RESUMEN
Availability of light and CO2, substrates of microalgae photosynthesis, is frequently far from optimal. Microalgae activate photoprotection under strong light, to prevent oxidative damage, and the CO2 Concentrating Mechanism (CCM) under low CO2, to raise intracellular CO2 levels. The two processes are interconnected; yet, the underlying transcriptional regulators remain largely unknown. Employing a large transcriptomic data compendium of Chlamydomonas reinhardtii's responses to different light and carbon supply, we reconstruct a consensus genome-scale gene regulatory network from complementary inference approaches and use it to elucidate transcriptional regulators of photoprotection. We show that the CCM regulator LCR1 also controls photoprotection, and that QER7, a Squamosa Binding Protein, suppresses photoprotection- and CCM-gene expression under the control of the blue light photoreceptor Phototropin. By demonstrating the existence of regulatory hubs that channel light- and CO2-mediated signals into a common response, our study provides an accessible resource to dissect gene expression regulation in this microalga.
Asunto(s)
Chlamydomonas reinhardtii , Chlamydomonas , Chlamydomonas reinhardtii/metabolismo , Dióxido de Carbono/metabolismo , Fotosíntesis/genética , Regulación de la Expresión Génica , Chlamydomonas/metabolismo , Carbono/metabolismoRESUMEN
Photosynthetic algae have evolved mechanisms to cope with suboptimal light and CO2 conditions. When light energy exceeds CO2 fixation capacity, Chlamydomonas reinhardtii activates photoprotection, mediated by LHCSR1/3 and PSBS, and the CO2 Concentrating Mechanism (CCM). How light and CO2 signals converge to regulate these processes remains unclear. Here, we show that excess light activates photoprotection- and CCM-related genes by altering intracellular CO2 concentrations and that depletion of CO2 drives these responses, even in total darkness. High CO2 levels, derived from respiration or impaired photosynthetic fixation, repress LHCSR3/CCM genes while stabilizing the LHCSR1 protein. Finally, we show that the CCM regulator CIA5 also regulates photoprotection, controlling LHCSR3 and PSBS transcript accumulation while inhibiting LHCSR1 protein accumulation. This work has allowed us to dissect the effect of CO2 and light on CCM and photoprotection, demonstrating that light often indirectly affects these processes by impacting intracellular CO2 levels.
Asunto(s)
Dióxido de Carbono , Chlamydomonas reinhardtii , Dióxido de Carbono/metabolismo , Complejo de Proteína del Fotosistema II/metabolismo , Fotosíntesis/genética , Proteínas/metabolismo , Chlamydomonas reinhardtii/metabolismoRESUMEN
Chloroplast-to-nucleus retrograde signalling (RS) is known to impact plant growth and development. In Arabidopsis, we and others have shown that RS affects seedling establishment by inhibiting deetiolation. In the presence of lincomycin, a chloroplast protein synthesis inhibitor that triggers RS, Arabidopsis light-grown seedlings display partial skotomorphogenesis with undeveloped plastids and closed cotyledons. By contrast, RS in monocotyledonous has been much less studied. Here, we show that emerging rice seedlings exposed to lincomycin do not accumulate chlorophyll but otherwise remain remarkably unaffected. However, by using high red (R) and blue (B) monochromatic lights in combination with lincomycin, we have uncovered a RS inhibition of length and a reduction in the B light-induced declination of the second leaf. Furthermore, we present data showing that seedlings grown in high B and R light display different non-photochemical quenching capacity. Our findings support the view that excess B and R light impact seedling photomorphogenesis differently to photoprotect and optimize the response to high-light stress. This article is part of the theme issue 'Retrograde signalling from endosymbiotic organelles'.
