RESUMEN
BACKGROUND: MADS-box genes are key regulators of plant reproductive development and members of most lineages of this gene family have been extensively studied. However, the function and diversification of the ancient TM8 lineage remains elusive to date. The available data suggest a possible function in flower development in tomato and fast evolution through numerous gene loss events in flowering plants. RESULTS: We show the broad conservation of TM8 within angiosperms and find that in contrast to other MADS-box gene lineages, no gene duplicates have been retained after major whole genome duplication events. Through knock-down of NbTM8 by virus induced gene silencing in Nicotiana benthamiana, we show that NbTM8 represses miR172 together with another MADS-box gene, SHORT VEGETATIVE PHASE (NbSVP). In the closely related species Petunia hybrida, PhTM8 is not expressed under the conditions we investigated and consistent with this, a knock-out mutant did not show a phenotype. Finally, we generated transgenic tomato plants in which TM8 was silenced or ectopically expressed, but these plants did not display a clear phenotype. Therefore, no clear function could be confirmed for Solanum lycopersium. CONCLUSIONS: While the presence of TM8 is generally conserved, it remains difficult to propose a general function in angiosperms. Based on all the available data to date, supplemented with our own results, TM8 function seems to have diversified quickly throughout angiosperms and acts as repressor of miR172 in Nicotiana benthamiana, together with NbSVP.
Asunto(s)
Proteínas de Dominio MADS/genética , Magnoliopsida/crecimiento & desarrollo , Nicotiana/crecimiento & desarrollo , Proteínas de Plantas/genética , Evolución Biológica , Secuencia Conservada/genética , Secuencia Conservada/fisiología , Genes de Plantas/genética , Genes de Plantas/fisiología , Proteínas de Dominio MADS/fisiología , Magnoliopsida/genética , Petunia/genética , Petunia/fisiología , Filogenia , Proteínas de Plantas/fisiología , Nicotiana/genética , TranscriptomaRESUMEN
Auxin steers numerous physiological processes in plants, making the tight control of its endogenous levels and spatiotemporal distribution a necessity. This regulation is achieved by different mechanisms, including auxin biosynthesis, metabolic conversions, degradation, and transport. Here, we introduce cis-cinnamic acid (c-CA) as a novel and unique addition to a small group of endogenous molecules affecting in planta auxin concentrations. c-CA is the photo-isomerization product of the phenylpropanoid pathway intermediate trans-CA (t-CA). When grown on c-CA-containing medium, an evolutionary diverse set of plant species were shown to exhibit phenotypes characteristic for high auxin levels, including inhibition of primary root growth, induction of root hairs, and promotion of adventitious and lateral rooting. By molecular docking and receptor binding assays, we showed that c-CA itself is neither an auxin nor an anti-auxin, and auxin profiling data revealed that c-CA does not significantly interfere with auxin biosynthesis. Single cell-based auxin accumulation assays showed that c-CA, and not t-CA, is a potent inhibitor of auxin efflux. Auxin signaling reporters detected changes in spatiotemporal distribution of the auxin response along the root of c-CA-treated plants, and long-distance auxin transport assays showed no inhibition of rootward auxin transport. Overall, these results suggest that the phenotypes of c-CA-treated plants are the consequence of a local change in auxin accumulation, induced by the inhibition of auxin efflux. This work reveals a novel mechanism how plants may regulate auxin levels and adds a novel, naturally occurring molecule to the chemical toolbox for the studies of auxin homeostasis.
