RESUMO
Short- and long-distance circadian communication is essential for integration of temporal information. However, a major challenge in plant biology is to decipher how individual clocks are interconnected to sustain rhythms in the whole plant. Here we show that the shoot apex is composed of an ensemble of coupled clocks that influence rhythms in roots. Live-imaging of single cells, desynchronization of dispersed protoplasts, and mathematical analysis using barycentric coordinates for high-dimensional space show a gradation in the strength of circadian communication in different tissues, with shoot apex clocks displaying the highest coupling. The increased synchrony confers robustness of morning and evening oscillations and particular capabilities for phase readjustments. Rhythms in roots are altered by shoot apex ablation and micrografting, suggesting that signals from the shoot apex are able to synchronize distal organs. Similarly to the mammalian suprachiasmatic nucleus, shoot apexes play a dominant role within the plant hierarchical circadian structure.
Assuntos
Arabidopsis/fisiologia , Relógios Circadianos , Animais , Ritmo Circadiano , Hipocótilo/fisiologia , Mamíferos/fisiologia , Células Vegetais/fisiologia , Raízes de Plantas/fisiologia , Brotos de Planta/fisiologiaRESUMO
We present unresolved questions in plant abiotic stress biology as posed by 15 research groups with expertise spanning eco-physiology to cell and molecular biology. Common themes of these questions include the need to better understand how plants detect water availability, temperature, salinity, and rising carbon dioxide (CO2) levels; how environmental signals interface with endogenous signaling and development (e.g. circadian clock and flowering time); and how this integrated signaling controls downstream responses (e.g. stomatal regulation, proline metabolism, and growth versus defense balance). The plasma membrane comes up frequently as a site of key signaling and transport events (e.g. mechanosensing and lipid-derived signaling, aquaporins). Adaptation to water extremes and rising CO2 affects hydraulic architecture and transpiration, as well as root and shoot growth and morphology, in ways not fully understood. Environmental adaptation involves tradeoffs that limit ecological distribution and crop resilience in the face of changing and increasingly unpredictable environments. Exploration of plant diversity within and among species can help us know which of these tradeoffs represent fundamental limits and which ones can be circumvented by bringing new trait combinations together. Better defining what constitutes beneficial stress resistance in different contexts and making connections between genes and phenotypes, and between laboratory and field observations, are overarching challenges.
Assuntos
Dióxido de Carbono , Mudança Climática , Estresse Fisiológico , Dióxido de Carbono/metabolismo , Transpiração Vegetal/fisiologia , Plantas/metabolismo , Água/metabolismoRESUMO
HISTONE MONOUBIQUITINATION1 (HUB1) and its paralog HUB2 act in a conserved heterotetrameric complex in the chromatin-mediated transcriptional modulation of developmental programs, such as flowering time, dormancy, and the circadian clock. The KHD1 and SPEN3 proteins were identified as interactors of the HUB1 and HUB2 proteins with in vitro RNA-binding activity. Mutants in SPEN3 and KHD1 had reduced rosette and leaf areas. Strikingly, in spen3 mutants, the flowering time was slightly, but significantly, delayed, as opposed to the early flowering time in the hub1-4 mutant. The mutant phenotypes in biomass and flowering time suggested a deregulation of their respective regulatory genes CIRCADIAN CLOCK-ASSOCIATED1 (CCA1) and FLOWERING LOCUS C (FLC) that are known targets of the HUB1-mediated histone H2B monoubiquitination (H2Bub). Indeed, in the spen3-1 and hub1-4 mutants, the circadian clock period was shortened as observed by luciferase reporter assays, the levels of the CCA1α and CCA1ß splice forms were altered, and the CCA1 expression and H2Bub levels were reduced. In the spen3-1 mutant, the delay in flowering time was correlated with an enhanced FLC expression, possibly due to an increased distal versus proximal ratio of its antisense COOLAIR transcript. Together with transcriptomic and double-mutant analyses, our data revealed that the HUB1 interaction with SPEN3 links H2Bub during transcript elongation with pre-mRNA processing at CCA1 Furthermore, the presence of an intact HUB1 at the FLC is required for SPEN3 function in the formation of the FLC-derived antisense COOLAIR transcripts.