Asunto(s)
Clorofila/metabolismo , Proteínas de Cloroplastos/metabolismo , Etiolado/efectos de la radiación , Luz , Lincomicina/farmacología , Oryza/efectos de la radiación , Inhibidores de la Síntesis de la Proteína/farmacología , Transducción de Señal/efectos de la radiación , Oryza/fisiología , Plantones/fisiología , Plantones/efectos de la radiaciónRESUMEN
Plant cells can change their identity based on positional information, a mechanism that confers developmental plasticity to plants. This ability, common to distinct multicellular organisms, is particularly relevant for plant phloem cells. Protophloem sieve elements (PSEs), one type of phloem conductive cells, act as the main organizers of the phloem pole, which comprises four distinct cell files organized in a conserved pattern. Here, we report how Arabidopsis roots generate a reservoir of meristematic phloem cells competent to swap their cell identities. Although PSE misspecification induces cell identity hybridism, the activity of RECEPTOR LIKE PROTEIN KINASE 2 (RPK2) by perceiving CLE45 peptide contributes to restrict PSE identity to the PSE position. By maintaining a spatiotemporal window when PSE and PSE-adjacent cells' identities are interchangeable, CLE45 signaling endows phloem cells with the competence to re-pattern a functional phloem pole when protophloem fails to form.
Asunto(s)
Arabidopsis/crecimiento & desarrollo , Floema/crecimiento & desarrollo , Raíces de Plantas/crecimiento & desarrollo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Proteínas de la Membrana/metabolismo , Meristema/crecimiento & desarrollo , Meristema/metabolismo , Floema/metabolismo , Raíces de Plantas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Transducción de SeñalRESUMEN
While light is a crucial energy source in photosynthetic organisms, if its intensity exceeds their photosynthetic capacity it may cause light-induced damage. A dominant photoprotective mechanism in plants and algae is the qE (quenching of energy), the major component of nonphotochemical quenching (NPQ). qE is a process that dissipates absorbed excitation energy as heat, ensuring cell survival even under adverse conditions. The present protocol gathers together a set of experimental approaches (in vivo chlorophyll fluorescence, western blotting, growth and cellular chlorophyll content at very strong light) that collectively allow for the characterization of the qE capacity of the model green algae Chlamydomonas reinhardtii.
Asunto(s)
Chlorophyta/fisiología , Procesos Fotoquímicos , Fotoquímica/métodos , Células Cultivadas , Clorofila/metabolismo , Fluorescencia , Luz , Microscopía Fluorescente , FotosíntesisRESUMEN
Protophloem and metaphloem sieve tubes are essential for transporting carbohydrates and signalling molecules towards sink tissues. OCTOPUS (OPS) was previously identified as an important regulator of protophloem differentiation in Arabidopsis roots. Here, we investigated the role of OCTOPUS-LIKE 2 (OPL2), a gene homologous to OPS. OPL2 expression patterns were analysed, and functional equivalence of OPS and OPL2 was tested. Mutant and double mutant phenotypes were investigated. OPS and OPL2 displayed overlapping expression patterns and a high degree of functional overlap. A mutation in OPL2 revealed redundant functions of OPS and OPL2 in developmental processes in which OPS was known to play a role, notably cotyledon vascular patterning and protophloem development. Moreover, we also uncovered redundant roles for OPS and OPL2 in leaf vascular patterning and, most interestingly, metaphloem sieve tube differentiation. Our results reveal a novel OPS-like protein that, together with OPS, is an important regulator of vascular patterning, root growth and phloem development. OPS and OPL2 are the first genes identified that play a role in metaphloem sieve tube differentiation.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/crecimiento & desarrollo , Proteínas de la Membrana/metabolismo , Raíces de Plantas/crecimiento & desarrollo , Haz Vascular de Plantas/crecimiento & desarrollo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de la Membrana/genética , Familia de Multigenes , Raíces de Plantas/citología , Haz Vascular de Plantas/citologíaRESUMEN
A wide diversity of isoprenoids is produced in different plant compartments. Most groups of isoprenoids synthesized in plastids, and some produced elsewhere in the plant cell derive from geranylgeranyl diphosphate (GGPP) synthesized by GGPP synthase (GGPPS) enzymes. In Arabidopsis (Arabidopsis thaliana), five genes appear to encode GGPPS isoforms localized in plastids (two), the endoplasmic reticulum (two), and mitochondria (one). However, the loss of function of the plastid-targeted GGPPS11 isoform (referred to as G11) is sufficient to cause lethality. Here, we show that the absence of a strong transcription initiation site in the G11 gene results in the production of transcripts of different lengths. The longer transcripts encode an isoform with a functional plastid import sequence that produces GGPP for the major groups of photosynthesis-related plastidial isoprenoids. However, shorter transcripts are also produced that lack the first translation initiation codon and rely on a second in-frame ATG codon to produce an enzymatically active isoform lacking this N-terminal domain. This short enzyme localizes in the cytosol and is essential for embryo development. Our results confirm that the production of differentially targeted enzyme isoforms from the same gene is a central mechanism to control the biosynthesis of isoprenoid precursors in different plant cell compartments.