Asunto(s)
Cinamatos/metabolismo , Ácidos Indolacéticos/metabolismo , Raíces de Plantas/efectos de los fármacos , Raíces de Plantas/crecimiento & desarrollo , Arabidopsis/efectos de los fármacos , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Bryopsida/efectos de los fármacos , Bryopsida/crecimiento & desarrollo , Cinamatos/química , Cinamatos/farmacología , Ciclina B/genética , Ciclina B/metabolismo , Regulación de la Expresión Génica de las Plantas , Isomerismo , Raíces de Plantas/metabolismo , Plantas Modificadas Genéticamente , Proteínas Qa-SNARE/genética , Proteínas Qa-SNARE/metabolismo , Selaginellaceae/efectos de los fármacos , Selaginellaceae/crecimiento & desarrollo , Transducción de SeñalRESUMEN
Phosphatidylinositol (PtdIns) is a structural phospholipid that can be phosphorylated into various lipid signaling molecules, designated polyphosphoinositides (PPIs). The reversible phosphorylation of PPIs on the 3, 4, or 5 position of inositol is performed by a set of organelle-specific kinases and phosphatases, and the characteristic head groups make these molecules ideal for regulating biological processes in time and space. In yeast and mammals, PtdIns3P and PtdIns(3,5)P2 play crucial roles in trafficking toward the lytic compartments, whereas the role in plants is not yet fully understood. Here we identified the role of a land plant-specific subgroup of PPI phosphatases, the suppressor of actin 2 (SAC2) to SAC5, during vacuolar trafficking and morphogenesis in Arabidopsis thaliana. SAC2-SAC5 localize to the tonoplast along with PtdIns3P, the presumable product of their activity. In SAC gain- and loss-of-function mutants, the levels of PtdIns monophosphates and bisphosphates were changed, with opposite effects on the morphology of storage and lytic vacuoles, and the trafficking toward the vacuoles was defective. Moreover, multiple sac knockout mutants had an increased number of smaller storage and lytic vacuoles, whereas extralarge vacuoles were observed in the overexpression lines, correlating with various growth and developmental defects. The fragmented vacuolar phenotype of sac mutants could be mimicked by treating wild-type seedlings with PtdIns(3,5)P2, corroborating that this PPI is important for vacuole morphology. Taken together, these results provide evidence that PPIs, together with their metabolic enzymes SAC2-SAC5, are crucial for vacuolar trafficking and for vacuolar morphology and function in plants.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimología , Fosfoproteínas Fosfatasas/metabolismo , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/genética , Cartilla de ADN/genética , Evolución Molecular , Microscopía Electrónica de Transmisión , Fenotipo , Fosfatos de Fosfatidilinositol/metabolismo , Fosfoproteínas Fosfatasas/genética , Plantas Modificadas Genéticamente , Reacción en Cadena en Tiempo Real de la Polimerasa , Vacuolas/metabolismo , Vacuolas/fisiología , Vacuolas/ultraestructuraRESUMEN
Plant development mediated by the phytohormone auxin depends on tightly controlled cellular auxin levels at its target tissue that are largely established by intercellular and intracellular auxin transport mediated by PIN auxin transporters. Among the eight members of the Arabidopsis PIN family, PIN6 is the least characterized candidate. In this study we generated functional, fluorescent protein-tagged PIN6 proteins and performed comprehensive analysis of their subcellular localization and also performed a detailed functional characterization of PIN6 and its developmental roles. The localization study of PIN6 revealed a dual localization at the plasma membrane (PM) and endoplasmic reticulum (ER). Transport and metabolic profiling assays in cultured cells and Arabidopsis strongly suggest that PIN6 mediates both auxin transport across the PM and intracellular auxin homeostasis, including the regulation of free auxin and auxin conjugates levels. As evidenced by the loss- and gain-of-function analysis, the complex function of PIN6 in auxin transport and homeostasis is required for auxin distribution during lateral and adventitious root organogenesis and for progression of these developmental processes. These results illustrate a unique position of PIN6 within the family of PIN auxin transporters and further add complexity to the developmentally crucial process of auxin transport.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Membrana Celular/metabolismo , Retículo Endoplásmico/metabolismo , Ácidos Indolacéticos/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Arabidopsis/genética , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/genética , Evolución Molecular , Homeostasis , Proteínas de Transporte de Membrana/genética , Filogenia , Raíces de Plantas/crecimiento & desarrollo , Raíces de Plantas/metabolismo , Plantas Modificadas GenéticamenteRESUMEN
In order to establish a reference for analysis of the function of auxin and the auxin biosynthesis regulators SHORT INTERNODE/STYLISH (SHI/STY) during Physcomitrella patens reproductive development, we have described male (antheridial) and female(archegonial) development in detail, including temporal and positional information of organ initiation. This has allowed us to define discrete stages of organ morphogenesis and to show that reproductive organ development in P. patens is highly organized and that organ phyllotaxis differs between vegetative and reproductive development. Using the PpSHI1 and PpSHI2 reporter and knockout lines, the auxin reporters GmGH3(pro):GUS and PpPINA(pro):GFP-GUS, and the auxin-conjugating transgene PpSHI2(pro):IAAL, we could show that the PpSHI genes, and by inference also auxin, play important roles for reproductive organ development in moss. The PpSHI genes are required for the apical opening of the reproductive organs, the final differentiation of the egg cell, and the progression of canal cells into a cell death program. The apical cells of the archegonium, the canal cells, and the egg cell are also sites of auxin responsiveness and are affected by reduced levels of active auxin, suggesting that auxin mediates PpSHI function in the reproductive organs.
