A low-cost open-source imaging platform reveals spatiotemporal insight into leaf elongation and movement.

Plant organs move throughout the diurnal cycle, changing leaf and petiole positions to balance light capture, leaf temperature, and water loss under dynamic environmental conditions. Upward movement of the petiole, called hyponasty, is one of several traits of the shade avoidance syndrome (SAS). SAS traits are elicited upon perception of vegetation shade signals such as far-red light (FR) and improve light capture in dense vegetation. Monitoring plant movement at a high temporal resolution allows studying functionality and molecular regulation of hyponasty. However, high temporal resolution imaging solutions are often very expensive, making this unavailable to many researchers. Here, we present a modular and low-cost imaging setup, based on small Raspberry Pi computers that can track leaf movements and elongation growth with high temporal resolution. We also developed an open-source, semiautomated image analysis pipeline. Using this setup, we followed responses to FR enrichment, light intensity, and their interactions. Tracking both elongation and the angle of the petiole, lamina, and entire leaf in Arabidopsis (Arabidopsis thaliana) revealed insight into R:FR sensitivities of leaf growth and movement dynamics and the interactions of R:FR with background light intensity. The detailed imaging options of this system allowed us to identify spatially separate bending points for petiole and lamina positioning of the leaf.

Far-red light enrichment affects gene expression and architecture as well as growth and photosynthesis in rice.

Plants use light as a resource and signal. Photons within the 400-700 nm waveband are considered photosynthetically active. Far-red photons (FR, 700-800 nm) are used by plants to detect nearby vegetation and elicit the shade avoidance syndrome. In addition, FR photons have also been shown to contribute to photosynthesis, but knowledge about these dual effects remains scarce. Here, we study shoot-architectural and photosynthetic responses to supplemental FR light during the photoperiod in several rice varieties. We observed that FR enrichment only mildly affected the rice transcriptome and shoot architecture as compared to established model species, whereas leaf formation, tillering and biomass accumulation were clearly promoted. Consistent with this growth promotion, we found that CO2-fixation in supplemental FR was strongly enhanced, especially in plants acclimated to FR-enriched conditions as compared to control conditions. This growth promotion dominates the effects of FR photons on shoot development and architecture. When substituting FR enrichment with an end-of-day FR pulse, this prevented photosynthesis-promoting effects and elicited shade avoidance responses. We conclude that FR photons can have a dual role, where effects depend on the environmental context: in addition to being an environmental signal, they are also a potent source of harvestable energy.

Lights, location, action: Shade avoidance signalling over spatial scales.

Plants growing in dense vegetation stands need to flexibly position their photosynthetic organs to ensure optimal light capture in a competitive environment. They do so through a suite of developmental responses referred to as the shade avoidance syndrome. Belowground, root development is also adjusted in response to aboveground neighbour proximity. Canopies are dynamic and complex environments with heterogenous light cues in the far-red, red, blue and UV spectrum, which can be perceived with photoreceptors by spatially separated plant tissues. Molecular regulation of plant architecture adjustment via PHYTOCHROME-INTERACTING FACTOR (PIF) transcription factors and growth-related hormones such as auxin, gibberellic acid, brassinosteroids and abscisic acid were historically studied without much attention to spatial or tissue-specific context. Recent developments and technologies have, however, sparked strong interest in spatially explicit understanding of shade avoidance regulation. Other environmental factors such as temperature and nutrient availability interact with the molecular shade avoidance regulation network, often depending on the spatial location of the signals, and the responding organs. Here, we aim to review recent advances in how plants respond to heterogenous light cues and integrate these with other environmental signals.

Evaluation of the roles of brassinosteroid, gibberellin and auxin for tomato internode elongation in response to low red:far-red light.

