RESUMO
C2 photosynthesis is a photosynthetic pathway in which photorespiratory CO2 release and refixation are enhanced in leaf bundle sheath (BS) tissues. The evolution of C2 photosynthesis has been hypothesized to be a major step in the origin of C4 photosynthesis, highlighting the importance of studying C2 evolution. In this study, physiological, anatomical, ultrastructural, and immunohistochemical properties of leaf photosynthetic tissues were investigated in six non-C4 Tribulus species and four C4 Tribulus species. At 42°C, T. cristatus exhibited a photosynthetic CO2 compensation point in the absence of respiration (C*) of 21 µmol mol-1, below the C3 mean C* of 73 µmol mol-1. Tribulus astrocarpus had a C* value at 42°C of 55 µmol mol-1, intermediate between the C3 species and the C2 T. cristatus. Glycine decarboxylase (GDC) allocation to BS tissues was associated with lower C*. Tribulus cristatus and T. astrocarpus allocated 86% and 30% of their GDC to the BS tissues, respectively, well above the C3 mean of 11%. Tribulus astrocarpus thus exhibits a weaker C2 (termed sub-C2) phenotype. Increased allocation of mitochondria to the BS and decreased length-to-width ratios of BS cells, were present in non-C4 species, indicating a potential role in C2 and C4 evolution.
Assuntos
Evolução Biológica , Fotossíntese , Folhas de Planta , Fotossíntese/fisiologia , Folhas de Planta/fisiologia , Folhas de Planta/metabolismo , Dióxido de Carbono/metabolismo , Glicina Desidrogenase (Descarboxilante)/metabolismoRESUMO
The C4 photosynthetic pathway is hypothesized to have evolved from the ancestral C3 pathway through progressive changes in leaf anatomy and biochemistry with extant C3-C4 photosynthetic intermediate species representing phenotypes between species demonstrating full C3 and full C4 states. The Australian endemic genus Neurachne is the only known grass group that contains distinct, closely related species that carry out C3, C3-C4 intermediate, or C4 photosynthesis. To explore and understand the molecular mechanisms underlying C4 photosynthesis evolution in this genus, leaf transcriptomes were generated from two C3, three photosynthetic intermediate (proto-Kranz, C2-like, and C2), and two C4 Neurachne species. The data were used to reconstruct phylogenetic relationships in Neurachne, which confirmed two independent C4 origins in the genus. Relative transcript abundances substantiated the photosynthetic phenotypes of individual species and highlighted transcriptional investment differences between species, including between the two C4 species. The data also revealed proteins potentially involved in C4 cycle intermediate transport and identified molecular mechanisms responsible for the evolution of C4-associated proteins in the genus.
RESUMO
The Calvin-Benson-Bassham (CBB) cycle is the ancestral CO2 assimilation pathway and is found in all photosynthetic organisms. Biochemical extensions to the CBB cycle have evolved that allow the resulting pathways to act as CO2 concentrating mechanisms, either spatially in the case of C4 photosynthesis or temporally in the case of Crassulacean acid metabolism (CAM). While the biochemical steps in the C4 and CAM pathways are known, questions remain on their integration and regulation with CBB cycle activity. The application of omic and transgenic technologies is providing a more complete understanding of the biochemistry of C4 and CAM species and will also provide insight into the CBB cycle in these plants. As the global population increases, new solutions are required to increase crop yields and meet demands for food and other bioproducts. Previous work in C3 species has shown that increasing carbon assimilation through genetic manipulation of the CBB cycle can increase biomass and yield. There may also be options to improve photosynthesis in species using C4 photosynthesis and CAM through manipulation of the CBB cycle in these plants. This is an underexplored strategy and requires more basic knowledge of CBB cycle operation in these species to enable approaches for increased productivity.