Asunto(s)
Bryopsida/crecimiento & desarrollo , Bryopsida/genética , Regulación de la Expresión Génica de las Plantas , Ácidos Indolacéticos/metabolismo , Flores/genética , Técnicas de Inactivación de Genes , Genes Reporteros , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Brotes de la Planta/genética , Plantas Modificadas GenéticamenteRESUMEN
Comparative genome biology has unveiled the polyploid origin of all angiosperms and the role of recurrent polyploidization in the amplification of gene families and the structuring of genomes. Which species share certain ancient polyploidy events, and which do not, is ill defined because of the limited number of sequenced genomes and transcriptomes and their uneven phylogenetic distribution. Previously, it has been suggested that most, but probably not all, of the eudicots have shared an ancient hexaploidy event, referred to as the gamma triplication. In this study, detailed phylogenies of subfamilies of MADS-box genes suggest that the gamma triplication has occurred before the divergence of Gunnerales but after the divergence of Buxales and Trochodendrales. Large-scale phylogenetic and K(S)-based approaches on the inflorescence transcriptomes of Gunnera manicata (Gunnerales) and Pachysandra terminalis (Buxales) provide further support for this placement, enabling us to position the gamma triplication in the stem lineage of the core eudicots. This triplication likely initiated the functional diversification of key regulators of reproductive development in the core eudicots, comprising 75% of flowering plants. Although it is possible that the gamma event triggered early core eudicot diversification, our dating estimates suggest that the event occurred early in the stem lineage, well before the rapid speciation of the earliest core eudicot lineages. The evolutionary significance of this paleopolyploidy event may thus rather lie in establishing a species lineage that was resilient to extinction, but with the genomic potential for later diversification. We consider that the traits generated from this potential characterize extant core eudicots both chemically and morphologically.
Asunto(s)
Evolución Molecular , Genoma de Planta/genética , Magnoliopsida/genética , Filogenia , Poliploidía , Secuencia de Bases , Clonación Molecular , Cartilla de ADN/genética , Perfilación de la Expresión Génica , Inflorescencia/genética , Inflorescencia/metabolismo , Funciones de Verosimilitud , Proteínas de Dominio MADS/genética , Modelos Genéticos , Datos de Secuencia Molecular , Sistemas de Lectura Abierta/genética , Alineación de Secuencia , Análisis de Secuencia de ADNRESUMEN
⢠An important evolutionary mechanism shaping the biodiversity of flowering plants is the transfer of function from one plant organ to another. To investigate whether and how transference of function is associated with the remodeling of the floral organ identity program we studied Davidia involucrata, a species with conspicuous, petaloid bracts subtending a contracted inflorescence with reduced flowers. ⢠A detailed ontogeny enabled the interpretation of expression patterns of B-, C- and E-class homeotic MADS-box genes using qRT-PCR and in situ hybridization techniques. We investigated protein-protein interactions using yeast two-hybrid assays. ⢠Although loss of organs does not appear to have affected organ identity in the retained organs of the reduced flowers of D. involucrata, the bracts express the B-class TM6 (Tomato MADS box gene 6) and GLOBOSA homologs, but not DEFICIENS, and the C-class AGAMOUS homolog, representing a subset of genes also involved in stamen identity. ⢠Our results may illustrate how petal identity can be partially transferred outside the flower by expressing a subset of stamen identity genes. This adds to the molecular mechanisms explaining the diversity of plant reproductive morphology.