In this study, we explore the interplay between the plant hormones gibberellins (GA), brassinosteroids (BR), and Indole-3-Acetic Acid (IAA) in their collective impact on plant shade avoidance elongation under varying light conditions. We focus particularly on low Red:Far-red (R:FR) light conditions achieved by supplementing the background light with FR. We characterized the tomato internode response to low R:FR and, with RNA-seq analysis, we were able to identify some of the potential regulatory hormonal pathways. Through a series of exogenous pharmacological modulations of GA, IAA, and BR, we demonstrate that GA and BR are sufficient but also necessary for inducing stem elongation under low R:FR light conditions. Intriguingly, while IAA alone shows limited effects, its combination with GA yields significant elongation, suggesting a nuanced hormonal balance. Furthermore, we unveil the complex interplay of these hormones under light with low R:FR, where the suppression of one hormone's effect can be compensated by the others. This study provides insights into the hormonal mechanisms governing plant adaptation to light, highlighting the intricate and adaptable nature of plant growth responses. Our findings have far-reaching implications for agricultural practices, offering potential strategies for optimizing plant growth and productivity in various lighting environments.

Mechanisms of far-red light-mediated dampening of defense against Botrytis cinerea in tomato leaves.

Plants detect neighboring competitors through a decrease in the ratio between red and far-red light (R:FR). This decreased R:FR is perceived by phytochrome photoreceptors and triggers shade avoidance responses such as shoot elongation and upward leaf movement (hyponasty). In addition to promoting elongation growth, low R:FR perception enhances plant susceptibility to pathogens: the growth-defense tradeoff. Although increased susceptibility in low R:FR has been studied for over a decade, the associated timing of molecular events is still unknown. Here, we studied the chronology of FR-induced susceptibility events in tomato (Solanum lycopersicum) plants pre-exposed to either white light (WL) or WL supplemented with FR light (WL+FR) prior to inoculation with the necrotrophic fungus Botrytis cinerea (B.c.). We monitored the leaf transcriptional changes over a 30-h time course upon infection and followed up with functional studies to identify mechanisms. We found that FR-induced susceptibility in tomato is linked to a general dampening of B.c.-responsive gene expression, and a delay in both pathogen recognition and jasmonic acid-mediated defense gene expression. In addition, we found that the supplemental FR-induced ethylene emissions affected plant immune responses under the WL+FR condition. This study improves our understanding of the growth-immunity tradeoff, while simultaneously providing leads to improve tomato resistance against pathogens in dense cropping systems.

A Gas-and-Brake Mechanism of bHLH Proteins Modulates Shade Avoidance.

Plants detect proximity of competitors through reduction in the ratio between red and far-red light that triggers the shade avoidance syndrome, inducing responses such as accelerated shoot elongation and early flowering. Shade avoidance is regulated by PHYTOCHROME INTERACTING FACTORs, a group of basic helix-loop-helix (bHLH) transcription factors. Another (b)HLH protein, KIDARI (KDR), which is non-DNA-binding, was identified in de-etiolation studies and proposed to interact with LONG HYPOCOTYL IN FAR-RED1 (HFR1), a (b)HLH protein that inhibits shade avoidance. Here, we established roles of KDR in regulating shade avoidance in Arabidopsis (Arabidopsis thaliana) and investigated how KDR regulates the shade avoidance network. We showed that KDR is a positive regulator of shade avoidance and interacts with several negative growth regulators. We identified KDR interactors using a combination of yeast two-hybrid screening and dedicated confirmations with bimolecular fluorescence complementation. We demonstrated that KDR is translocated primarily to the nucleus when coexpressed with these interactors. A genetic approach confirmed that several of these interactions play a functional role in shade avoidance; however, we propose that KDR does not interact with HFR1 to regulate shade avoidance. Based on these observations, we propose that shade avoidance is regulated by a three-layered gas-and-brake mechanism of bHLH protein interactions, adding a layer of complexity to what was previously known.

Reducing shade avoidance can improve Arabidopsis canopy performance against competitors.