Assuntos
Dióxido de Carbono , Metabolismo Ácido das Crassuláceas , Dióxido de Carbono/metabolismo , Fotossíntese/fisiologiaRESUMO
A key feature of C4 Kranz anatomy is the presence of an enlarged, photosynthetically highly active bundle sheath whose cells contain large numbers of chloroplasts. With the aim to identify novel candidate regulators of C4 bundle sheath development, we performed an activation tagging screen with Arabidopsis thaliana. The reporter gene used encoded a chloroplast-targeted GFP protein preferentially expressed in the bundle sheath, and the promoter of the C4 phosphoenolpyruvate carboxylase gene from Flaveria trinervia served as activation tag because of its activity in all chlorenchymatous tissues of A. thaliana. Primary mutants were selected based on their GFP signal intensity, and one stable mutant named kb-1 with a significant increase in GFP fluorescence intensity was obtained. Despite the increased GFP signal, kb-1 showed no alterations to bundle sheath anatomy. The causal locus, AT1G29480, is specific to the Brassicaceae with its second exon being conserved. Overexpression and reconstitution studies confirmed that AT1G29480, and specifically its second exon, were sufficient for the enhanced GFP phenotype, which was not dependent on translation of the locus or its parts into protein. We conclude, therefore, that the AT1G29480 locus enhances the GFP reporter gene activity via an RNA-based mechanism.
RESUMO
C4 photosynthesis concentrates CO2 around Rubisco in the bundle sheath, favouring carboxylation over oxygenation and decreasing photorespiration. This complex trait evolved independently in >60 angiosperm lineages. Its evolution can be investigated in genera such as Flaveria (Asteraceae) that contain species representing intermediate stages between C3 and C4 photosynthesis. Previous studies have indicated that the first major change in metabolism probably involved relocation of glycine decarboxylase and photorespiratory CO2 release to the bundle sheath and establishment of intercellular shuttles to maintain nitrogen stoichiometry. This was followed by selection for a CO2-concentrating cycle between phosphoenolpyruvate carboxylase in the mesophyll and decarboxylases in the bundle sheath, and relocation of Rubisco to the latter. We have profiled 52 metabolites in nine Flaveria species and analysed 13CO2 labelling patterns for four species. Our results point to operation of multiple shuttles, including movement of aspartate in C3-C4 intermediates and a switch towards a malate/pyruvate shuttle in C4-like species. The malate/pyruvate shuttle increases from C4-like to complete C4 species, accompanied by a rise in ancillary organic acid pools. Our findings support current models and uncover further modifications of metabolism along the evolutionary path to C4 photosynthesis in the genus Flaveria.
Assuntos
Flaveria , Flaveria/genética , Flaveria/metabolismo , Glicina Desidrogenase (Descarboxilante)/genética , Glicina Desidrogenase (Descarboxilante)/metabolismo , Metaboloma , Fotossíntese , Ribulose-Bifosfato Carboxilase/genética , Ribulose-Bifosfato Carboxilase/metabolismoRESUMO
C4 photosynthesis is a remarkable complex trait, elucidations of the evolutionary trajectory of C4 photosynthesis from its ancestral C3 pathway can help us better understand the generic principles of the evolution of complex traits and guide the engineering of C3 crops for higher yields. Here, we used the genus Flaveria that contains C3, C3-C4, C4-like and C4 species as a system to study the evolution of C4 photosynthesis. We first mapped transcript abundance, protein sequence and morphological features onto the phylogenetic tree of the genus Flaveria, and calculated the evolutionary correlation of different features; we then predicted the relative changes of ancestral nodes of those features to illustrate the major events during the evolution of C4 photosynthesis. We found that gene expression and protein sequence showed consistent modification patterns in the phylogenetic tree. High correlation coefficients ranging from 0.46 to 0.9 among gene expression, protein sequence and morphology were observed. The greatest modification of those different features consistently occurred at the transition between C3-C4 species and C4-like species. Our results show highly coordinated changes in gene expression, protein sequence and morphological features, which support evolutionary major events during the evolution of C4 metabolism.
