Final Report Summary - SMARTPLANT (Environmental control of carotenoid biosynthesis: a novel strategy to improve photosynthetic capacity)
Carotenoids are pigments of isoprenoid nature essential to protect the photosynthetic apparatus against irreversible photodestruction. They also act as light harvesting agents, supplementing the light capturing ability of chlorophyll. Beyond their critical roles in photosynthesis carotenoids are known to be important precursors of plant hormones (e.g. absicisc acid and strigolactones) as well as key regulators of growth and stress responses.
Carotenoid biosynthesis is tightly linked to photosynthesis and is therefore a key determinant of plant biomass and agricultural productivity. Carotenoid production is also strongly regulated by light and temperature, so improved knowledge of these regulatory pathways is fundamental to design new strategies to optimize carotenoid production in a changing environment. In SmartPlant we aimed to determine the molecular pathway that drives environmental control of carotenoid production. We identified a major regulatory module through which light and temperature signals converge to control carotenoid biosynthesis. We determined the functional properties of this molecular mechanism and its influence on carotenoid pathway dynamics.
The ability to sense and integrate light and temperature signals is essential for plant biomass production. The phytochrome red and far-red light photoreceptors are central molecular regulators of plasticity, altering plant growth and biomass build-up according to the light environment. Recent work has shown that the light pathways also act as a conduit for temperature signals, suggesting that these environmental cues utilize common pathways to regulate plant growth and development. Within the phytochrome-mediated changes during this critical time is the assembly of the photosynthetic apparatus, and the production of the photosynthetic pigments (chlorophylls/carotenoids). Carotenoids are integral accessory pigments in the light-absorbing antenna. During greening, carotenoids play an essential photoprotective role minimizing the potentially damaging effects of light on the photosynthetic apparatus. Therefore, to maximize light capture but minimize the damaging effects of light over the emerging photosynthetic machinery, the production of carotenoids and chlorophylls has to take place in a tightly controlled/interdependent manner. However, we do not currently understand how this regulation takes place. Improved knowledge of these regulatory pathways is fundamental to design new strategies to optimize carotenoid production.
In our project we aimed to define the roles of key phytochrome signaling components in photoprotection and maintenance of photosynthetic capacity over a range of light and temperature conditions that are experienced in nature. Experimental and modeling approaches were combined to determine mechanistic insights.
We previously showed that the Phytochrome Interacting Factors (PIFs) repress carotenoid accumulation by directly down-regulating the expression of the gene encoding for PHYTOENE SYNTHASE (PSY), the main rate-determining enzyme in carotenogenesis. Recent studies have shown that PIFs also participate in temperature signaling. During our characterization of PIFs in carotenogenesis we postulated the existence of an unidentified PSY-activator that becomes more effective with PIF depletion. Our analysis showed the bZIP transcription factor Long Hypocotyl 5 (HY5) fulfilled this role, operating as a potent PIF antagonist in the control of carotenoid accumulation. HY5 is required for the light induction of PSY, and carotenoid and chlorophyll synthesis. In chromatin immunoprecipitation assays we demonstrated that PIFs and HY5 target directly a common cis-element (G-box). We assessed the role of temperature in the PIFs-HY5 mediated control of photopigment content (Figure 1A). Our current results indicate that carotenoid and chlorophyll levels increase with temperature. PIFs are necessary to maintain control of photopigment levels over a temperature range. However, HY5 impact is more relevant at cooler temperatures (Figure 1B). In adult plants we demonstrated that HY5 is required to increase chlorophyll synthesis and carbon uptake in the cool (Figure 1C). Indeed, HY5 is required for dynamic light and temperature induced photopigment adjustments over the life span of the plant. Because of this the hy5 null mutant has lower carbon assimilation and a marked reduction in biomass at lower temperatures. Thus, we have uncovered an important role of HY5 in maintaining photosynthetic capacity at lower temperatures. Our research illustrated as well that abrupt changes in HY5 and PIF1 induced by light and temperature lead to corresponding adjustments in signaling by switching from transcriptional repression to activation, finely tuning photopigment gene expression (Figure 1D). However through diurnal cycles, PIFs and HY5 do not show strong diurnal shifts but are coordinated with the circadian clock to moderate rhythmic gene expression. This work is summarized in our publication: Toledo-Ortiz et al. PLoS Genet. 2014 10(6):e1004416. We are currently utilizing modeling approaches test a new hypothesis,that PIFs operate in partnership with phyA and the clock to set the timing and amplitude of output genes incl. photopigment genes. This work will be prepared for publication later this year.
