEETs) have been reported to play pivotal roles in diverse physiological events in the context of carbon assimilate transport and utilization. Theoretically, SWEETs would participate in Tk growth, development and response to environmental cues with relation to the accumulation of rubber and biomass, both of which rely on the input of carbon assimilates. Here, we identified 22 TkSWEETs through homology searching of the Tk genomes and bioinformatics analyses. RNA-seq and qRT-PCR analysis revealed these TkSWEETs to have overlapping yet distinct tissue expression patterns. Two TkSWEET isofroms, TkSWEET1 and TkSWEET12 expressed substantially in the latex, the cytoplasm of rubber-producing laticifers as well as the rubber source. As revealed by the transient expression analysis using Tk mesophyll protoplasts, both TkSWEET1 and TkSWEET12 were located in the plasma membrane. Heterologous expressions of the two TkSWEETs in a yeast mutant revealed that only TkSWEET1 exhibited apparent sugar transport activities, with a preference for monosaccharides. Interestingly, TkSWEET12, the latex-predominant TkSWEET isoform, seemed to have evolved from a tandem duplication event that results in a cluster of six TkSWEET genes with the TkSWEET12 therein, suggesting its specialized roles in the laticifers.
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Látex , Taraxacum , Goma/metabolismo , Taraxacum/genética , Taraxacum/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Proteínas de Transporte de Membrana/genética , Saccharomyces cerevisiae/metabolismo , Isoformas de Proteínas/metabolismo , Azúcares/metabolismo , Regulación de la Expresión Génica de las PlantasRESUMEN
Trehalose 6-phosphate (T6P), the intermediate of trehalose biosynthesis and a signaling molecule, affects crop yield via targeting sucrose allocation and utilization. As there have been no reports of T6P signaling affecting secondary metabolism in a crop plant, the rubber tree Hevea brasiliensis serves as an ideal model in this regard. Sucrose metabolism critically influences the productivity of natural rubber, a secondary metabolite of industrial importance. Here, we report on the characterization of the T6P synthase (TPS) gene family and the T6P/SNF1-related protein kinase1 (T6P/SnRK1) signaling components in Hevea laticifers under tapping (rubber harvesting), an agronomic manipulation that itself stimulates rubber production. A total of fourteen TPS genes were identified, among which a class II TPS gene, HbTPS5, seemed to have evolved with a function specialized in laticifers. T6P and trehalose increased when the trees were tapped, this being consistent with the observed enhanced activities of TPS and T6P phosphatase (TPP) and expression of an active TPS-encoding gene, HbTPS1. On the other hand, SnRK1 activities decreased, suggesting the inhibition of elevated T6P on SnRK1. Expression profiles of the SnRK1 marker genes coincided with elevated T6P and depressed SnRK1. Interestingly, HbTPS5 expression decreased significantly with the onset of tapping, suggesting a regulatory function in the T6P pathway associated with latex production in laticifers. In brief, transcriptional, enzymatic, and metabolic evidence supports the participation of T6P/SnRK1 signaling in rubber formation, thus providing a possible avenue to increasing the yield of a valuable secondary metabolite by targeting T6P in specific cells.
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Natural rubber, an important industrial raw material with wide applications, is harvested in the form of latex (cytoplasm of rubber-producing laticifers) from Hevea brasiliensis (para rubber tree) by the way of tapping. Conspicuous stimulation on latex production is observed for the first few tappings conducted on virgin (untapped before) or resting (tapped before but no tapping for a period) rubber trees. To understand the underlying mechanisms, an integrative analysis of the latex transcriptome and proteome was conducted on virgin or resting Hevea trees for the first five tappings. A total of 505 non-redundant differentially expressed (DE) transcript-derived fragments (TDFs) were identified by silver-staining cDNA-AFLP, with 217 exhibiting patterns of upregulated, 180 downregulated and 108 irregularly-regulated. Meanwhile, 117 two dimensional gel electrophoresis DE-protein spots were isolated and subjected to mass spectrometry analysis, with 89 and 57 being successfully identified by MALDI-TOF and MALDI-TOF/TOF, respectively. About 72.5% DE-TDFs and 76.1% DE-proteins were functionally annotated and categorized. Noteworthily, most of the DE-TDFs implicated in sugar transport and metabolism as well as rubber biosynthesis were upregulated by the tapping treatment. The importance of sugar metabolism in harvesting-induced latex production was reinforced by the identification of abundant relevant DE-protein spots. About 83.8% of the randomly selected DE-TDFs were validated for expression patterns by semi-quantitative RT-PCR, and an 89.7% consistency for the 29 latex regeneration-related DE-TDFs examined by quantitative RT-PCR analysis. In brief, our results reveal extensive physiological and molecular changes in Hevea laticifers incurred by the tapping treatment, and the vast number of DE genes and proteins identified here contribute to unraveling the gene regulatory network of tapping-stimulated latex production.