Assuntos
Flaveria , Fotossíntese , Filogenia , Evolução Biológica , Cloroplastos/metabolismoRESUMO
Background: Lifestyle intervention programs comprise the first-choice therapy to reduce the cardiovascular risk factors in metabolic syndrome (MetS). Our aim was to compare the effects of three lifestyle modification programs on the number of diagnostic criteria and clinical parameters of MetS. Methods: Twelve-week clinical trial, including 125 adults who presented at least three of the criteria defined by the revised NCEP ATP III (National Cholesterol Education Program Adult Panel III) for MetS. Individuals were randomized into three multidisciplinary intervention groups: Standard Intervention (SI), Group Intervention (GI) and Individual Intervention (II). Results: Seventy-one individuals, aged 34-59 years, concluded the study: SI: 20, GI: 25, and II: 26. The GI and II groups presented a significant decrease of body mass index, abdominal circumference, diastolic and systolic arterial pressure after intervention. The number of diagnostic criteria for MetS decreased significantly. Within the GI and II groups, 16.0% and 15.4% of the individuals, respectively, did not meet the criteria for the clinical condition studied at the end of the interventions. In the II group, the percentage of individuals with five criteria reduced 83.5%. In the GI group, the percentage of individuals with five criteria remained the same, but the number of individuals with four criteria presented a 50.0% reduction. Conclusions: Results reinforce that nonpharmacological strategies for changing lifestyle affect the reduction of cardiovascular risk factors existing in MetS. They are also able to remove the population from this clinical condition by decreasing the diagnostic criteria. II or GI lead to a successful treatment of MetS, especially when conducted by multidisciplinary team. Brazilian Registry of Clinical Trials-ReBEC number: RBR-9wz5fc.
Assuntos
Doenças Cardiovasculares/prevenção & controle , Dieta Saudável , Exercício Físico , Estilo de Vida Saudável , Síndrome Metabólica/terapia , Comportamento de Redução do Risco , Adulto , Brasil , Fatores de Risco Cardiometabólico , Doenças Cardiovasculares/diagnóstico , Doenças Cardiovasculares/etiologia , Comportamento Alimentar , Feminino , Humanos , Masculino , Síndrome Metabólica/complicações , Síndrome Metabólica/diagnóstico , Pessoa de Meia-Idade , Valor Nutritivo , Medição de Risco , Fatores de Tempo , Resultado do TratamentoRESUMO
Introduction of a C4 photosynthetic mechanism into C3 crops offers an opportunity to improve photosynthetic efficiency, biomass and yield in addition to potentially improving nitrogen and water use efficiency. To create a two-cell metabolic prototype for an NADP-malic enzyme type C4 rice, we transformed Oryza sativa spp. japonica cultivar Kitaake with a single construct containing the coding regions of carbonic anhydrase, phosphoenolpyruvate (PEP) carboxylase, NADP-malate dehydrogenase, pyruvate orthophosphate dikinase and NADP-malic enzyme from Zea mays, driven by cell-preferential promoters. Gene expression, protein accumulation and enzyme activity were confirmed for all five transgenes, and intercellular localization of proteins was analysed. 13 CO2 labelling demonstrated a 10-fold increase in flux though PEP carboxylase, exceeding the increase in measured in vitro enzyme activity, and estimated to be about 2% of the maize photosynthetic flux. Flux from malate via pyruvate to PEP remained low, commensurate with the low NADP-malic enzyme activity observed in the transgenic lines. Physiological perturbations were minor and RNA sequencing revealed no substantive effects of transgene expression on other endogenous rice transcripts associated with photosynthesis. These results provide promise that, with enhanced levels of the C4 proteins introduced thus far, a functional C4 pathway is achievable in rice.
Assuntos
Oryza , Malato Desidrogenase/genética , Malato Desidrogenase/metabolismo , Oryza/genética , Oryza/metabolismo , Fosfoenolpiruvato Carboxilase/genética , Fosfoenolpiruvato Carboxilase/metabolismo , Fotossíntese , Piruvato Ortofosfato Diquinase/genética , Piruvato Ortofosfato Diquinase/metabolismo , Zea mays/metabolismoRESUMO
Abstract The present study is the translation and adaptation of an intervention based on the Transtheoretical Model of Change directed to patients at cardiovascular risk. The original 29-session manual was translated into Portuguese, extended to 36 sessions, and applied in a pilot study. The protocol was reduced to 12 sessions through the grouping based on the processes of change. The products of the present study were the processes in each behavior, the versatility of the protocol, the interdisciplinary work, the great cost-benefit, and the potential of effectiveness. Finally, the importance of consistent training in Motivational Interview, Transtheoretical Model and group management for proper use are highlighted, and the costs and benefits of protocols versus more flexible treatments are discussed.