Assuntos
Proteínas de Arabidopsis , Regulação da Expressão Gênica de Plantas , Histonas , RNA de Plantas , Ubiquitina-Proteína Ligases , Ubiquitinação , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Relógios Circadianos/genética , Relógios Circadianos/fisiologia , Flores/genética , Flores/fisiologia , Regulação da Expressão Gênica de Plantas/genética , Regulação da Expressão Gênica de Plantas/fisiologia , Histonas/genética , Histonas/metabolismo , Domínios Proteicos/genética , Precursores de RNA/genética , Precursores de RNA/metabolismo , RNA de Plantas/genética , RNA de Plantas/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação/genética , Ubiquitinação/fisiologiaRESUMO
The Evening Complex (EC) is a core component of the Arabidopsis (Arabidopsis thaliana) circadian clock, which represses target gene expression at the end of the day and integrates temperature information to coordinate environmental and endogenous signals. Here we show that the EC induces repressive chromatin structure to regulate the evening transcriptome. The EC component ELF3 directly interacts with a protein from the SWI2/SNF2-RELATED (SWR1) complex to control deposition of H2A.Z-nucleosomes at the EC target genes. SWR1 components display circadian oscillation in gene expression with a peak at dusk. In turn, SWR1 is required for the circadian clockwork, as defects in SWR1 activity alter morning-expressed genes. The EC-SWR1 complex binds to the loci of the core clock genes PSEUDO-RESPONSE REGULATOR7 (PRR7) and PRR9 and catalyzes deposition of nucleosomes containing the histone variant H2A.Z coincident with the repression of these genes at dusk. This provides a mechanism by which the circadian clock temporally establishes repressive chromatin domains to shape oscillatory gene expression around dusk.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Cromatina/metabolismo , Histonas/metabolismo , Arabidopsis/genética , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Relógios Circadianos/fisiologia , Histonas/genética , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
The rhythms of steady-state mRNA expression pervade nearly all circadian systems. However, the mechanisms behind the rhythmic transcriptional synthesis and its correlation with circadian expression remain fully unexplored, particularly in plants. Here, we discovered a multifunctional protein complex that orchestrates the rhythms of transcriptional activity in Arabidopsis thaliana The expression of the circadian oscillator genes TIMING OF CAB EXPRESSION1/PSEUDO-RESPONSE REGULATOR1 and PSEUDO-RESPONSE REGULATOR5 initially relies on the modular function of the clock-related factor REVEILLE8: its MYB domain provides the DNA binding specificity, while its LCL domain recruits the clock components, NIGHT LIGHT-INDUCIBLE AND CLOCK-REGULATED proteins (LNKs), to target promoters. LNKs, in turn, specifically interact with RNA Polymerase II and the transcript elongation FACT complex to rhythmically co-occupy the target loci. The functional interaction of these components is central for chromatin status, transcript initiation, and elongation as well as for proper rhythms in nascent RNAs. Thus, our findings explain how genome readout of environmental information ultimately results in rhythmic changes of gene expression.
Assuntos
Arabidopsis/genética , Ritmo Circadiano/genética , Complexos Multiproteicos/metabolismo , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Cromatina/genética , Complexos Multiproteicos/genética , Regiões Promotoras Genéticas/genética , RNA Mensageiro/genética , RNA de Plantas/genéticaRESUMO
The Elongator complex (hereafter Elongator) promotes RNA polymerase II-mediated transcript elongation through epigenetic activities such as histone acetylation. Elongator regulates growth, development, immune response and sensitivity to drought and abscisic acid. We demonstrate that elo mutants exhibit defective hypocotyl elongation but have a normal apical hook in darkness and are hyposensitive to light during photomorphogenesis. These elo phenotypes are supported by transcriptome changes, including downregulation of circadian clock components, positive regulators of skoto- or photomorphogenesis, hormonal pathways and cell wall biogenesis-related factors. The downregulated genes LHY, HFR1 and HYH are selectively targeted by Elongator for histone H3K14 acetylation in darkness. The role of Elongator in early seedling development in darkness and light is supported by hypocotyl phenotypes of mutants defective in components of the gene network regulated by Elongator, and by double mutants between elo and mutants in light or darkness signaling components. A model is proposed in which Elongator represses the plant immune response and promotes hypocotyl elongation and photomorphogenesis via transcriptional control of positive photomorphogenesis regulators and a growth-regulatory network that converges on genes involved in cell wall biogenesis and hormone signaling.This article has an associated First Person interview with the first author of the paper.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Escuridão , Morfogênese/efeitos da radiação , Complexos Multiproteicos/metabolismo , Acetilação , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Ritmo Circadiano/fisiologia , Epistasia Genética , Regulação da Expressão Gênica de Plantas , Histonas/metabolismo , Hipocótilo/crescimento & desenvolvimento , Modelos Biológicos , Mutação/genética , Fenótipo , Receptores de Superfície Celular/metabolismo , Plântula/crescimento & desenvolvimento , Plântula/efeitos da radiação , Transcriptoma/genéticaRESUMO