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Natural rubber is an important industrial raw material and is commercially produced by rubber trees (Hevea brasiliensis). The sucrose transporter HbSUT3 plays an essential role in rubber production. Its expression in latex (cytoplasm of rubber-producing laticifers) is induced by bark treatment with Ethrel, an ethylene releaser, and the inducing effect correlates well with Ethrel-stimulated rubber yield increase. However, the mechanisms of ethylene induction on HbSUT3 expression are not known. Here, five Ethylene Response Factor (ERF) genes were identified from the cDNA library of Hevea latex by yeast one-hybrid screening with the promoter of HbSUT3 gene as bait. As revealed in a tobacco (Nicotiana tabacum) protoplast transient expression system, these HbERFs were mainly localized in the nucleus and four of them exhibited apparent transactivation activity. Of the five HbERF genes, HbERF-IXc4 was the most frequently screened in yeast one-hybrid, accounting for 65% of the ERF clones obtained. Moreover, among the five HbERFs, HbERF-IXc4 showed the strongest transactivation capacity when expressed in tobacco protoplast, the highest transcript abundance in latex and a close expressional correlation with its target gene, HbSUT3, in response to the Ethrel treatment. Taken together, our results indicate that ERFs, especially HbERF-IXc4, are critically involved in the activation of HbSUT3 expression in latex after Ethrel treatment on Hevea bark, and thus the stimulated latex yield.
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Hevea , Etilenos , Regulación de la Expresión Génica de las Plantas , Hevea/genética , Hevea/metabolismo , Látex , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , SacarosaRESUMEN
Vascular cambium proliferation in plants is crucial for the generation of vascular tissues and for mechanical strength. Phytohormones and mobile peptides are key regulators of vascular cambial activity during secondary growth; however, the signalling cross-talk underlying their coordinated action is largely unknown. Here, we reveal that BIN2-LIKE 1 (BIL1), a glycogen synthase kinase 3, integrates the PHLOEM INTERCALATED WITH XYLEM/tracheary element differentiation inhibitory factor (TDIF) RECEPTOR (PXY/TDR) module into MONOPTEROS/AUXIN RESPONSE FACTOR 5 (MP/ARF5) transcription factor action during secondary growth. BIL1-mediated phosphorylation of MP/ARF5 enhances its negative effect on vascular cambial activity, which upregulates the negative regulators of cytokinin signalling ARABIDOPSIS RESPONSE REGULATOR 7 (ARR7) and ARR15. PXY/TDR inhibits BIL1 activity, which attenuates the effect of MP/ARF5 on ARR7 and ARR15 expression, thus increasing vascular cambial activity. Together, these results suggest that BIL1 is a key mediator that links peptide signalling with auxin-cytokinin signalling for the maintenance of cambial activity.