A second paper (Bou-Torrent, Toledo-Ortiz, et al., Plant Physiol. 2015) focused on the changing roles of HY5 and PIF1 in the FR light-rich conditions of vegetation shade, that is commonly experienced by plants in natural habitats. FR-rich light induces the so called “shade avoidance response” that alters growth giving plants a competitive advantage in vegetation dense conditions. Shade also markedly reduces carotenoid accumulation, a process that we have shown is regulated by PIFs and HY5 under non shade conditions (see above; Toledo-Ortiz, PLoS Genet. 2014). Our study demonstrated that PIFs remain major repressors of carotenoid production in FR-rich shade light (Figure 2A). These light conditions stimulate PIF accumulation and activity, which reduces carotenoid accumulation. Although we and others have shown that HY5 opposes PIF action in response to light and temperature cues (refs), unexpectedly, HY5 does not antagonize PIF-control of carotenoids under shade conditions. Instead, we demonstrated that Phytochrome Rapidly Regulated 1 (PAR1), a transcriptional co-factor (and shade responses antagonist) fulfilled this role in shaded environments. Figure 2B illustrates that PAR1 positively regulates carotenoid accumulation in shade, accomplishing this by directly promoting PSY gene expression. PIF1-PAR1 form heterodimers lack capacity to bind DNA. Therefore PAR1 presence in the nucleus titrates out the amount of PIF1 bound to the promoter (Figure 2B). We showed that this novel module has an important function in preventing a complete shade induced blockage in PSY expression and ensuring carotenoid production (Figure 2C). Our results indicate that different modules formed by transcriptional activators and repressors are recruited to finely control photopigment biosynthesis in response to environmental cues (Figure 1A and 2D).
In summary, during Smartplant we have demonstrated that signal convergence of antagonistic regulators (PIFs-HY5, PIFs-PAR1) provide a very effective mechanism to integrate light and temperature signals. These operational modules are also key to link transcriptional responses to the circadian clock for optimization of growth and development. This project allowed us to develop a very productive collaborative with the Rodríguez-Concepción lab (CRAG, Barcelona) that culminated in two in high impact journals (Toledo-Ortiz et al. PLoS Genet. 2014; Bou-Torrent, Toledo-Ortiz, et al., Plant Physiol. 2015). We envisage a third publication that will combined experimental and theoretical analysis. Here we have been working inderdisciplinarily with theoretical scientists based in Synthsys Edinburgh. Our research findings have been presented at national and international conferences, including two oral presentations at the most relevant conference for plant photobiology (International Symposium of Plant Photobiology 2013 and 2015) and established new international collaborative efforts and solid collaborations with theoretical scientists at Synthsys Edinburgh. This research has been instrumental reinforcing the CV of Dr Toledo-Ortiz who has recently accepted a faculty position: Lecturer in Plant Sciences, at Lancaster University, UK (starting October 2015). This work has provided a mechanistic understanding of environmental control of photosynthesis and photoprotection. It therefore has broader relevance to applications that seek to improve photosynthetic capacity in the face of climate change. Such initiatives are central to the European Strategic Research Agenda to address climate change and impact in photosynthesis and agricultural productivity.
Website: http://hallidaylab.bio.ed.ac.uk/(si apre in una nuova finestra)
Carotenoid biosynthesis is tightly linked to photosynthesis and is therefore a key determinant of plant biomass and agricultural productivity. Carotenoid production is also strongly regulated by light and temperature, so improved knowledge of these regulatory pathways is fundamental to design new strategies to optimize carotenoid production in a changing environment. In SmartPlant we aimed to determine the molecular pathway that drives environmental control of carotenoid production. We identified a major regulatory module through which light and temperature signals converge to control carotenoid biosynthesis. We determined the functional properties of this molecular mechanism and its influence on carotenoid pathway dynamics.
The ability to sense and integrate light and temperature signals is essential for plant biomass production. The phytochrome red and far-red light photoreceptors are central molecular regulators of plasticity, altering plant growth and biomass build-up according to the light environment. Recent work has shown that the light pathways also act as a conduit for temperature signals, suggesting that these environmental cues utilize common pathways to regulate plant growth and development. Within the phytochrome-mediated changes during this critical time is the assembly of the photosynthetic apparatus, and the production of the photosynthetic pigments (chlorophylls/carotenoids). Carotenoids are integral accessory pigments in the light-absorbing antenna. During greening, carotenoids play an essential photoprotective role minimizing the potentially damaging effects of light on the photosynthetic apparatus. Therefore, to maximize light capture but minimize the damaging effects of light over the emerging photosynthetic machinery, the production of carotenoids and chlorophylls has to take place in a tightly controlled/interdependent manner. However, we do not currently understand how this regulation takes place. Improved knowledge of these regulatory pathways is fundamental to design new strategies to optimize carotenoid production.
In our project we aimed to define the roles of key phytochrome signaling components in photoprotection and maintenance of photosynthetic capacity over a range of light and temperature conditions that are experienced in nature. Experimental and modeling approaches were combined to determine mechanistic insights.