Circadian clocks sustain 24-h rhythms in physiology and metabolism that are synchronized with the day/night cycle. In plants, the regulatory network responsible for the generation of rhythms has been broadly investigated over the past years. However, little is known about the intersecting pathways that link the environmental signals with rhythms in cellular metabolism. Here, we examine the role of the circadian components REVEILLE8/LHY-CCA1-LIKE5 (RVE8/LCL5) and NIGHT LIGHT-INDUCIBLE AND CLOCK-REGULATED genes (LNK) shaping the diurnal oscillation of the anthocyanin metabolic pathway. Around dawn, RVE8 up-regulates anthocyanin gene expression by directly associating to the promoters of a subset of anthocyanin biosynthetic genes. The up-regulation is overcome at midday by the repressing activity of LNK proteins, as inferred by the increased anthocyanin gene expression in lnk1/lnk2 double mutant plants. Chromatin immunoprecipitation assays using LNK and RVE8 misexpressing plants show that RVE8 binding to target promoters is precluded in LNK overexpressing plants and conversely, binding is enhanced in the absence of functional LNKs, which provides a mechanism by which LNKs antagonize RVE8 function in the regulation of anthocyanin accumulation. Based on their previously described transcriptional coactivating function, our study defines a switch in the regulatory activity of RVE8-LNK interaction, from a synergic coactivating role of evening-expressed clock genes to a repressive antagonistic function modulating anthocyanin biosynthesis around midday.
Assuntos
Antocianinas/biossíntese , Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Ritmo Circadiano , Fatores de Transcrição/metabolismo , Arabidopsis/genética , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Regiões Promotoras Genéticas , Ligação Proteica/genética , Fatores de Tempo , Regulação para Cima/genéticaRESUMO
The circadian clock is a cellular time-keeper mechanism that regulates biological rhythms with a period of ~24 h. The circadian rhythms in metabolism, physiology, and development are synchronized by environmental cues such as light and temperature. In plants, proper matching of the internal circadian time with the external environment confers fitness advantages on plant survival and propagation. Accordingly, plants have evolved elaborated regulatory mechanisms that precisely control the circadian oscillations. Transcriptional feedback regulation of several clock components has been well characterized over the past years. However, the importance of additional regulatory mechanisms such as chromatin remodeling, protein complexes, protein phosphorylation, and stability is only starting to emerge. The multiple layers of circadian regulation enable plants to properly synchronize with the environmental cycles and to fine-tune the circadian oscillations. This review focuses on the diverse posttranslational events that regulate circadian clock function. We discuss the mechanistic insights explaining how plants articulate a high degree of complexity in their regulatory networks to maintain circadian homeostasis and to generate highly precise waveforms of circadian expression and activity.
Assuntos
Arabidopsis/genética , Ritmo Circadiano/genética , Regulação da Expressão Gênica de Plantas , Processamento de Proteína Pós-Traducional , Arabidopsis/metabolismo , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Modelos Genéticos , Fosforilação , Proteólise , UbiquitinaçãoRESUMO
Circadian rhythms allow organisms to time biological processes to the most appropriate phases of the day-night cycle. Post-transcriptional regulation is emerging as an important component of circadian networks, but the molecular mechanisms linking the circadian clock to the control of RNA processing are largely unknown. Here we show that PROTEIN ARGININE METHYL TRANSFERASE 5 (PRMT5), which transfers methyl groups to arginine residues present in histones and Sm spliceosomal proteins, links the circadian clock to the control of alternative splicing in plants. Mutations in PRMT5 impair several circadian rhythms in Arabidopsis thaliana and this phenotype is caused, at least in part, by a strong alteration in alternative splicing of the core-clock gene PSEUDO RESPONSE REGULATOR 9 (PRR9). Furthermore, genome-wide studies show that PRMT5 contributes to the regulation of many pre-messenger-RNA splicing events, probably by modulating 5'-splice-site recognition. PRMT5 expression shows daily and circadian oscillations, and this contributes to the mediation of the circadian regulation of expression and alternative splicing of a subset of genes. Circadian rhythms in locomotor activity are also disrupted in dart5-1, a mutant affected in the Drosophila melanogaster PRMT5 homologue, and this is associated with alterations in splicing of the core-clock gene period and several clock-associated genes. Our results demonstrate a key role for PRMT5 in the regulation of alternative splicing and indicate that the interplay between the circadian clock and the regulation of alternative splicing by PRMT5 constitutes a common mechanism that helps organisms to synchronize physiological processes with daily changes in environmental conditions.