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Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/fisiología , Arabidopsis/metabolismo , Citocininas/metabolismo , Proteínas de Unión al ADN/metabolismo , Glucógeno Sintasa Quinasa 3/fisiología , Proteínas Quinasas/metabolismo , Factores de Transcripción/metabolismo , Arabidopsis/genética , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/genética , Glucógeno Sintasa Quinasa 3/genética , Glucógeno Sintasa Quinasa 3/metabolismo , Meristema/citología , Meristema/crecimiento & desarrollo , Meristema/metabolismo , Fosforilación , Reguladores del Crecimiento de las Plantas/metabolismo , Transducción de Señal , Xilema/metabolismoRESUMEN
Spatial organization of signalling events of the phytohormone auxin is fundamental for maintaining a dynamic transition from plant stem cells to differentiated descendants. The cambium, the stem cell niche mediating wood formation, fundamentally depends on auxin signalling but its exact role and spatial organization is obscure. Here we show that, while auxin signalling levels increase in differentiating cambium descendants, a moderate level of signalling in cambial stem cells is essential for cambium activity. We identify the auxin-dependent transcription factor ARF5/MONOPTEROS to cell-autonomously restrict the number of stem cells by directly attenuating the activity of the stem cell-promoting WOX4 gene. In contrast, ARF3 and ARF4 function as cambium activators in a redundant fashion from outside of WOX4-expressing cells. Our results reveal an influence of auxin signalling on distinct cambium features by specific signalling components and allow the conceptual integration of plant stem cell systems with distinct anatomies.
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Proteínas de Arabidopsis/metabolismo , Arabidopsis/crecimiento & desarrollo , Cámbium/citología , Proteínas de Unión al ADN/metabolismo , Proteínas de Homeodominio/metabolismo , Ácidos Indolacéticos/metabolismo , Proteínas Nucleares/metabolismo , Factores de Transcripción/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/biosíntesis , Proteínas de Arabidopsis/genética , Proliferación Celular/fisiología , Perfilación de la Expresión Génica , Regulación de la Expresión Génica de las Plantas/genética , Proteínas de Homeodominio/biosíntesis , Proteínas de Homeodominio/genética , Reguladores del Crecimiento de las Plantas/metabolismo , Plantas Modificadas Genéticamente/metabolismo , Transducción de Señal , Células Madre/citología , Madera/citología , Madera/crecimiento & desarrolloRESUMEN
Metallothioneins (MTs) as reactive oxygen species (ROS) scavengers play important roles in stress response and heavy metal homeostasis. In Hevea brasiliensis (the para rubber tree that is the source of commercial natural rubber) and in other trees, the functions of MTs are not well understood. Latex exudes when the rubber tree is tapped. The flow of latex and its regeneration can be enhanced by tapping, wounding and ethylene treatment, all of which produce ROS as a by-product. Here, we show the presence of four MT genes in H. brasiliensis, comprising three Type 2 (HbMT2, -2a and -2b) and one Type 3 (HbMT3L) isoforms, representing one of the smallest MT gene families among angiosperms. The four HbMTs exhibited distinct tissue expression patterns: HbMT2 and HbMT3L mainly in leaves, HbMT2a specifically in flowers and HbMT2b in diverse tissues. The expression of HbMT2b, an isoform present in latex, decreased significantly in the latex following the stress-inducing treatments of tapping, wounding and ethephon (an ethylene generator). The expressions of the leaf-abundant isoforms, HbMT2 and -3L were up-regulated following pathogenic fungus infection and high-temperature stress, but down-regulated by low-temperature stress. These reactions were consistent with multiple defense- and hormone-responsive cis-acting elements in the HbMT promoters. Nine transcription factors were shown to implicate in the high-temperature responsiveness of HbMT2 and -3L in leaves. Overexpression of HbMT2 in Escherichia coli enhanced the bacterium's tolerance to heavy metals and ROS, consistent with its predicted role as an ROS scavenger. Taken together, our results, along with other relevant studies, suggest an important role of HbMTs in latex regeneration as well as species adaptation via the regulation of ROS homeostasis.