Resumo Trata-se da tradução e adaptação de uma intervenção baseada no Modelo Transteórico de Mudança e direcionada para pacientes com risco cardiovascular. Inicialmente, o manual original com 29 sessões foi traduzido para a língua portuguesa, sendo ampliado para 36 sessões, e posteriormente aplicado em um estudo piloto. A partir de agrupamentos pelos processos de mudança, foi reduzido para 12 sessões. Evidencia-se como produtos deste trabalho o aprofundamento dos processos em cada comportamento, a versatilidade do protocolo, o trabalho interdisciplinar, o ótimo custo-benefício e o potencial de efetividade. Por fim, salienta-se a importância de um treinamento consistente em Entrevista Motivacional, Modelo Transteórico e manejo de grupos para sua utilização adequada, e discute-se os custos e os benefícios dos protocolos versus tratamentos mais flexíveis.
RESUMO
O objetivo deste estudo foi identificar fatores potenciais de impacto sobre a desistência em um ensaio clínico randomizado. Para participar das 12 semanas de intervenção, foram selecionados adolescentes com sobrepeso ou obesidade (IMC percentil ≥ 85), dos sexos masculino e feminino, com média de idade 16,44±1,09. Utilizou-se regressão logística binária para responder pelo desfecho de não conclusão. As regressões foram realizadas por blocos e as variáveis estudadas foram características demográficas e antropométricas, marcadores metabólicos, aspectos motivacionais, funcionamento psicológico e percepção das práticas alimentares parentais. Foi constatada uma taxa de desistência de 48,8%. A maior chance de desistência foi explicada no modelo final de regressão pela autoeficácia para mudança de estilo de vida (OR: 0,732; IC95%: 0,581-0,921; p=0,008), pelo suporte familiar (OR: 0,807; IC95%: 0,685-0,950; p=0,010), pela HbA1c (OR: 11,366; IC95%: 3,123-37,702; p=0,010) e por fatores relacionados às práticas alimentares parentais. Concluiu-se que a família e os aspectos relacionados à motivação para mudança podem ser determinantes de desistência em programas para perda de peso para adolescentes.
The number of overweight or obese adolescents has increased, but the rate of dropping out in programs for this population is high. The objective of this study was to identify potential factors of impact on dropping out in a randomized clinical trial. To take part in a 12-week intervention, overweight or obese adolescents (BMI percentile ≥ 85), males and females, with mean age 17.99±1.09 were selected. Binary logistic regression was used to account for the non-completion outcome. Regressions were performed by blocks and the variables studied were demographic and anthropometric traits, metabolic markers, motivational aspects, psychological functioning, and perception of parental food practices. The drop-out rate was 48.8%. The highest chance of withdrawal was explained in the final regression model by self-efficacy to change lifestyle (OR: 0.732; IC95%: 0.581-0.921; p=0.008), by family support (OR: 0.807; IC95%: 0.685-0.950; p=0.010), by HbA1c (OR: 11.366; IC95%: 3.123-37.702; p=0.010), and by factors associated with parental food practices. It was concluded that the family and aspects related to motivation for change can be determinants of dropping-out in weight-loss programs for teenagers.
El objetivo de este estudio fue identificar factores potenciales del impacto sobre la desistencia en un ensayo clínico aleatorizado. Se seleccionaron para participar en 12 semanas de intervención, adolescentes con sobrepeso u obesidad (IMC percentil ≥ 85), del sexo masculino y femenino con media de edad 16,44±1,09. Se utilizó regresión logística binaria para responder por el resultado de conclusión incompleta. Las regresiones fueron realizadas por bloques y las variables estudiadas fueron características demográficas y antropométricas, marcadores metabólicos, aspectos motivacionales, funcionamiento psicológico y percepción de las prácticas alimentarias parentales. La tasa de desistimiento fue de 48,8%. La mayor probabilidad de desistimiento fue explicada en el modelo final de regresión por auto-eficacia para el cambio de estilo de vida (OR: 0,732, IC95%: 0,581-0,921, p=0,008), por el soporte familiar (OR: 0,807, IC95%: 0,685-0,950; p=0,010), por la HbA1c (OR: 11,366, IC95%: 3,123-37,702; p=0,010) y por factores relacionados con las prácticas alimentarias parentales. Se concluyó que la familia y los aspectos relacionados con la motivación para el cambio pueden ser determinantes de la desistencia de programas para la pérdida de peso para los adolescentes.
