RESUMO
Understanding the mechanisms regulating root development under drought conditions is an important question for plant biology and world agriculture. We examine the effect of osmotic stress on abscisic acid (ABA), cytokinin and ethylene responses and how they mediate auxin transport, distribution and root growth through effects on PIN proteins. We integrate experimental data to construct hormonal crosstalk networks to formulate a systems view of root growth regulation by multiple hormones. Experimental analysis shows: that ABA-dependent and ABA-independent stress responses increase under osmotic stress, but cytokinin responses are only slightly reduced; inhibition of root growth under osmotic stress does not require ethylene signalling, but auxin can rescue root growth and meristem size; osmotic stress modulates auxin transporter levels and localization, reducing root auxin concentrations; PIN1 levels are reduced under stress in an ABA-dependent manner, overriding ethylene effects; and the interplay among ABA, ethylene, cytokinin and auxin is tissue-specific, as evidenced by differential responses of PIN1 and PIN2 to osmotic stress. Combining experimental analysis with network construction reveals that ABA regulates root growth under osmotic stress conditions via an interacting hormonal network with cytokinin, ethylene and auxin.
Assuntos
Ácido Abscísico/metabolismo , Arabidopsis/metabolismo , Etilenos/metabolismo , Ácidos Indolacéticos/metabolismo , Raízes de Plantas/crescimento & desenvolvimento , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Citocininas/metabolismo , Regulação da Expressão Gênica de Plantas , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Meristema/crescimento & desenvolvimento , Meristema/metabolismo , Pressão Osmótica , Raízes de Plantas/genética , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas , Plântula/metabolismo , Transdução de SinaisRESUMO
Patterning in Arabidopsis root development is coordinated via a localized auxin concentration maximum in the root tip, requiring the regulated expression of specific genes. However, little is known about how hormone and gene expression patterning is generated. Using a variety of experimental data, we develop a spatiotemporal hormonal crosstalk model that describes the integrated action of auxin, ethylene and cytokinin signalling, the POLARIS protein, and the functions of PIN and AUX1 auxin transporters. We also conduct novel experiments to confirm our modelling predictions. The model reproduces auxin patterning and trends in wild-type and mutants; reveals that coordinated PIN and AUX1 activities are required to generate correct auxin patterning; correctly predicts shoot to root auxin flux, auxin patterning in the aux1 mutant, the amounts of cytokinin, ethylene and PIN protein, and PIN protein patterning in wild-type and mutant roots. Modelling analysis further reveals how PIN protein patterning is related to the POLARIS protein through ethylene signalling. Modelling prediction of the patterning of POLARIS expression is confirmed experimentally. Our combined modelling and experimental analysis reveals that a hormonal crosstalk network regulates the emergence of patterns and levels of hormones and gene expression in wild-type and mutants.
Assuntos
Arabidopsis/genética , Padronização Corporal/genética , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Mutação/genética , Reguladores de Crescimento de Plantas/farmacologia , Raízes de Plantas/genética , Análise Espaço-Temporal , Arabidopsis/efeitos dos fármacos , Proteínas de Arabidopsis/metabolismo , Padronização Corporal/efeitos dos fármacos , Citocininas/metabolismo , Etilenos/metabolismo , Ácidos Indolacéticos/metabolismo , Modelos Biológicos , Raízes de Plantas/efeitos dos fármacos , Raízes de Plantas/crescimento & desenvolvimentoRESUMO
The ABACUS1-2 µ (ABscisic Acid Concentration and Uptake Sensor 1-2 µ) and GPS1 (Gibberellin Perception Sensor 1) are direct Förster resonance energy transfer (FRET) biosensors that can be used to measure hormone levels in planta. We provide detailed protocols to image FRET biosensors under exogenously applied hormones in roots, either as a single time point or for treatment time courses before and after hormone application. A new, free, open-source analysis toolset for Fiji is introduced and used to get full 3D segmentation of images of nuclear localized FRET biosensors and calculate emission ratios on a per nucleus basis allowing in-depth analysis of biosensor data.
Assuntos
Técnicas Biossensoriais , Transferência Ressonante de Energia de Fluorescência , Técnicas Biossensoriais/métodos , Transferência Ressonante de Energia de Fluorescência/métodos , Giberelinas , Hormônios , Reguladores de Crescimento de PlantasRESUMO
In recent years, plant biologists interested in quantifying molecules and molecular events in vivo have started to complement reporter systems with genetically encoded fluorescent biosensors (GEFBs) that directly sense an analyte. Such biosensors can allow measurements at the level of individual cells and over time. This information is proving valuable to mathematical modellers interested in representing biological phenomena in silico, because improved measurements can guide improved model construction and model parametrisation. Advances in synthetic biology have accelerated the pace of biosensor development, and the simultaneous expression of spectrally compatible biosensors now allows quantification of multiple nodes in signalling networks. For biosensors that directly respond to stimuli, targeting to specific cellular compartments allows the observation of differential accumulation of analytes in distinct organelles, bringing insights to reactive oxygen species/calcium signalling and photosynthesis research. In conjunction with improved image analysis methods, advances in biosensor imaging can help close the loop between experimentation and mathematical modelling.