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Hevea/genética , Metalotioneína/genética , Familia de Multigenes , Proteínas de Plantas/genética , Secuencia de Aminoácidos , Escherichia coli/genética , Escherichia coli/metabolismo , Hevea/metabolismo , Metalotioneína/química , Metalotioneína/metabolismo , Metales Pesados/metabolismo , Microorganismos Modificados Genéticamente/genética , Microorganismos Modificados Genéticamente/metabolismo , Filogenia , Proteínas de Plantas/química , Proteínas de Plantas/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Alineación de SecuenciaRESUMEN
Sucrose-metabolizing enzymes in plant leaves have hitherto been investigated mainly in temperate plants, and rarely conducted in tandem with gene expression and sugar analysis. Here, we investigated the sugar content, gene expression, and the activity of sucrose-metabolizing enzymes in the leaves of Hevea brasiliensis, a tropical tree widely cultivated for natural rubber. Sucrose, fructose and glucose were the major sugars detected in Hevea leaves at four developmental stages (I to IV), with starch and quebrachitol as minor saccharides. Fructose and glucose contents increased until stage III, but decreased strongly at stage IV (mature leaves). On the other hand, sucrose increased continuously throughout leaf development. Activities of all sucrose-cleaving enzymes decreased markedly at maturation, consistent with transcript decline for most of their encoding genes. Activity of sucrose phosphate synthase (SPS) was low in spite of its high transcript levels at maturation. Hence, the high sucrose content in mature leaves was not due to increased sucrose-synthesizing activity, but more to the decline in sucrose cleavage. Gene expression and activities of sucrose-metabolizing enzymes in Hevea leaves showed striking differences compared with other plants. Unlike in most other species where vacuolar invertase predominates in sucrose cleavage in developing leaves, cytoplasmic invertase and sucrose synthase (cleavage direction) also featured prominently in Hevea. Whereas SPS is normally responsible for sucrose synthesis in plant leaves, sucrose synthase (synthesis direction) was comparable or higher than that of SPS in Hevea leaves. Mature Hevea leaves had an unusually high sucrose:starch ratio of about 11, the highest reported to date in plants.
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SWEET proteins play an indispensable role as a sugar efflux transporter in plant development and stress responses. The SWEET genes have previously been characterized in several plants. Here, we present a comprehensive analysis of this gene family in the rubber tree, Hevea brasiliensis. There are 36 members of the SWEET gene family in this species, making it one of the largest families in plant genomes sequenced so far. Structure and phylogeny analyses of these genes in Hevea and in other species demonstrated broad evolutionary conservation. RNA-seq analyses revealed that SWEET2, 16, and 17 might represent the main evolutionary direction of SWEET genes in plants. Our results in Hevea suggested the involvement of HbSWEET1a, 2e, 2f, and 3b in phloem loading, HbSWEET10a and 16b in laticifer sugar transport, and HbSWEET9a in nectary-specific sugar transport. Parallel studies of RNA-seq analyses extended to three other plant species (Manihot esculenta, Populus trichocarpa, and Arabidopsis thaliana) produced findings which implicated MeSWEET10a, 3a, and 15b in M. esculenta storage root development, and the involvement of PtSWEET16b and PtSWEET16d in P. trichocarpa xylem development. RT-qPCR results further revealed that HbSWEET10a, 16b, and 1a play important roles in phloem sugar transport. The results from this study provide a foundation not only for further investigation into the functionality of the SWEET gene family in Hevea, especially in its sugar transport for latex production, but also for related studies of this gene family in the plant kingdom.
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How appendages, such as plant leaves or animal limbs, develop asymmetric shapes remains a fundamental question in biology. Although ongoing research has revealed the genetic regulation of organ pattern formation, how gene activity ultimately directs organ shape remains unclear. Here, we show that leaf dorsoventral (adaxial-abaxial) polarity signals lead to mechanical heterogeneity of the cell wall, related to the methyl-esterification of cell-wall pectins in tomato and Arabidopsis. Numerical simulations predicate that mechanical heterogeneity is sufficient to produce the asymmetry seen in planar leaves. Experimental tests that alter pectin methyl-esterification, and therefore cell wall mechanical properties, support this model and lead to polar changes in gene expression, suggesting the existence of a feedback mechanism for mechanical signals in morphogenesis. Thus, mechanical heterogeneity within tissue may underlie organ shape asymmetry.