RESUMO
The Australian grass subtribe Neurachninae contains closely related species that use C3, C4, and C2 photosynthesis. To gain insight into the evolution of C4 photosynthesis in grasses, we examined leaf gas exchange, anatomy and ultrastructure, and tissue localization of Gly decarboxylase subunit P (GLDP) in nine Neurachninae species. We identified previously unrecognized variation in leaf structure and physiology within Neurachne that represents varying degrees of C3-C4 intermediacy in the Neurachninae. These include inverse correlations between the apparent photosynthetic carbon dioxide (CO2) compensation point in the absence of day respiration (C * ) and chloroplast and mitochondrial investment in the mestome sheath (MS), where CO2 is concentrated in C2 and C4 Neurachne species; width of the MS cells; frequency of plasmodesmata in the MS cell walls adjoining the parenchymatous bundle sheath; and the proportion of leaf GLDP invested in the MS tissue. Less than 12% of the leaf GLDP was allocated to the MS of completely C3 Neurachninae species with C * values of 56-61 µmol mol-1, whereas two-thirds of leaf GLDP was in the MS of Neurachne lanigera, which exhibits a newly-identified, partial C2 phenotype with C * of 44 µmol mol-1 Increased investment of GLDP in MS tissue of the C2 species was attributed to more MS mitochondria and less GLDP in mesophyll mitochondria. These results are consistent with a model where C4 evolution in Neurachninae initially occurred via an increase in organelle and GLDP content in MS cells, which generated a sink for photorespired CO2 in MS tissues.
Assuntos
Folhas de Planta/metabolismo , Proteínas de Plantas/metabolismo , Fotossíntese/genética , Fotossíntese/fisiologia , Folhas de Planta/fisiologia , Proteínas de Plantas/genética , Plasmodesmos/metabolismo , Plasmodesmos/fisiologia , Poaceae/genética , Poaceae/fisiologiaRESUMO
Low atmospheric CO2 in recent geological time led to the evolution of carbon-concentrating mechanisms (CCMs) such as C4 photosynthesis in >65 terrestrial plant lineages. We know little about the impact of low CO2 on the Calvin-Benson cycle (CBC) in C3 species that did not evolve CCMs, representing >90% of terrestrial plant species. Metabolite profiling provides a top-down strategy to investigate the operational balance in a pathway. We profiled CBC intermediates in a panel of C4 (Zea mays, Setaria viridis, Flaveria bidentis, and F. trinervia) and C3 species (Oryza sativa, Triticium aestivum, Arabidopsis thaliana, Nicotiana tabacum, and Manihot esculenta). Principal component analysis revealed differences between C4 and C3 species that were driven by many metabolites, including lower ribulose 1,5-bisphosphate in C4 species. Strikingly, there was also considerable variation between C3 species. This was partly due to different chlorophyll and protein contents, but mainly to differences in relative levels of metabolites. Correlation analysis indicated that one contributory factor was the balance between fructose-1,6-bisphosphatase, sedoheptulose-1,7-bisphosphatase, phosphoribulokinase, and Rubisco. Our results point to the CBC having experienced different evolutionary trajectories in C3 species since the ancestors of modern plant lineages diverged. They underline the need to understand CBC operation in a wide range of species.
Assuntos
Ciclo do Carbono , Magnoliopsida/metabolismo , Fotossíntese , Especificidade da EspécieRESUMO
Sesuvium sesuvioides (Sesuvioideae, Aizoaceae) is a perennial, salt-tolerant herb distributed in flats, depressions, or disturbed habitats of southern Africa and the Cape Verdes. Based on carbon isotope values, it is considered a C4 species, despite a relatively high ratio of mesophyll to bundle sheath cells (2.7:1) in the portulacelloid leaf anatomy. Using leaf anatomy, immunocytochemistry, gas exchange measurements, and enzyme activity assays, we sought to identify the biochemical subtype of C4 photosynthesis used by S. sesuvioides and to explore the anatomical, physiological, and biochemical traits of young, mature, and senescing leaves, with the aim to elucidate the plasticity and possible limitations of the photosynthetic efficiency in this species. Assays indicated that S. sesuvioides employs the NADP-malic enzyme as the major decarboxylating enzyme. The activity of C4 enzymes, however, declined as leaves aged, and the proportion of water storage tissue increased while air space decreased. These changes suggest a functional shift from photosynthesis to water storage in older leaves. Interestingly, S. sesuvioides demonstrated CO2 compensation points ranging between C4 and C3-C4 intermediate values, and immunocytochemistry revealed labeling of the Rubisco large subunit in mesophyll cells. We hypothesize that S. sesuvioides represents a young C4 lineage with C4-like photosynthesis in which C3 and C4 cycles are running simultaneously in the mesophyll.