Assuntos
Processamento Alternativo/genética , Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Relógios Circadianos/fisiologia , Ritmo Circadiano/fisiologia , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/fisiologia , Proteínas Metiltransferases/metabolismo , Proteína-Arginina N-Metiltransferases/metabolismo , Animais , Arabidopsis/enzimologia , Arabidopsis/genética , Arabidopsis/efeitos da radiação , Proteínas de Arabidopsis/genética , Sequência de Bases , Relógios Circadianos/genética , Ritmo Circadiano/genética , Escuridão , Proteínas de Drosophila/genética , Drosophila melanogaster/enzimologia , Drosophila melanogaster/genética , Drosophila melanogaster/efeitos da radiação , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Luz , Metilação , Mutação , Proteínas Circadianas Period/genética , Fenótipo , Proteínas Metiltransferases/genética , Proteína-Arginina N-Metiltransferases/genética , Precursores de RNA/genética , Precursores de RNA/metabolismo , Sítios de Splice de RNA/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Spliceossomos/metabolismo , Fatores de Transcrição/genéticaRESUMO
Light signaling pathways and the circadian clock interact to help organisms synchronize physiological and developmental processes with periodic environmental cycles. The plant photoreceptors responsible for clock resetting have been characterized, but signaling components that link the photoreceptors to the clock remain to be identified. Here we describe a family of night light-inducible and clock-regulated genes (LNK) that play a key role linking light regulation of gene expression to the control of daily and seasonal rhythms in Arabidopsis thaliana. A genomewide transcriptome analysis revealed that most light-induced genes respond more strongly to light during the subjective day, which is consistent with the diurnal nature of most physiological processes in plants. However, a handful of genes, including the homologous genes LNK1 and LNK2, are more strongly induced by light in the middle of the night, when the clock is most responsive to this signal. Further analysis revealed that the morning phased LNK1 and LNK2 genes control circadian rhythms, photomorphogenic responses, and photoperiodic dependent flowering, most likely by regulating a subset of clock and flowering time genes in the afternoon. LNK1 and LNK2 themselves are directly repressed by members of the TIMING OF CAB1 EXPRESSION/PSEUDO RESPONSE REGULATOR family of core-clock genes in the afternoon and early night. Thus, LNK1 and LNK2 integrate early light signals with temporal information provided by core oscillator components to control the expression of afternoon genes, allowing plants to keep track of seasonal changes in day length.
Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/fisiologia , Relógios Circadianos/fisiologia , Regulação da Expressão Gênica de Plantas/fisiologia , Transdução de Sinal Luminoso/fisiologia , Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Imunoprecipitação da Cromatina , Relógios Circadianos/genética , Primers do DNA/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas/genética , Sequenciamento de Nucleotídeos em Larga Escala , Transdução de Sinal Luminoso/genética , Análise em Microsséries , Fotoperíodo , Filogenia , Reação em Cadeia da Polimerase em Tempo Real , Estações do Ano , Fatores de Transcrição/metabolismoRESUMO
Circadian clocks are endogenous mechanisms that translate environmental cues into temporal information to generate the 24-h rhythms in metabolism and physiology. The circadian function relies on the precise regulation of rhythmic gene expression at the core of the oscillator, which temporally modulates the genome transcriptional activity in virtually all multicellular organisms examined to date. Emerging evidence in plants suggests a highly sophisticated interplay between the circadian patterns of gene expression and the rhythmic changes in chromatin remodeling and histone modifications. Alternative precursor messenger RNA (pre-mRNA) splicing has also been recently defined as a fundamental pillar within the circadian system, providing the required plasticity and specificity for fine-tuning the circadian clock. This review highlights the relationship between the plant circadian clock with both chromatin remodeling and alternative splicing and compares the similarities and divergences with analogous studies in animal circadian systems.