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Arabidopsis/crecimiento & desarrollo , Hojas de la Planta/crecimiento & desarrollo , Solanum lycopersicum/crecimiento & desarrollo , Arabidopsis/anatomía & histología , Arabidopsis/genética , Fenómenos Biomecánicos , Pared Celular/metabolismo , Regulación de la Expresión Génica de las Plantas , Solanum lycopersicum/efectos adversos , Solanum lycopersicum/anatomía & histología , Solanum lycopersicum/genética , Hojas de la Planta/anatomía & histología , Hojas de la Planta/genéticaRESUMEN
Calcium-dependent protein kinases (CDPKs or CPKs) play important roles in various physiological processes of plants, including growth and development, stress responses and hormone signaling. Although the CDPK gene family has been characterized in several model plants, little is known about this gene family in Hevea brasiliensis (the Para rubber tree). Here, we characterize the entire H. brasiliensis CDPK and CDPK-related kinase (CRK) gene families comprising 30 CDPK genes (HbCPK1 to 30) and nine CRK genes (HbCRK1 to 9). Structure and phylogeny analyses of these CDPK and CRK genes demonstrate evolutionary conservation in these gene families across H. brasiliensis and other plant species. The expression of HbCPK and HbCRK genes was investigated via Solexa sequencing in a range of experimental conditions (different tissues, phases of leaf development, ethylene treatment, and various abiotic stresses). The results suggest that HbCPK and HbCRK genes are important components in growth, development, and stress responses of H. brasiliensis. Parallel studies on the CDPK and CRK gene families were also extended to five other plant species (Arabidopsis thaliana, Oryza sativa, Populus trichocarpa, Manihot esculenta, and Ricinus communis). The CDPK and CRK genes from different plant species that exhibit similar expression patterns tend to cluster together, suggesting a coevolution of gene structure and expression behavior in higher plants. The results serve as a foundation to further functional studies of these gene families in H. brasiliensis as well as in the whole plant kingdom.
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Spatial organization is fundamental for the performance of living organisms and is reflected in a distinct distribution of structures and molecules down to the subcellular level. In particular, eukaryotic cells harbor a vast range of possibilities for distributing organelles, the cytoskeleton or the extracellular matrix in an active and highly regulated manner. An asymmetric or polar distribution is rather the rule than the exception and often reflects a particular position or orientation of a cell within a multicellular body. Here, we highlight recent insights into the regulation of cell polarity in plants and reveal the interactive nature of underlying molecular processes.
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Polaridad Celular/fisiología , Células Vegetales/fisiología , Fenómenos Fisiológicos de las PlantasRESUMEN
The Para rubber tree (Hevea brasiliensis) is an economically important tropical tree species that produces natural rubber, an essential industrial raw material. Here we present a high-quality genome assembly of this species (1.37â Gb, scaffold N50â =â 1.28 Mb) that covers 93.8% of the genome (1.47â Gb) and harbours 43,792 predicted protein-coding genes. A striking expansion of the REF/SRPP (rubber elongation factor/small rubber particle protein) gene family and its divergence into several laticifer-specific isoforms seem crucial for rubber biosynthesis. The REF/SRPP family has isoforms with sizes similar to or larger than SRPP1 (204 amino acids) in 17 other plants examined, but no isoforms with similar sizes to REF1 (138 amino acids), the predominant molecular variant. A pivotal point in Hevea evolution was the emergence of REF1, which is located on the surface of large rubber particles that account for 93% of rubber in the latex (despite constituting only 6% of total rubber particles, large and small). The stringent control of ethylene synthesis under active ethylene signalling and response in laticifers resolves a longstanding mystery of ethylene stimulation in rubber production. Our study, which includes the re-sequencing of five other Hevea cultivars and extensive RNA-seq data, provides a valuable resource for functional genomics and tools for breeding elite Hevea cultivars.
