RESUMO
Light is both the main source of energy and a key environmental signal for plants. It regulates not only gene expression but also the tightly related processes of splicing and alternative splicing (AS). Two main pathways have been proposed to link light sensing with the splicing machinery. One occurs through a photosynthesis-related signal, and the other is mediated by photosensory proteins, such as red light-sensing phytochromes. Here, we evaluated the relative contribution of each of these pathways by performing a transcriptome-wide analysis of light regulation of AS in plants that do not express any functional phytochrome (phyQ). We found that an acute 2-h red-light pulse in the middle of the night induces changes in the splicing patterns of 483 genes in wild-type plants. Approximately 30% of these genes also showed strong light regulation of splicing patterns in phyQ mutant plants, revealing that phytochromes are important but not essential for the regulation of AS by R light. We then performed a meta-analysis of related transcriptomic datasets and found that different light regulatory pathways can have overlapping targets in terms of AS regulation. All the evidence suggests that AS is regulated simultaneously by various light signaling pathways, and the relative contribution of each pathway is highly dependent on the plant developmental stage.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Fitocromo , Arabidopsis/genética , Arabidopsis/metabolismo , Fitocromo/genética , Fitocromo/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Processamento Alternativo/genética , Splicing de RNA , Plantas/metabolismoRESUMO
Red/far-red light-sensing bacteriophytochrome photoreceptor (BphP) pathways play key roles in bacterial physiology and ecology. These bilin-binding proteins photoswitch between two states, Pr (red absorbing) and Pfr (far-red absorbing). The isomerization of the chromophore and the downstream structural changes result in the light signal transduction. The agricultural pathogen Xanthomonas campestris pv. campestris (Xcc) code for a single bathy-like type BphP (XccBphP), previously shown to negatively regulate several light-mediated biological processes involved in virulence. Here, we generated three different full-length variants with single amino acid changes within its GAF domain that affect the XccBphP photocycle favouring its Pr state: L193Q, L193N and D199A. While D199A recombinant protein locks XccBphP in a Pr-like state, L193Q and L193N exhibit a significant enrichment of the Pr form in thermal equilibrium. The X-ray crystal structures of the three variants were solved, resembling the wild-type protein in the Pr state. Finally, we studied the effects of altering the XccBphP photocycle on the exopolysaccharide xanthan production and stomatal aperture assays as readouts of its bacterial signalling pathway. Null-mutant complementation assays show that the photoactive Pr-favoured XccBphP variants L193Q and L193N tend to negatively regulate xanthan production in vivo. In addition, our results indicate that strains expressing these variants also promote stomatal apertures in challenged plant epidermal peels, compared to wild-type Xcc. The findings presented in this work provide new evidence on the Pr state of XccBphP as a negative regulator of the virulence-associated mechanisms by light in Xcc.
Assuntos
Arabidopsis/microbiologia , Pigmentos Biliares/metabolismo , Fitocromo/química , Fitocromo/genética , Doenças das Plantas/microbiologia , Virulência , Xanthomonas campestris/fisiologia , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Cristalografia por Raios X , Luz , Modelos Moleculares , Mutagênese Sítio-Dirigida , Mutação , Fitocromo/metabolismoRESUMO
Light cues from neighboring vegetation rapidly initiate plant shade-avoidance responses. Despite our detailed knowledge of the early steps of this response, the molecular events under prolonged shade are largely unclear. Here we show that persistent neighbor cues reinforce growth responses in addition to promoting auxin-responsive gene expression in Arabidopsis and soybean. However, while the elevation of auxin levels is well established as an early event, in Arabidopsis, the response to prolonged shade occurs when auxin levels have declined to the prestimulation values. Remarkably, the sustained low activity of phytochrome B under prolonged shade led to (i) decreased levels of PHYTOCHROME INTERACTING FACTOR 4 (PIF4) in the cotyledons (the organs that supply auxin) along with increased levels in the vascular tissues of the stem, (ii) elevated expression of the PIF4 targets INDOLE-3-ACETIC ACID 19 (IAA19) and IAA29, which in turn reduced the expression of the growth-repressive IAA17 regulator, (iii) reduced abundance of AUXIN RESPONSE FACTOR 6, (iv) reduced expression of MIR393 and increased abundance of its targets, the auxin receptors, and (v) elevated auxin signaling as indicated by molecular markers. Mathematical and genetic analyses support the physiological role of this system-level rearrangement. We propose that prolonged shade rewires the connectivity between light and auxin signaling to sustain shade avoidance without enhanced auxin levels.