Assuntos
Processamento Alternativo , Proteínas de Arabidopsis/genética , Arabidopsis/fisiologia , Montagem e Desmontagem da Cromatina , Cromatina , Relógios Circadianos/fisiologia , Regulação da Expressão Gênica de Plantas , RNA Mensageiro/genética , Proteínas de Arabidopsis/metabolismo , Histonas/genética , Histonas/metabolismo , Fotoperíodo , RNA Mensageiro/metabolismo , Transdução de Sinais , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Transcrição GênicaRESUMO
Circadian clock function in Arabidopsis thaliana relies on a complex network of reciprocal regulations among oscillator components. Here, we demonstrate that chromatin remodeling is a prevalent regulatory mechanism at the core of the clock. The peak-to-trough circadian oscillation is paralleled by the sequential accumulation of H3 acetylation (H3K56ac, K9ac), H3K4 trimethylation (H3K4me3), and H3K4me2. Inhibition of acetylation and H3K4me3 abolishes oscillator gene expression, indicating that both marks are essential for gene activation. Mechanistically, blocking H3K4me3 leads to increased clock-repressor binding, suggesting that H3K4me3 functions as a transition mark modulating the progression from activation to repression. The histone methyltransferase SET DOMAIN GROUP 2/ARABIDOPSIS TRITHORAX RELATED 3 (SDG2/ATXR3) might contribute directly or indirectly to this regulation because oscillator gene expression, H3K4me3 accumulation, and repressor binding are altered in plants misexpressing SDG2/ATXR3. Despite divergences in oscillator components, a chromatin-dependent mechanism of clock gene activation appears to be common to both plant and mammal circadian systems.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Relógios Circadianos , Histonas/metabolismo , Processamento de Proteína Pós-Traducional , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Relógios Circadianos/genética , Regulação da Expressão Gênica de Plantas , Genes de Plantas/genética , Histona Metiltransferases , Histona-Lisina N-Metiltransferase/metabolismo , Lisina/metabolismo , Metilação , Ligação Proteica/genética , Processamento de Proteína Pós-Traducional/genética , Proteínas Repressoras/metabolismoRESUMO
The increasing development of novel methods and techniques facilitates the measurement of high-dimensional time series but challenges our ability for accurate modeling and predictions. The use of a general mathematical model requires the inclusion of many parameters, which are difficult to be fitted for relatively short high-dimensional time series observed. Here, we propose a novel method to accurately model a high-dimensional time series. Our method extends the barycentric coordinates to high-dimensional phase space by employing linear programming, and allowing the approximation errors explicitly. The extension helps to produce free-running time-series predictions that preserve typical topological, dynamical, and/or geometric characteristics of the underlying attractors more accurately than the radial basis function model that is widely used. The method can be broadly applied, from helping to improve weather forecasting, to creating electronic instruments that sound more natural, and to comprehensively understanding complex biological data.
RESUMO
Research on the functional properties of nucleosome structure and composition dynamics has revealed that chromatin-level regulation is an essential component of light signalling and clock function in plants, two processes that rely extensively on transcriptional controls. In particular, several types of histone post-translational modifications and chromatin-bound factors act sequentially or in combination to establish transcriptional patterns and to fine-tune the transcript abundance of a large repertoire of light-responsive genes and clock components. Cytogenetic approaches have also identified light-induced higher-order chromatin changes that dynamically organize the condensation of chromosomal domains into sub-nuclear foci containing silenced repeat elements. In this review, we report recently identified molecular actors that establish chromatin state dynamics in response to light signals such as photoperiod, intensity, and spectral quality. We also highlight the chromatin-dependent mechanisms that contribute to the 24-h circadian gene expression and its impact on plant physiology and development. The commonalities and contrasts of light- and clock-associated chromatin-based mechanisms are discussed, with particular emphasis on their impact on the selective regulation and rapid modulation of responsive genes.
Assuntos
Cromatina/metabolismo , Relógios Circadianos/efeitos da radiação , Luz , Fenômenos Fisiológicos Vegetais/efeitos da radiação , Plantas/genética , Plantas/efeitos da radiação , Modelos BiológicosRESUMO
For over a decade, the animal field has led the way in using DNA methylation measurements to construct epigenetic clocks of aging. These clocks can predict organismal age with a level of accuracy that surpasses any other molecular proxy known to date. Evidence is finally emerging that epigenetic clocks also exist in plants. However, these clocks appear to differ from those in animals in some key aspects, including in their ability to measure time beyond the life span of an individual. Clock-like epigenetic changes can be found in plant circadian rhythms (scale: 24 h), during plant aging (scale: weeks/centuries), and across plant lineage evolution (scale: decades/millennia). Here, we provide a first classification of these different types of epigenetic clocks, highlight their main features, and discuss their biological basis.