Asunto(s)
Inflorescencia/anatomía & histología , Nyssaceae/anatomía & histología , Árboles/anatomía & histología , Clorofila/metabolismo , Clonación Molecular , ADN Complementario/genética , Regulación del Desarrollo de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Genes de Plantas/genética , Hibridación in Situ , Inflorescencia/citología , Inflorescencia/crecimiento & desarrollo , Inflorescencia/ultraestructura , Proteínas de Dominio MADS/genética , Proteínas de Dominio MADS/metabolismo , Datos de Secuencia Molecular , Nyssaceae/citología , Nyssaceae/genética , Nyssaceae/ultraestructura , Especificidad de Órganos , Epidermis de la Planta/citología , Epidermis de la Planta/ultraestructura , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Unión Proteica , Reproducción/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Árboles/citología , Árboles/genética , Árboles/ultraestructura , Técnicas del Sistema de Dos HíbridosRESUMEN
As wheat (Triticum aestivum) is an important staple food across the world, preservation of stable yields and increased productivity are major objectives in breeding programs. Drought is a global concern because its adverse impact is expected to be amplified in the future due to the current climate change. Here, we analyzed the effects of edaphic, environmental, and host factors on the wheat root microbiomes collected in soils from six regions in Belgium. Amplicon sequencing analysis of unplanted soil and wheat root endosphere samples indicated that the microbial community variations can be significantly explained by soil pH, microbial biomass, wheat genotype, and soil sodium and iron levels. Under drought stress, the biodiversity in the soil decreased significantly, but increased in the root endosphere community, where specific soil parameters seemingly determine the enrichment of bacterial groups. Indeed, we identified a cluster of drought-enriched bacteria that significantly correlated with soil compositions. Interestingly, integration of a functional analysis further revealed a strong correlation between the same cluster of bacteria and ß-glucosidase and osmoprotectant proteins, two functions known to be involved in coping with drought stress. By means of this in silico analysis, we identified amplicon sequence variants (ASVs) that could potentially protect the plant from drought stress and validated them in planta. Yet, ASVs based on 16S rRNA sequencing data did not completely distinguish individual isolates because of their intrinsic short sequences. Our findings support the efforts to maintain stable crop yields under drought conditions through implementation of root microbiome analyses.
RESUMEN
Basal asterid families, and to a lesser extent the asterids as a whole, are characterized by a high variation in petal and stamen morphology. Moreover, the stamen number, the adnation of stamens to petals, and the degree of sympetaly vary considerably among basal asterid taxa. The B group genes, members of the APETALA3 (AP3) and PISTILLATA (PI) gene lineages, have been shown to specify petal and stamen identities in several core eudicot species. Duplicate genes in these lineages have been shown in some cases to have diversified in their function; for instance in Petunia, a PI paralog is required for the fusion of stamens to the corolla tube, illustrating that such genes belonging to this lineage are not just involved in specifying the identity of the stamens and petals but can also specify novel floral morphologies. This motivated us to study the duplication history of class B genes throughout asterid lineages, which comprise approximately one-third of all flowering plants. The evolutionary history of the PI gene subfamily indicates that the two genes in Petunia result from an ancient duplication event, coinciding with the origin of core asterids. A second duplication event occurred before the speciation of basal asterid Ericales families. These and other duplications in the PI lineage are not correlated with duplications in the AP3 lineage. To understand the molecular evolution of the Ericales PI genes after duplication, we have described their expression patterns using reverse transcription polymerase chain reaction and in situ hybridization, reconstructed how selection shaped their protein sequences and tested their protein interaction specificity with other class B proteins. We find that after duplication, PI paralogs have acquired multiple different expression patterns and negative selective pressure on their codons is relaxed, whereas substitutions in sites putatively involved in protein-protein interactions show positive selection, allowing for a change in the interaction behavior of the PI paralogs after duplication. Together, these observations suggest that the asterids have preferentially recruited PI duplicate genes to diverse and potentially novel roles in asterid flower development.