Assuntos
Aizoaceae/fisiologia , Carbono/metabolismo , Fotossíntese , Folhas de Planta/fisiologia , Ciclo do Carbono , Células do Mesofilo/metabolismoRESUMO
A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.
RESUMO
Assessment is a central component of course curriculums and is used to certify student learning, but it can also be used as a tool to improve teaching and learning. Many laboratory courses are structured such that there is only a grade for a particular laboratory, which limits the insights that can be gained in student learning. We developed a laboratory program that incorporates assessments designed to probe student understanding of different components of the individual modules making up the program. The challenge was to analyze and present grades from these assessment tasks in a format that was readily interpretable by academics. We show that a simplified synthesis of grade distributions (grade distribution digests) provides sufficient information to make decisions about changes in course components. The main feature of the digests is its data visualization approach, where student grades for individual laboratory practicals, individual assessment tasks or individual assessment items are graphically presented as an overall average grade, an average top quartile grade and an average bottom quartile grade, and relative averages across all assessments. This ability to visualize student grades in variety of contexts enables academics with many other demands on their time (e.g. research and administration) to more efficiently identify ways to improve teaching delivery and learning outcomes. Examples are presented of the use of such data to identify and improve deficiencies in both student skills and teaching practice, resulting in improved learning outcomes. © 2017 by The International Union of Biochemistry and Molecular Biology, 46(2):130-140, 2018.
Assuntos
Avaliação Educacional , Laboratórios , Aprendizagem , Ensino/educação , Humanos , Estudantes , UniversidadesRESUMO
The past few years have witnessed a paradigm shift in molecular systematics from phylogenetic methods (using one or a few genes) to those that can be described as phylogenomics (phylogenetic inference with entire genomes). One approach that has recently emerged is phylo-transcriptomics (transcriptome-based phylogenetic inference). As in any phylogenetics experiment, accurate orthology inference is critical to phylo-transcriptomics. To date, most analyses have inferred orthology based either on pure sequence similarity or using gene-tree approaches. The use of conserved genome synteny in orthology detection has been relatively under-employed in phylogenetics, mainly due to the cost of sequencing genomes. While current trends focus on the quantity of genes included in an analysis, the use of synteny is likely to improve the quality of ortholog inference. In this study, we combine de novo transcriptome data and sequenced genomes from an economically important group of grass species, the tribe Paniceae, to make phylogenomic inferences. This method, which we call "genome-guided phylo-transcriptomics", is compared to other recently published orthology inference pipelines, and benchmarked using a set of sequenced genomes from across the grasses. These comparisons provide a framework for future researchers to evaluate the costs and benefits of adding sequenced genomes to transcriptome data sets.
Assuntos
Genoma de Planta , Poaceae/genética , Perfilação da Expressão Gênica , Filogenia , Poaceae/classificação , RNA de Plantas/química , RNA de Plantas/isolamento & purificação , RNA de Plantas/metabolismo , Análise de Sequência de RNARESUMO
Neurachne is the only known grass lineage containing closely related C3, C3-C4 intermediate, and C4 species, making it an ideal taxon with which to study the evolution of C4 photosynthesis in the grasses. To begin dissecting the molecular changes that led to the evolution of C4 photosynthesis in this group, the complementary DNAs encoding four distinct ß-carbonic anhydrase (CA) isoforms were characterized from leaf tissue of Neurachne munroi (C4), Neurachne minor (C3-C4), and Neurachne alopecuroidea (C3). Two genes (CA1 and CA2) each encode two different isoforms: CA1a/CA1b and CA2a/CA2b. Transcript analyses found that CA1 messenger RNAs were significantly more abundant than transcripts from the CA2 gene in the leaves of each species examined, constituting â¼99% of all ß-CA transcripts measured. Localization experiments using green fluorescent protein fusion constructs showed that, while CA1b is a cytosolic CA in all three species, the CA1a proteins are differentially localized. The N. alopecuroidea and N. minor CA1a isoforms were imported into chloroplasts of Nicotiana benthamiana leaf cells, whereas N. munroi CA1a localized to the cytosol. Sequence analysis indicated an 11-amino acid deletion in the amino terminus of N. munroi CA1a relative to the C3 and C3-C4 proteins, suggesting that chloroplast targeting of CA1a is the ancestral state and that loss of a functional chloroplast transit peptide in N. munroi CA1a is associated with the evolution of C4 photosynthesis in Neurachne spp. Remarkably, this mechanism is homoplastic with the evolution of the C4-associated CA in the dicotyledonous genus Flaveria, although the actual mutations in the two lineages differ.