Asunto(s)
Transferasas Alquil y Aril/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimología , Arabidopsis/genética , Genes de Plantas , Transferasas Alquil y Aril/genética , Alelos , Proteínas de Arabidopsis/genética , Secuencia de Bases , Vías Biosintéticas/genética , Pruebas de Enzimas , Isoenzimas/genética , Isoenzimas/metabolismo , Ácido Mevalónico/metabolismo , Fenotipo , Plastidios/metabolismo , Biosíntesis de Proteínas/genética , Semillas/metabolismo , Fracciones Subcelulares/metabolismo , Terpenos/química , Terpenos/metabolismo , Sitio de Iniciación de la TranscripciónRESUMEN
Most plastid isoprenoids, including photosynthesis-related metabolites such as carotenoids and the side chain of chlorophylls, tocopherols (vitamin E), phylloquinones (vitamin K), and plastoquinones, derive from geranylgeranyl diphosphate (GGPP) synthesized by GGPP synthase (GGPPS) enzymes. Seven out of 10 functional GGPPS isozymes in Arabidopsis thaliana reside in plastids. We aimed to address the function of different GGPPS paralogues for plastid isoprenoid biosynthesis. We constructed a gene co-expression network (GCN) using GGPPS paralogues as guide genes and genes from the upstream and downstream pathways as query genes. Furthermore, knock-out and/or knock-down ggpps mutants were generated and their growth and metabolic phenotypes were analyzed. Also, interacting protein partners of GGPPS11 were searched for. Our data showed that GGPPS11, encoding the only plastid isozyme essential for plant development, functions as a hub gene among GGPPS paralogues and is required for the production of all major groups of plastid isoprenoids. Furthermore, we showed that the GGPPS11 protein physically interacts with enzymes that use GGPP for the production of carotenoids, chlorophylls, tocopherols, phylloquinone, and plastoquinone. GGPPS11 is a hub isozyme required for the production of most photosynthesis-related isoprenoids. Both gene co-expression and protein-protein interaction likely contribute to the channeling of GGPP by GGPPS11.