Asunto(s)
Flores/genética , Magnoliopsida/genética , Proteínas de Plantas/genética , Evolución Molecular , Duplicación de Gen , Especiación Genética , Hibridación in Situ , Filogenia , Proteínas de Plantas/clasificación , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Técnicas del Sistema de Dos HíbridosRESUMEN
BACKGROUND: Because of their known role as transcriptional regulators of key plant developmental processes, the diversification of MADS-box gene function is thought to be a major driving force in the developmental evolution of plants. Yet the function of some MADS-box gene subfamilies has remained elusive thus far. One such lineage, AGL6, has now been functionally characterized in three angiosperm species, but a phylogenetic framework for comparison of AGL6 gene function is currently missing. RESULTS: Based on phylogenetic analyses of newly isolated and EST-based sequences, we describe the duplication history of the AGL6 subfamily in angiosperms. Our analyses provide support for four ancient duplications in the evolution of the AGL6 lineage: one at the base of core eudicots resulting in euAGL6 and AGL6-like gene clades, one during basal angiosperm diversification and two in monocot evolution. To investigate whether the spatial domains in which AGL6 genes function have diverged after duplication, we use quantitative Real Time PCR. We show that the core eudicot AGL6-like clade acquired expression in vegetative tissues, while its paralog euAGL6 remains predominantly confined to reproductive tissues. CONCLUSIONS: These and previous data lead us to propose that the AGL6 lineage in core eudicots, in addition to functions related to the expression in reproductive structures, may have acquired a function in developmental transitions of vegetative shoots.
Asunto(s)
Regulación de la Expresión Génica de las Plantas , Magnoliopsida/genética , Magnoliopsida/metabolismo , Clonación Molecular , Duplicación de Gen , Proteínas de Dominio MADS/genética , Proteínas de Dominio MADS/metabolismo , Magnoliopsida/clasificación , Filogenia , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
BACKGROUND: When maize (Zea mays L.) is grown in the Northern hemisphere, its development is heavily arrested by chilling temperatures, especially at the juvenile phase. As some endophytes are beneficial for plants under stress conditions, we analyzed the impact of chilling temperatures on the root microbiome and examined whether microbiome-based analysis might help to identify bacterial strains that could promote growth under these temperatures. RESULTS: We investigated how the maize root microbiome composition changed by means of 16S rRNA gene amplicon sequencing when maize was grown at chilling temperatures in comparison to ambient temperatures by repeatedly cultivating maize in field soil. We identified 12 abundant and enriched bacterial families that colonize maize roots, consisting of bacteria recruited from the soil, whereas seed-derived endophytes were lowly represented. Chilling temperatures modified the root microbiome composition only slightly, but significantly. An enrichment of several chilling-responsive families was detected, of which the Comamonadaceae and the Pseudomonadaceae were the most abundant in the root endosphere of maize grown under chilling conditions, whereas only three were strongly depleted, among which the Streptomycetaceae. Additionally, a collection of bacterial strains isolated from maize roots was established and a selection was screened for growth-promoting effects on juvenile maize grown under chilling temperatures. Two promising strains that promoted maize growth under chilling conditions were identified that belonged to the root endophytic bacterial families, from which the relative abundance remained unchanged by variations in the growth temperature. CONCLUSIONS: Our analyses indicate that chilling temperatures affect the bacterial community composition within the maize root endosphere. We further identified two bacterial strains that boost maize growth under chilling conditions. Their identity revealed that analyzing the chilling-responsive families did not help for their identification. As both strains belong to root endosphere enriched families, visualizing and comparing the bacterial diversity in these communities might still help to identify new PGPR strains. Additionally, a strain does not necessarely need to belong to a high abundant family in the root endosphere to provoke a growth-promoting effect in chilling conditions. Video abstract.
Asunto(s)
Bacterias/clasificación , Frío , Raíces de Plantas/microbiología , Zea mays/crecimiento & desarrollo , Bacterias/aislamiento & purificación , Endófitos/clasificación , Microbiota , Filogenia , ARN Ribosómico 16S/genética , Rizosfera , Semillas/microbiología , Microbiología del Suelo , Zea mays/microbiologíaRESUMEN
PIN-FORMED (PIN) transporters mediate directional, intercellular movement of the phytohormone auxin in land plants. To elucidate the evolutionary origins of this developmentally crucial mechanism, we analysed the single PIN homologue of a simple green alga Klebsormidium flaccidum. KfPIN functions as a plasma membrane-localized auxin exporter in land plants and heterologous models. While its role in algae remains unclear, PIN-driven auxin export is probably an ancient and conserved trait within streptophytes.