RESUMO
Despite our increasing knowledge on the transcriptional networks connecting abscisic acid (ABA) signalling with the circadian clock, the molecular nodes in which both pathways converge to translate the environmental information into a physiological response are not known. Here, we provide evidence of a feedback mechanism linking the circadian clock with plant responses to drought. A key clock component (TOC1, timing of CAB expression 1) binds to the promoter of the ABA-related gene (ABAR/CHLH/GUN5) and controls its circadian expression. TOC1 is in turn acutely induced by ABA and this induction advances the phase of TOC1 binding and modulates ABAR circadian expression. Moreover, the gated induction of TOC1 by ABA is abolished in ABAR RNAi plants suggesting that the reciprocal regulation between ABAR and TOC1 expression is important for sensitized ABA activity. Genetic studies with TOC1 and ABAR over-expressing and RNAi plants showed defective responses to drought, which support the notion that clock-dependent gating of ABA function is important for cellular homeostasis under dry environments.
Assuntos
Ácido Abscísico/fisiologia , Proteínas de Arabidopsis/fisiologia , Ritmo Circadiano , Secas , Retroalimentação Fisiológica/fisiologia , Regulação da Expressão Gênica de Plantas , Reguladores de Crescimento de Plantas/fisiologia , Estômatos de Plantas/fisiologia , Fatores de Transcrição/fisiologia , Ácido Abscísico/biossíntese , Ácido Abscísico/genética , Arabidopsis/enzimologia , Arabidopsis/genética , Arabidopsis/fisiologia , Proteínas de Arabidopsis/biossíntese , Proteínas de Arabidopsis/genética , Clorofila/genética , Ritmo Circadiano/genética , Liases/genética , Liases/fisiologia , Fenótipo , Fosfoproteínas Fosfatases/genética , Fosfoproteínas Fosfatases/fisiologia , Reguladores de Crescimento de Plantas/biossíntese , Reguladores de Crescimento de Plantas/genética , Estômatos de Plantas/genética , Ligação Proteica/genética , Proteína Fosfatase 2C , Transdução de Sinais/genética , Fatores de Transcrição/biossíntese , Fatores de Transcrição/genéticaRESUMO
Circadian rhythms are daily biological oscillations driven by an endogenous mechanism known as circadian clock. The protein kinase CK2 is one of the few clock components that is evolutionary conserved among different taxonomic groups. CK2 regulates the stability and nuclear localization of essential clock proteins in mammals, fungi, and insects. Two CK2 regulatory subunits, CKB3 and CKB4, have been also linked with the Arabidopsis thaliana circadian system. However, the biological relevance and the precise mechanisms of CK2 function within the plant clockwork are not known. By using ChIP and Double-ChIP experiments together with in vivo luminescence assays at different temperatures, we were able to identify a temperature-dependent function for CK2 modulating circadian period length. Our study uncovers a previously unpredicted mechanism for CK2 antagonizing the key clock regulator CIRCADIAN CLOCK-ASSOCIATED 1 (CCA1). CK2 activity does not alter protein accumulation or subcellular localization but interferes with CCA1 binding affinity to the promoters of the oscillator genes. High temperatures enhance the CCA1 binding activity, which is precisely counterbalanced by the CK2 opposing function. Altering this balance by over-expression, mutation, or pharmacological inhibition affects the temperature compensation profile, providing a mechanism by which plants regulate circadian period at changing temperatures. Therefore, our study establishes a new model demonstrating that two opposing and temperature-dependent activities (CCA1-CK2) are essential for clock temperature compensation in Arabidopsis.