Assuntos
Anidrases Carbônicas/genética , Proteínas de Cloroplastos/genética , Fotossíntese/genética , Poaceae/genética , Sinais Direcionadores de Proteínas/genética , Sequência de Aminoácidos , Anidrase Carbônica I/genética , Anidrase Carbônica II/genética , Citoplasma/enzimologia , Citosol/enzimologia , Evolução Molecular , Deleção de Genes , Regulação Enzimológica da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Isoenzimas/genética , Microscopia Confocal , Folhas de Planta/enzimologia , Folhas de Planta/genética , Poaceae/classificação , Poaceae/enzimologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência de DNA , Homologia de Sequência de AminoácidosRESUMO
The first two reactions of C4 photosynthesis are catalysed by carbonic anhydrase (CA) and phosphoenolpyruvate carboxylase (PEPC) in the leaf mesophyll (M) cell cytosol. Translatome experiments using a tagged ribosomal protein expressed under the control of M and bundle-sheath (BS) cell-specific promoters showed transcripts encoding CA3 from the C4 species Flaveria bidentis were highly enriched in polysomes from M cells relative to those of the BS. Localisation experiments employing a CA3-green fluorescent protein fusion protein showed F. bidentis CA3 is a cytosolic enzyme. A motif showing high sequence homology to that of the Flaveria M expression module 1 (MEM1) element was identified approximately 2 kb upstream of the F. bidentis and F. trinervia ca3 translation start sites. MEM1 is located in the promoter of C4 Flaveria ppcA genes, which encode the C4-associated PEPC, and is necessary for M-specific expression. No MEM1-like sequence was found in the 4 kb upstream of the C3 species F. pringlei ca3 translation start site. Promoter-reporter fusion experiments demonstrated the region containing the ca3 MEM1-like element also directs M-specific expression. These results support the idea that a common regulatory switch drives the expression of the C4 Flaveria ca3 and ppcA1 genes specifically in M cells.
Assuntos
Flaveria/enzimologia , Regulação da Expressão Gênica de Plantas , Células do Mesofilo/enzimologia , Sequência de Bases , Flaveria/genética , Dados de Sequência MolecularRESUMO
Carbonic anhydrases (CAs) are zinc metalloenzymes that catalyze the interconversion of CO2 and HCO3- and are ubiquitous in nature. Higher plants contain three evolutionarily distinct CA families, αCAs, ßCAs, and γCAs, where each family is represented by multiple isoforms in all species. Alternative splicing of CA transcripts appears common; consequently, the number of functional CA isoforms in a species may exceed the number of genes. CAs are expressed in numerous plant tissues and in different cellular locations. The most prevalent CAs are those in the chloroplast, cytosol, and mitochondria. This diversity in location is paralleled in the many physiological and biochemical roles that CAs play in plants. In this review, the number and types of CAs in C3, C4, and crassulacean acid metabolism (CAM) plants are considered, and the roles of the α and γCAs are briefly discussed. The remainder of the review focuses on plant ßCAs and includes the identification of homologs between species using phylogenetic approaches, a consideration of the inter- and intracellular localization of the proteins, along with the evidence for alternative splice forms. Current understanding of ßCA tissue-specific expression patterns and what controls them are reviewed, and the physiological roles for which ßCAs have been implicated are presented.