Asunto(s)
Chlorophyta/metabolismo , Evolución Molecular , Ácidos Indolacéticos/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Transporte Biológico , Chlorophyta/genética , Proteínas de Transporte de Membrana/genética , Plantas/genética , Plantas/metabolismoRESUMEN
Here we discuss the advantages of the majority of this versatile and diverse group of microorganisms for plant health and growth as demonstrated by their contribution to disease-suppressive soils, their antifungal and antibacterial activities, their ability to produce volatile compounds and their capacity to enhance plant biomass. Although much is still to be discovered about the colonization strategies and molecular interactions between plant roots and these microorganisms, they are destined to become important players in the field of plant growth-promoting rhizobacteria for agriculture.
Asunto(s)
Desarrollo de la Planta/fisiología , Raíces de Plantas/microbiología , Plantas/microbiología , Streptomyces/crecimiento & desarrollo , Simbiosis/fisiología , Agricultura , Agentes de Control Biológico , Suelo , Microbiología del SueloRESUMEN
The emergence and radiation of multicellular land plants was driven by crucial innovations to their body plans. The directional transport of the phytohormone auxin represents a key, plant-specific mechanism for polarization and patterning in complex seed plants. Here, we show that already in the early diverging land plant lineage, as exemplified by the moss Physcomitrella patens, auxin transport by PIN transporters is operational and diversified into ER-localized and plasma membrane-localized PIN proteins. Gain-of-function and loss-of-function analyses revealed that PIN-dependent intercellular auxin transport in Physcomitrella mediates crucial developmental transitions in tip-growing filaments and waves of polarization and differentiation in leaf-like structures. Plasma membrane PIN proteins localize in a polar manner to the tips of moss filaments, revealing an unexpected relation between polarization mechanisms in moss tip-growing cells and multicellular tissues of seed plants. Our results trace the origins of polarization and auxin-mediated patterning mechanisms and highlight the crucial role of polarized auxin transport during the evolution of multicellular land plants.
RESUMEN
Polarized auxin transport is crucial for many developmental processes in flowering plants and requires the PIN-FORMED (PIN) family of auxin efflux carriers. However, the impact of polar auxin transport and PIN proteins on the development of non-seed plant species and green algal lineages is largely unknown. Using recently available sequence information from streptophyte algae and other non-seed plant species, we have constructed a preliminary phylogenetic framework and present several hypotheses for PIN protein evolution. We postulate that PIN proteins originated in streptophyte algae at the endoplasmic reticulum (ER) and that plasma membrane localization was acquired during land plant evolution. We also suggest that PIN proteins are evolutionarily distinct from another family of auxin transporters at the ER, the PIN-LIKES (PILS) proteins.
Asunto(s)
Proteínas Algáceas/genética , Evolución Molecular , Ácidos Indolacéticos/metabolismo , Proteínas de Transporte de Membrana/genética , Proteínas de Plantas/genética , Streptophyta/genética , Transporte Biológico , Membrana Celular/metabolismo , Retículo Endoplásmico/metabolismo , Filogenia , Análisis de Secuencia de ProteínaRESUMEN
A consistent challenge in studying the evolution of developmental processes has been the problem of explicitly assessing the function of developmental control genes in diverse species. In recent years, virus-induced gene silencing (VIGS) has proved to be remarkably adaptable and efficient in silencing developmental control genes in species across the angiosperms. Here we describe proven protocols for Nicotiana benthamiana and Papaver somniferum, representing a core and basal eudicot species.
Asunto(s)
Nicotiana/genética , Papaver/genética , Virus de Plantas/genética , Interferencia de ARN , Agrobacterium tumefaciens/virología , Regulación de la Expresión Génica de las Plantas , Técnicas de Silenciamiento del Gen/métodos , Vectores Genéticos , Crecimiento y Desarrollo/genética , Papaver/crecimiento & desarrollo , Papaver/virología , Hojas de la Planta/virología , Nicotiana/crecimiento & desarrollo , Nicotiana/virología , Transformación GenéticaRESUMEN
Evolution of class B genes through gene duplication has been proposed as an evolutionary mechanism that contributed to the enormous floral diversity. Frameshift mutations are a likely mechanism to explain the divergent C-terminal sequences of MIKC gene subfamilies. So far, the inferences for frameshifts and selective pressures on the C-terminal domain are made for old duplications for which the exact selective pressures are obscured by evolutionary time. This motivated us to study an example of a recent duplication, which allows us to consider in more detail the selective pressures that are involved after duplication. We find that after duplication and frameshift of Impatiens class B genes, the individual codons show no evidence for adaptive selection. It is rather the length of the C-terminal domain that either is strictly conserved or varies strongly. This suggests a role for the length of the C-terminal domain in the retention of duplicated genes.