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Ácidos Indolacéticos/farmacologia , Luz , Fitocromo/metabolismo , Fenômenos Fisiológicos Vegetais , Arabidopsis/efeitos dos fármacos , Arabidopsis/crescimento & desenvolvimento , Proteínas de Arabidopsis/genética , Fitocromo/genética , Reguladores de Crescimento de Plantas/farmacologia , Transdução de SinaisRESUMO
The growing collection of sequenced or genotyped Arabidopsis thaliana accessions includes mostly individuals from the native Eurasian and N. African range and introduced North American populations. Here, we describe the genetic and phenotypic diversity, along with habitats and life history, of A. thaliana plants collected at the southernmost end of its worldwide distribution. Seed samples were harvested from plants growing in four sites within a ~3500-km2 -area in Patagonia, Argentina, and represent the first germplasm to be collected in South America for this species. Whole-genome resequencing revealed that plants from the four sites and a Patagonia herbarium specimen collected in 1967 formed a single haplogroup (Pat), indicating that the phenotypic variation observed in the field reflected plastic responses to the environment. admixture and principal components analyses suggest that the ancestor of the Pat haplogroup either came from Italy or the Balkan/Caucasus regions of Eurasia. In the laboratory, plants from the Pat haplogroup were hyposensitive to continuous red (Rc) and shade light, with corresponding changes in the expression of phytochrome signalling genes. Pat had higher PIF3 and PIF5 and lower HY5 expression under Rc light; and lower expression of PIL1, ATHB2 and HFR1 under shade compared to Col-0. In addition, Pat plants had a strong vernalization requirement associated with high levels of FLC expression. We conclude that including Pat in studies of natural variation and in comparison with other introduced populations will provide additional information for association studies and allow for a more detailed assessment of the demographic events following colonization.
Assuntos
Arabidopsis/genética , Genética Populacional , Haplótipos , Luz , Adaptação Fisiológica , Arabidopsis/efeitos da radiação , Argentina , Regulação da Expressão Gênica de Plantas , Fenótipo , Fitocromo/genéticaRESUMO
Plants have developed sophisticated systems to monitor and rapidly acclimate to environmental fluctuations. Light is an essential source of environmental information throughout the plant's life cycle. The model plant Arabidopsis thaliana possesses five phytochromes (phyA-phyE) with important roles in germination, seedling establishment, shade avoidance, and flowering. However, our understanding of the phytochrome signaling network is incomplete, and little is known about the individual roles of phytochromes and how they function cooperatively to mediate light responses. Here, we used a bottom-up approach to study the phytochrome network. We added each of the five phytochromes to a phytochrome-less background to study their individual roles and then added the phytochromes by pairs to study their interactions. By analyzing the 16 resulting genotypes, we revealed unique roles for each phytochrome and identified novel phytochrome interactions that regulate germination and the onset of flowering. Furthermore, we found that ambient temperature has both phytochrome-dependent and -independent effects, suggesting that multiple pathways integrate temperature and light signaling. Surprisingly, none of the phytochromes alone conferred a photoperiodic response. Although phyE and phyB were the strongest repressors of flowering, both phyB and phyC were needed to confer a flowering response to photoperiod. Thus, a specific combination of phytochromes is required to detect changes in photoperiod, whereas single phytochromes are sufficient to respond to light quality, indicating how phytochromes signal different light cues.