Asunto(s)
Transferasas Alquil y Aril/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimología , Terpenos/metabolismo , Transferasas Alquil y Aril/genética , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Carotenoides/metabolismo , Clorofila/metabolismo , Isoenzimas , Fenotipo , Fotosíntesis , Plastidios/enzimología , Fosfatos de Poliisoprenilo/metabolismo , Mapeo de Interacción de ProteínasRESUMEN
Carotenoids are isoprenoid compounds that are essential for plants to protect the photosynthetic apparatus against excess light. They also function as health-promoting natural pigments that provide colors to ripe fruit, promoting seed dispersal by animals. Work in Arabidopsis thaliana unveiled that transcription factors of the phytochrome-interacting factor (PIF) family regulate carotenoid gene expression in response to environmental signals (i.e. light and temperature), including those created when sunlight reflects from or passes though nearby vegetation or canopy (referred to as shade). Here we show that PIFs use a virtually identical mechanism to modulate carotenoid biosynthesis during fruit ripening in tomato (Solanum lycopersicum). However, instead of integrating environmental information, PIF-mediated signaling pathways appear to fulfill a completely new function in the fruit. As tomatoes ripen, they turn from green to red due to chlorophyll breakdown and carotenoid accumulation. When sunlight passes through the flesh of green fruit, a self-shading effect within the tissue maintains high levels of PIFs that directly repress the master gene of the fruit carotenoid pathway, preventing undue production of carotenoids. This effect is attenuated as chlorophyll degrades, causing degradation of PIF proteins and boosting carotenoid biosynthesis as ripening progresses. Thus, shade signaling components may have been co-opted in tomato fruit to provide information on the actual stage of ripening (based on the pigment profile of the fruit at each moment) and thus finely coordinate fruit color change. We show how this mechanism may be manipulated to obtain carotenoid-enriched fruits.
Asunto(s)
Carotenoides/metabolismo , Regulación de la Expresión Génica de las Plantas/efectos de la radiación , Proteínas de Plantas/metabolismo , Transducción de Señal/efectos de la radiación , Solanum lycopersicum/fisiología , Clorofila/metabolismo , Ambiente , Etilenos/metabolismo , Frutas/genética , Frutas/fisiología , Frutas/efectos de la radiación , Luz , Solanum lycopersicum/genética , Solanum lycopersicum/efectos de la radiación , Fitocromo/metabolismo , Proteínas de Plantas/genética , Plantas Modificadas Genéticamente , Terpenos/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
Abscisic acid (ABA) is a hormone that plays a vital role in mediating abiotic stress responses in plants. Salt exposure induces the synthesis of ABA through the cleavage of carotenoid precursors (xanthophylls), which are found at very low levels in roots. Here we show that de novo ABA biosynthesis in salt-treated Arabidopsis thaliana roots involves an organ-specific induction of the carotenoid biosynthetic pathway. Upregulation of the genes encoding phytoene synthase (PSY) and other enzymes of the pathway producing ABA precursors was observed in roots but not in shoots after salt exposure. A pharmacological block of the carotenoid pathway substantially reduced ABA levels in stressed roots, confirming that an increase in carotenoid accumulation contributes to fuel hormone production after salt exposure. Treatment with exogenous ABA was also found to upregulate PSY expression only in roots, suggesting an organ-specific feedback regulation of the carotenoid pathway by ABA. Taken together, our results show that the presence of high concentrations of salt in the growth medium rapidly triggers a root-specific activation of the carotenoid pathway, probably to ensure a proper supply of ABA precursors required for a sustained production of the hormone.
Asunto(s)
Ácido Abscísico/metabolismo , Arabidopsis/fisiología , Carotenoides/metabolismo , Raíces de Plantas/fisiología , Cloruro de Sodio/metabolismo , Ácido Abscísico/genética , Arabidopsis/genética , Vías Biosintéticas , Carotenoides/genética , Regulación de la Expresión Génica de las Plantas , Raíces de Plantas/genética , Estrés FisiológicoRESUMEN
Carotenoids are plastidial isoprenoids essential for the protection of photosynthetic tissues against excess light. They also serve as precursors of apocarotenoid hormones such as abscisic acid (ABA) and strigolactones. The first enzyme of the carotenoid pathway, phytoene synthase (PSY), is also the main rate-limiting step. Unlike that observed in most plants, PSY is encoded by a single gene in Arabidopsis thaliana. Whereas the PSY gene is induced by light in photosynthetic tissues, a root-specific upregulation of PSY expression by salt stress and ABA has been recently demonstrated. Here we report that transcription factors of the Phytochrome-Interacting Factor (PIF) family, previously shown to repress PSY expression in etiolated seedlings and mature leaves, do not influence PSY expression in roots. Together, our results suggest that organ-specific pathways regulate PSY expression and hence carotenoid production in response to different environmental cues.