Asunto(s)
Impatiens/genética , Proteínas de Dominio MADS/genética , Proteínas de Plantas/genética , Selección Genética , Secuencia de Aminoácidos , Secuencia de Bases , Mutación del Sistema de Lectura , Duplicación de Gen , Proteínas de Dominio MADS/clasificación , Datos de Secuencia Molecular , Filogenia , Proteínas de Plantas/clasificación , Alineación de SecuenciaRESUMEN
APETALA3 (AP3)/DEFICIENS (DEF) is a MADS-box transcription factor that is involved in establishing the identity of petal and stamen floral organs. The AP3/DEF gene lineage has been extensively examined throughout the angiosperms in order to better understand its role in floral diversity and evolution. As a result, a large number of cloned AP3/DEF orthologues are available, which can be used for the design of taxon specific primers for phylogeny reconstruction of close relatives of the group of interest. Following this reasoning, we investigated the phylogenetic utility of the two AP3/DEF paralogues (ImpDEF1 and ImpDEF2) that were recently identified in the genus Impatiens (Balsaminaceae). K-domain introns 4 and 5 of both AP3/DEF duplicates were amplified and sequenced for 59 Impatiens species. Phylogenetic analyses of the separated and combined ImpDEF1 and ImpDEF2 data sets result in highly congruent topologies with the previously obtained chloroplast atpB-rbcL data set. Combination of chloroplast and nuclear matrices results in a well-supported evolutionary hypothesis of Impatiens. Our results show that introns 4 and 5 in AP3/DEF-like genes are a valuable source of characters for phylogenetic studies at the infrageneric level.
Asunto(s)
Proteína DEFICIENS/genética , Evolución Molecular , Impatiens/genética , Filogenia , Secuencia de Aminoácidos , Secuencia de Bases , Teorema de Bayes , Cartilla de ADN , ADN de Cloroplastos/genética , Dosificación de Gen/genética , Variación Genética , Intrones/genética , Modelos Genéticos , Datos de Secuencia Molecular , Análisis de Secuencia de ADN , Especificidad de la EspecieRESUMEN
Impatiens and Marcgravia have striking morphological innovations associated with the flowers. One of the sepals in Impatiens is spurred and petaloid, while in Marcgravia the petals are fused into a cap and nectary cups are associated with the inflorescence. Balsaminaceae (Impatiens) and Marcgraviaceae have surprisingly been shown to be closely related, since both belong to the balsaminoid clade of Ericales (basal asterids). However, several thorough morphological studies thus far have not revealed shared derived characters (synapomorphies) that support a close relationship between these families. In the balsaminoid clade, transitions from entirely green flowers to flowers with heterotopic petaloid organs can be observed. The primary role of class B genes in core eudicots is to specify the identity of petal and stamen floral organs. E-class genes, of which SEP3 is a representative, have been identified as redundant mediators that confer transcriptional activation potential on protein complexes that specify organ identity. Given the conserved function of organ-identity MADS-box genes in model plants, but the rapid molecular evolution in angiosperms, it remains controversial whether these genes have been involved in shaping floral diversity. We have identified a SEP3-like gene and a total of five class B genes from Impatiens hawkeri and Marcgravia umbellata and report their quantitative expression in the floral organs. In Impatiens, two AP3/DEF-like genes were identified with strongly divergent C-terminal domains, one truncated and one unusually long. Both genes show a gradual decrease in expression towards the outer perianth organs, but no GLO-like gene expression is observed in the petaloid sepal. Remarkably, SEP3-like gene expression in the Impatiens perianth is absent from the green sepals but present in the petaloid sepal and in the petals. Dimeric protein interactions of the cloned Impatiens genes were studied in yeast and by using gel retardation. In Marcgravia, strong overlapping class B gene expression is limited to the stamens, but a SEP3-like gene is strongly expressed in the Marcgravia nectary, indicating that both Impatiens and Marcgravia show heterotopic expression of a SEP3-like gene. We discuss several candidate mechanisms for heterotopic petaloidy involving modified gene expression and protein interaction of SEP3-like and class B genes.