Assuntos
Apoproteínas/genética , Proteínas de Arabidopsis/genética , Arabidopsis/genética , Fitocromo B/genética , Fitocromo/genética , Apoproteínas/metabolismo , Arabidopsis/crescimento & desenvolvimento , Proteínas de Arabidopsis/metabolismo , Genótipo , Germinação/genética , Luz , Fitocromo/metabolismo , Fitocromo A , Fitocromo B/metabolismo , Folhas de Planta/genética , Folhas de Planta/crescimento & desenvolvimento , Plântula/genética , Plântula/crescimento & desenvolvimento , Transdução de Sinais/genética , TemperaturaRESUMO
In several species, seed germination is regulated by light in a way that restricts seedling emergence to the environmental conditions that are likely to be favourable for the success of the new individual, and therefore, this behaviour is recognized to have adaptive value. The phytochromes are one of the most relevant photoreceptors involved in light perception by plants. We explored the redundancy and diversity functions of the phytochrome family in the control of seed responsiveness to light and gibberellins (GA) by using a set of phytochrome mutants of Arabidopsis. Our data show that, in addition to the well-known role of phyB in the promotion of germination in response to high red to far-red ratios (R/FR), phyE and phyD stimulate germination at very low R/FR ratios, probably by promoting the action of phyA. Further, we show that phyC regulates negatively the seed responsiveness to light, unravelling unexpected functions for phyC in seed germination. Finally, we find that seed responsiveness to GA is mainly controlled by phyB, with phyC, phyD and phyE having relevant roles when acting in a phyB-deficient background. Our results indicate that phytochromes have multiple and complex roles during germination depending on the active photoreceptor background.
Assuntos
Arabidopsis/crescimento & desenvolvimento , Arabidopsis/genética , Germinação/efeitos da radiação , Giberelinas/farmacologia , Luz , Família Multigênica , Fitocromo/genética , Arabidopsis/efeitos dos fármacos , Arabidopsis/efeitos da radiação , Germinação/efeitos dos fármacos , Modelos Biológicos , Mutação/genética , Fitocromo/metabolismo , Sementes/efeitos dos fármacos , Sementes/crescimento & desenvolvimento , Sementes/efeitos da radiaçãoRESUMO
Ever since the discovery of the role of bacteriophytochrome (BphP) in inducing carotenoid synthesis in Deinococcus radiodurans in response to light the role of BphPs in other non-photosynthetic bacteria is not clear yet. Azospirillum brasilense, a non-photosynthetic rhizobacterium, harbours a pair of BphPs out of which AbBphP1 is a homolog of AtBphP1 of Agrobacterium tumefaciens. By overexpression, purification, biochemical and spectral characterization we have shown that AbBphP1 is a photochromic bacteriophytochrome. Phenotypic study of the ΔAbBphP1 mutant showed that it is required for the survival of A. brasilense on minimal medium under red light. The mutant also showed reduced chemotaxis towards dicarboxylates and increased sensitivity to the photooxidative stress. Unlike D. radiodurans, AbBphP1 was not involved in controlling carotenoid synthesis. Proteome analysis of the ΔAbBphP1 indicated that AbBphP1 is involved in inducing a cellular response that enables A. brasilense in regenerating proteins that might be damaged due to photodynamic stress.
Assuntos
Azospirillum brasilense/metabolismo , Proteínas de Bactérias/metabolismo , Carotenoides/metabolismo , Fitocromo/metabolismo , Azospirillum brasilense/crescimento & desenvolvimento , Proteínas de Bactérias/classificação , Proteínas de Bactérias/genética , Dimerização , Luz , Mutação , Filogenia , Fitocromo/classificação , Fitocromo/genética , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , Cloreto de Tolônio/químicaRESUMO
A common molecular regulatory pathway that involves PHYA, PHYB and floral integrator genes CONSTANS (CO), FLOWERING LOCUS T (FT) and SUPRESSOR OF OVEREXPRESSION OF CO1 (SOC1) has been suggested to participate in the regulation of photoperiod dependent processes such as flowering and dormancy. In grapevines (Vitis vinifera L.), decreasing photoperiod and low temperatures trigger the transition of buds into endodormancy (ED), a process that is accompanied by drastic changes in gene expression of VvPHYA and B in leaves. To analyse the relationship of VvPHYA, VvPHYB, and grape homologues of Arabidopsis floral integrator genes VvCO, VvFT, VvMADS8, with ED, a comparative expression analysis of these genes was performed in grapevine-leaves and buds before, during and after the transition of buds into ED. The expression of all the above genes in the bud-tissue, and the fact that photoperiod regulates differently the expression of VvPHYA and B in buds than in leaves, suggests that the bud might be an autonomous or semi-autonomous organ in perceiving and transducing the photoperiod signal. On the other hand, the coordinated down-regulation of VvFT in leaves and buds during the transition of buds into ED, and its subsequent up-regulation following the application of dormancy-breaking compounds, hydrogen cyanamide (HC) and sodium azide, suggests that VvFT could play a key role in stimulating bud-growth by repressing their entry into ED.