Assuntos
Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimologia , Arabidopsis/genética , Caseína Quinase II/metabolismo , Relógios Circadianos/genética , Regulação da Expressão Gênica de Plantas , Temperatura , Fatores de Transcrição/genética , Núcleo Celular/enzimologia , Flores/fisiologia , Genes de Plantas/genética , Hipocótilo/crescimento & desenvolvimento , Fosforilação , Regiões Promotoras Genéticas/genética , Ligação Proteica , Isoformas de Proteínas/metabolismo , Fatores de Tempo , Fatores de Transcrição/metabolismo , Transcrição GênicaRESUMO
Despite our increasing understanding of the molecular determinants essential for circadian clock function, we still lack a complete picture of the mechanisms contributing to clock progression in plants. Here, we explore the role of REVEILLE8/LHY-CCA1-LIKE5 (RVE8/LCL5) within the Arabidopsis circadian system. RVE8/LCL5 encodes a MYB-like transcription factor similar to CIRCADIAN CLOCK-ASSOCIATED1 (CCA1) and ELONGATED HYPOCOTYL (LHY), which are essential regulators of the Arabidopsis circadian clock. Consistent with the sequence similarity, the rhythmic expression of RVE8/LCL5 shows a morning acrophase comparable to that of CCA1 and LHY. Plants mis-expressing RVE8/LCL5 display a variety of circadian phenotypes, including altered circadian gene expression and photoperiodic flowering time. Similar to CCA1, RVE8/LCL5 regulates the expression of the oscillator gene TOC1 (TIMING OF CAB EXPRESSION1) by associating with the TOC1 promoter and by modulating the pattern of histone 3 (H3) acetylation. However, the mechanisms of RVE8/LCL5 and CCA1 activity in this regulation differ markedly. Indeed, the use of chromatin immunoprecipitation and pharmacological inhibition assays reveals that RVE8/LCL5 favours a hyper-acetylated state of H3 at the TOC1 promoter, which may facilitate the rising phase of TOC1. In contrast, CCA1 represses TOC1 expression by promoting histone deacetylation. Thus, despite the sequence homology and the similar morning phase of expression, RVE8/LCL5 and CCA1 have opposing regulatory functions within the Arabidopsis circadian clock, although CCA1 has a more predominant role. We propose that contrasting chromatin compaction and transcriptional modulation through the opposing activities of RVE8/LCL5 and CCA1 might provide a fine-tuning mechanism for precisely shaping the TOC1 circadian waveform in Arabidopsis.
Assuntos
Ritmo Circadiano , Acetilação , Arabidopsis/genética , Arabidopsis/metabolismo , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Flores/fisiologia , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Histonas/metabolismo , Ácidos Hidroxâmicos/farmacologia , Mutagênese Insercional , Fotoperíodo , Regiões Promotoras Genéticas , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Our understanding of the circadian clock function in plants has been markedly assisted by studies with the model species Arabidopsis thaliana. Molecular and genetics approaches have delivered a comprehensive view of the transcriptional regulatory networks underlying the Arabidopsis circadian system. The use of the luciferase as a reporter allowed the precise in vivo determination of circadian periods, phases, and amplitudes of clock promoter activities with unprecedented temporal resolution. An increasing repertoire of fine-tuned luciferases together with additional applications such as translational fusions or bioluminescence molecular complementation assays have considerably expanded our view of circadian protein expression and activity, far beyond transcriptional regulation. Further applications have focused on the in vivo simultaneous examination of rhythms in different parts of the plant. The use of intact versus excised plant organs has also provided a glimpse on both the organ-specific and autonomy of the clocks and the importance of long distance communication for circadian function. This chapter provides a basic protocol for in vivo high-throughput monitoring of circadian rhythms in Arabidopsis seedlings using bioluminescent reporters and a microplate luminometer.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Relógios Circadianos , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Ritmo Circadiano/fisiologia , Regulação da Expressão Gênica de Plantas , Luciferases/genética , Luciferases/metabolismoRESUMO
The plant circadian clock regulates essential biological processes including flowering time or petal movement. However, little is known about how the clock functions in flowers. Here, we identified the circadian components and transcriptional networks contributing to the generation of rhythms in pistils, the female reproductive organ. When detached from the rest of the flower, pistils sustain highly precise rhythms, indicating organ-specific circadian autonomy. Analyses of clock mutants and chromatin immunoprecipitation assays showed distinct expression patterns and specific regulatory functions for clock activators and repressors in pistils. Genetic interaction studies also suggested a hierarchy of the repressing activities that provide robustness and precision to the pistil clock. Globally, the circadian function in pistils primarily governs responses to environmental stimuli and photosynthesis and controls pistil growth and seed weight and production. Understanding the circadian intricacies in reproductive organs may prove useful for optimizing plant reproduction and productivity.