Asunto(s)
Arabidopsis/genética , Carotenoides/biosíntesis , Regulación de la Expresión Génica de las Plantas , Geranilgeranil-Difosfato Geranilgeraniltransferasa/genética , Luz , Raíces de Plantas/metabolismo , Factores de Transcripción/metabolismo , Ácido Abscísico/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Ambiente , Expresión Génica , Genes de Plantas , Geranilgeranil-Difosfato Geranilgeraniltransferasa/metabolismo , Fitocromo/metabolismo , Tolerancia a la Sal/genética , Cloruro de Sodio/metabolismo , Estrés Fisiológico/genética , Regulación hacia ArribaRESUMEN
Geranylgeranyl diphosphate (GGPP) is a key precursor of various isoprenoids that have diverse functions in plant metabolism and development. The annotation of the Arabidopsis thaliana genome predicts 12 genes to encode geranylgeranyl diphosphate synthases (GGPPS). In this study we analyzed GGPPS activity as well as the subcellular localization and tissue-specific expression of the entire protein family in A. thaliana. GGPPS2 (At2g18620), GGPPS3 (At2g18640), GGPPS6 (At3g14530), GGPPS7 (At3g14550), GGPPS8 (At3g20160), GGPPS9 (At3g29430), GGPPS10 (At3g32040) and GGPPS11 (At4g36810) showed GGPPS activity in Escherichia coli, similar to activities reported earlier for GGPPS1 (At1g49530) and GGPPS4 (At2g23800) (Zhu et al. in Plant Cell Physiol 38(3):357-361, 1997a; Plant Mol Biol 35(3):331-341, b). GGPPS12 (At4g38460) did not produce GGPP in E. coli. Based on DNA sequence analysis we propose that GGPPS5 (At3g14510) is a pseudogene. GGPPS-GFP (green fluorescent protein) fusion proteins of the ten functional GGPP synthases localized to plastids, mitochondria and the endoplasmic reticulum, with the majority of the enzymes located in plastids. Gene expression analysis using quantitative real time-PCR, GGPPS promoter-GUS (ß-glucuronidase) assays and publicly available microarray data revealed a differential spatio-temporal expression of GGPPS genes. The results suggest that plastids and mitochondria are key subcellular compartments for the synthesis of ubiquitous GGPP-derived isoprenoid species. GGPPS11 and GGPPS1 are the major isozymes responsible for their biosynthesis. All remaining paralogs, encoding six plastidial isozymes and two cytosolic isozymes, were expressed in specific tissues and/or at specific developmental stages, suggesting their role in developmentally regulated isoprenoid biosynthesis. Our results show that of the 12 predicted GGPPS encoded in the A. thaliana genome 10 are functional proteins that can synthesize GGPP. Their specific subcellular location and differential expression pattern suggest subfunctionalization in providing GGPP to specific tissues, developmental stages, or metabolic pathways.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimología , Farnesiltransferasa/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/genética , Farnesiltransferasa/química , Farnesiltransferasa/genética , Regulación de la Expresión Génica de las Plantas/genética , Regulación de la Expresión Génica de las Plantas/fisiología , Reacción en Cadena de la PolimerasaRESUMEN
Plant carotenoids are a family of pigments that participate in light harvesting and are essential for photoprotection against excess light. Furthermore, they act as precursors for the production of apocarotenoid hormones such as abscisic acid and strigolactones. In this review, we summarize the current knowledge on the genes and enzymes of the carotenoid biosynthetic pathway (which is now almost completely elucidated) and on the regulation of carotenoid biosynthesis at both transcriptional and post-transcriptional levels. We also discuss the relevance of Arabidopsis as a model system for the study of carotenogenesis and how metabolic engineering approaches in this plant have taught important lessons for carotenoid biotechnology.