Assuntos
Cianamida/farmacologia , Flores/crescimento & desenvolvimento , Fitocromo/genética , Proteínas de Plantas/genética , Azida Sódica/farmacologia , Vitis/genética , Temperatura Baixa , Regulação para Baixo/genética , Flores/genética , Flores/fisiologia , Regulação da Expressão Gênica no Desenvolvimento , Regulação da Expressão Gênica de Plantas , Genes de Plantas/genética , Especificidade de Órgãos , Fotoperíodo , Fitocromo/metabolismo , Folhas de Planta/genética , Proteínas de Plantas/metabolismo , Regiões Promotoras Genéticas , RNA de Plantas/genética , Transdução de Sinais , Regulação para Cima/genética , Vitis/crescimento & desenvolvimento , Vitis/fisiologiaRESUMO
Plants use light as a source of energy for photosynthesis and as a source of environmental information perceived by photoreceptors. Testing whether plants can complete their cycle if light provides energy but no information about the environment requires a plant devoid of phytochromes because all photosynthetically active wavelengths activate phytochromes. Producing such a quintuple mutant of Arabidopsis thaliana has been challenging, but we were able to obtain it in the flowering locus T (ft) mutant background. The quintuple phytochrome mutant does not germinate in the FT background, but it germinates to some extent in the ft background. If germination problems are bypassed by the addition of gibberellins, the seedlings of the quintuple phytochrome mutant exposed to red light produce chlorophyll, indicating that phytochromes are not the sole red-light photoreceptors, but they become developmentally arrested shortly after the cotyledon stage. Blue light bypasses this blockage, rejecting the long-standing idea that the blue-light receptors cryptochromes cannot operate without phytochromes. After growth under white light, returning the quintuple phytochrome mutant to red light resulted in rapid senescence of already expanded leaves and severely impaired expansion of new leaves. We conclude that Arabidopsis development is stalled at several points in the presence of light suitable for photosynthesis but providing no photomorphogenic signal.
Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Mutação , Fitocromo/genética , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Clorofila/metabolismo , Ritmo Circadiano , Germinação/efeitos dos fármacos , Germinação/efeitos da radiação , Giberelinas/farmacologia , Luz , Morfogênese/efeitos dos fármacos , Morfogênese/efeitos da radiação , Fototropismo/efeitos dos fármacos , Fototropismo/efeitos da radiação , Reguladores de Crescimento de Plantas/farmacologia , Folhas de Planta/genética , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/metabolismo , Plântula/genética , Plântula/crescimento & desenvolvimento , Plântula/metabolismo , Sementes/genética , Sementes/crescimento & desenvolvimentoRESUMO
Despite the crucial role that phytochromes (Phys) play in light perception and in the entrainment of the circadian clock to local time, the photoperiodic regulation of PHYA and PHYB gene expression has been poorly studied, especially in woody perennials. Here the dynamic of Vitis vinifera PHYA (VvPHYA) and PHYB (VvPHYB) transcript accumulation was studied in field-grown grapevine leaves throughout daily cycles under decreasing natural photoperiods. Given that in grapevine the entrance of buds into endodormancy (ED) is a photoperiod-driven process, increases in BR(50) values, a parameter that measures the depth of dormancy in single bud cuttings assays was used to determine the critical daylength at which grapevine discriminates between long day (LD) and short day (SD) photoperiod. Therefore, we monitored the daily expression profile of VvPHYA and VvPHYB transcripts before, during and after the defined critical daylength. Results showed that under LD photoperiod (21 December, daylength 14 h 40 min) the abundance of both transcripts oscillated with diurnal rhythms, attaining maximum and minimum levels before dawn and after dusk, respectively. However, under SD photoperiod (12 April, daylength 11 h 40 min) the rhythmic expression disappeared, and both transcripts expressed uniformly at high levels. Our results showing that photoperiod regulates VvPHYA and VvPHYB gene expression, contrast with those reported in Arabidopsis and in other herbaceous plants in which PHYA and PHYB gene expression is regulated by the circadian clock.