We previously showed that the Phytochrome Interacting Factors (PIFs) repress carotenoid accumulation by directly down-regulating the expression of the gene encoding for PHYTOENE SYNTHASE (PSY), the main rate-determining enzyme in carotenogenesis. Recent studies have shown that PIFs also participate in temperature signaling. During our characterization of PIFs in carotenogenesis we postulated the existence of an unidentified PSY-activator that becomes more effective with PIF depletion. Our analysis showed the bZIP transcription factor Long Hypocotyl 5 (HY5) fulfilled this role, operating as a potent PIF antagonist in the control of carotenoid accumulation. HY5 is required for the light induction of PSY, and carotenoid and chlorophyll synthesis. In chromatin immunoprecipitation assays we demonstrated that PIFs and HY5 target directly a common cis-element (G-box). We assessed the role of temperature in the PIFs-HY5 mediated control of photopigment content (Figure 1A). Our current results indicate that carotenoid and chlorophyll levels increase with temperature. PIFs are necessary to maintain control of photopigment levels over a temperature range. However, HY5 impact is more relevant at cooler temperatures (Figure 1B). In adult plants we demonstrated that HY5 is required to increase chlorophyll synthesis and carbon uptake in the cool (Figure 1C). Indeed, HY5 is required for dynamic light and temperature induced photopigment adjustments over the life span of the plant. Because of this the hy5 null mutant has lower carbon assimilation and a marked reduction in biomass at lower temperatures. Thus, we have uncovered an important role of HY5 in maintaining photosynthetic capacity at lower temperatures. Our research illustrated as well that abrupt changes in HY5 and PIF1 induced by light and temperature lead to corresponding adjustments in signaling by switching from transcriptional repression to activation, finely tuning photopigment gene expression (Figure 1D). However through diurnal cycles, PIFs and HY5 do not show strong diurnal shifts but are coordinated with the circadian clock to moderate rhythmic gene expression. This work is summarized in our publication: Toledo-Ortiz et al. PLoS Genet. 2014 10(6):e1004416. We are currently utilizing modeling approaches test a new hypothesis,that PIFs operate in partnership with phyA and the clock to set the timing and amplitude of output genes incl. photopigment genes. This work will be prepared for publication later this year.
A second paper (Bou-Torrent, Toledo-Ortiz, et al., Plant Physiol. 2015) focused on the changing roles of HY5 and PIF1 in the FR light-rich conditions of vegetation shade, that is commonly experienced by plants in natural habitats. FR-rich light induces the so called “shade avoidance response” that alters growth giving plants a competitive advantage in vegetation dense conditions. Shade also markedly reduces carotenoid accumulation, a process that we have shown is regulated by PIFs and HY5 under non shade conditions (see above; Toledo-Ortiz, PLoS Genet. 2014). Our study demonstrated that PIFs remain major repressors of carotenoid production in FR-rich shade light (Figure 2A). These light conditions stimulate PIF accumulation and activity, which reduces carotenoid accumulation. Although we and others have shown that HY5 opposes PIF action in response to light and temperature cues (refs), unexpectedly, HY5 does not antagonize PIF-control of carotenoids under shade conditions. Instead, we demonstrated that Phytochrome Rapidly Regulated 1 (PAR1), a transcriptional co-factor (and shade responses antagonist) fulfilled this role in shaded environments. Figure 2B illustrates that PAR1 positively regulates carotenoid accumulation in shade, accomplishing this by directly promoting PSY gene expression. PIF1-PAR1 form heterodimers lack capacity to bind DNA. Therefore PAR1 presence in the nucleus titrates out the amount of PIF1 bound to the promoter (Figure 2B). We showed that this novel module has an important function in preventing a complete shade induced blockage in PSY expression and ensuring carotenoid production (Figure 2C). Our results indicate that different modules formed by transcriptional activators and repressors are recruited to finely control photopigment biosynthesis in response to environmental cues (Figure 1A and 2D).
In summary, during Smartplant we have demonstrated that signal convergence of antagonistic regulators (PIFs-HY5, PIFs-PAR1) provide a very effective mechanism to integrate light and temperature signals. These operational modules are also key to link transcriptional responses to the circadian clock for optimization of growth and development. This project allowed us to develop a very productive collaborative with the Rodríguez-Concepción lab (CRAG, Barcelona) that culminated in two in high impact journals (Toledo-Ortiz et al. PLoS Genet. 2014; Bou-Torrent, Toledo-Ortiz, et al., Plant Physiol. 2015). We envisage a third publication that will combined experimental and theoretical analysis. Here we have been working inderdisciplinarily with theoretical scientists based in Synthsys Edinburgh. Our research findings have been presented at national and international conferences, including two oral presentations at the most relevant conference for plant photobiology (International Symposium of Plant Photobiology 2013 and 2015) and established new international collaborative efforts and solid collaborations with theoretical scientists at Synthsys Edinburgh. This research has been instrumental reinforcing the CV of Dr Toledo-Ortiz who has recently accepted a faculty position: Lecturer in Plant Sciences, at Lancaster University, UK (starting October 2015). This work has provided a mechanistic understanding of environmental control of photosynthesis and photoprotection. It therefore has broader relevance to applications that seek to improve photosynthetic capacity in the face of climate change. Such initiatives are central to the European Strategic Research Agenda to address climate change and impact in photosynthesis and agricultural productivity.
Website: http://hallidaylab.bio.ed.ac.uk/(si apre in una nuova finestra)