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Etilenos/farmacología , Genoma de Planta , Hevea/genética , Hevea/metabolismo , Reguladores del Crecimiento de las Plantas/farmacología , Goma/metabolismo , Adaptación BiológicaRESUMEN
In rubber tree, latex regeneration is one of the decisive factors influencing the rubber yield, although its molecular regulation is not well known. Quantitative real-time PCR (qPCR) is a popular and powerful tool used to understand the molecular mechanisms of latex regeneration. However, the suitable reference genes required for qPCR are not available to investigate the expressions of target genes during latex regeneration. In this study, 20 candidate reference genes were selected and evaluated for their expression stability across the samples during the process of latex regeneration. All reference genes showed a relatively wide range of the threshold cycle values, and their stability was validated by four different algorithms (comparative delta Ct method, Bestkeeper, NormFinder and GeNorm). Three softwares (comparative delta Ct method, NormFinder and GeNorm) exported similar results that identify UBC4, ADF, UBC2a, eIF2 and ADF4 as the top five suitable references, and 18S as the least suitable one. The application of the screened references would improve accuracy and reliability of gene expression analysis in latex regeneration experiments.
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Genes de Plantas/genética , Hevea/genética , Látex/metabolismo , Regeneración/genética , Algoritmos , Perfilación de la Expresión Génica/métodos , Regulación de la Expresión Génica de las Plantas/genética , Hevea/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa/métodos , Estándares de Referencia , Reproducibilidad de los ResultadosRESUMEN
In Hevea brasiliensis, an alkaline/neutral invertase (A/N-Inv) is responsible for sucrose catabolism in latex (essentially the cytoplasm of rubber-producing laticifers, the source of natural rubber) and implicated in rubber yield. However, neither the gene encoding this enzyme nor its molecular and biochemical properties have been well documented. Three Hevea A/N-Inv genes, namely HbNIN1, 2 and 3, were first cloned and characterized in planta and in Escherichia coli. Cellular localizations of HbNIN2 mRNA and protein were probed. From latex, active A/N-Inv proteins were purified, identified, and explored for enzymatic properties. HbNIN2 was identified as the major A/N-Inv gene functioning in latex based on its functionality in E. coli, its latex-predominant expression, the conspicuous localization of its mRNA and protein in the laticifers, and its expressional correlation with rubber yield. An active A/N-Inv protein was partially purified from latex, and determined as HbNIN2. The enhancement of HbNIN2 enzymatic activity by pyridoxal is peculiar to A/N-Invs in other plants. We conclude that HbNIN2, a cytosolic A/N-Inv, is responsible for sucrose catabolism in rubber laticifers. The results contribute to the studies of sucrose catabolism in plants as a whole and natural rubber synthesis in particular.
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Hevea/enzimología , Sacarosa/metabolismo , beta-Fructofuranosidasa/metabolismo , Secuencia de Aminoácidos , Citosol/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Expresión Génica , Hevea/citología , Hevea/genética , Látex/metabolismo , Datos de Secuencia Molecular , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Tallos de la Planta/citología , Tallos de la Planta/enzimología , Tallos de la Planta/genética , Alineación de Secuencia , beta-Fructofuranosidasa/genéticaRESUMEN
In plants, ethanolic fermentation occurs not only under anaerobic conditions but also under aerobic conditions, and involves carbohydrate and energy metabolism. Pyruvate decarboxylase (PDC) is the first and the key enzyme of ethanolic fermentation, which branches off the main glycolytic pathway at pyruvate. Here, four PDC genes were isolated and identified in a rubber tree, and the protein sequences they encode are very similar. The expression patterns of HbPDC4 correlated well with tapping-simulated rubber productivity in virgin rubber trees, indicating it plays an important role in regulating glycometabolism during latex regeneration. HbPDC1, HbPDC2 and HbPDC3 had striking expressional responses in leaves and bark to drought, low temperature and high temperature stresses, indicating that the HbPDC genes are involve in self-protection and defense in response to various abiotic and biotic stresses during rubber tree growth and development. To understand ethanolic fermentation in rubber trees, it will be necessary to perform an in-depth study of the regulatory pathways controlling the HbPDCs in the future.