Plants that grow in high density communities activate shade avoidance responses to consolidate light capture by individuals. Although this is an evolutionary successful strategy, it may not enhance performance of the community as a whole. Resources are invested in shade responses at the expense of other organs and light penetration through the canopy is increased, allowing invading competitors to grow better. Here we investigate if suppression of shade avoidance responses would enhance group performance of a monoculture community that is invaded by a competitor. Using different Arabidopsis genotypes, we show that suppression of shade-induced upward leaf movement in the pif7 mutant increases the pif7 communal performance against invaders as compared to a wild-type canopy. The invaders were more severely suppressed and the community grew larger as compared to wild type. Using computational modelling, we show that leaf angle variations indeed strongly affect light penetration and growth of competitors that invade the canopy. Our data thus show that modifying specific shade avoidance aspects can improve plant community performance. These insights may help to suppress weeds in crop stands.

Light signalling shapes plant-plant interactions in dense canopies.

Plants growing at high densities interact via a multitude of pathways. Here, we provide an overview of mechanisms and functional consequences of plant architectural responses initiated by light cues that occur in dense vegetation. We will review the current state of knowledge about shade avoidance, as well as its possible applications. On an individual level, plants perceive neighbour-associated changes in light quality and quantity mainly with phytochromes for red and far-red light and cryptochromes and phototropins for blue light. Downstream of these photoreceptors, elaborate signalling and integration takes place with the PHYTOCHROME INTERACTING FACTORS, several hormones and other regulators. This signalling leads to the shade avoidance responses, consisting of hyponasty, stem and petiole elongation, apical dominance and life cycle adjustments. Architectural changes of the individual plant have consequences for the plant community, affecting canopy structure, species composition and population fitness. In this context, we highlight the ecological, evolutionary and agricultural importance of shade avoidance.

Far-Red Light Detection in the Shoot Regulates Lateral Root Development through the HY5 Transcription Factor.

Plants in dense vegetation compete for resources and detect competitors through reflection of far-red (FR) light from surrounding plants. This reflection causes a reduced red (R):FR ratio, which is sensed through phytochromes. Low R:FR induces shade avoidance responses of the shoot and also changes the root system architecture, although this has received little attention so far. Here, we investigate the molecular mechanisms through which light detection in the shoot regulates root development in Arabidopsis thaliana We do so using a combination of microscopy, gene expression, and mutant study approaches in a setup that allows root imaging without exposing the roots to light treatment. We show that low R:FR perception in the shoot decreases the lateral root (LR) density by inhibiting LR emergence. This decrease in LR emergence upon shoot FR enrichment is regulated by phytochrome-dependent accumulation of the transcription factor ELONGATED HYPOCOTYL5 (HY5) in the LR primordia. HY5 regulates LR emergence by decreasing the plasma membrane abundance of PIN-FORMED3 and LIKE-AUX1 3 auxin transporters. Accordingly, FR enrichment reduces the auxin signal in the overlaying cortex cells, and this reduces LR outgrowth. This shoot-to-root communication can help plants coordinate resource partitioning under competition for light in high density fields.

The role of seasonality in reproduction of multiannual delayed gametophytes of Saccharina latissima.

Delayed gametophytes are able to grow vegetatively for prolonged periods of time. As such, they are potentially very valuable for kelp aquaculture given their great promise in opening up novel opportunities for kelp breeding and farming. However, large-scale application would require more in-depth understanding of how to control reproduction in delayed gametophytes. For newly formed gametophytes, many environmental factors for reproduction have been identified, with key drivers being light intensity, temperature, and the initial gametophyte density. However, the question of whether delayed gametophytes react similarly to these life cycle controls remains open for exploration. In this study, we performed a full factorial experiment on the influences of light intensity, temperature, and density on the reproduction of multiannual delayed gametophytes of Saccharina latissima, during which the number of sporophytes formed was counted. We demonstrate that delayed gametophytes of S. latissima can reliably reproduce sexually after more than a year of vegetative growth, depending on the effects between light intensity and temperature. Under higher light intensities (≥29 µmol photons · m-2 · s-1 ), optimal reproduction was observed at lower temperatures (10.2°C), while at lower light intensities (≤15 µmol photons · m-2 · s-1 ), optimal reproduction was observed at higher temperatures (≥12.6°C). Given the seasonal lag between solar radiation and sea surface temperature in natural systems, these conditions resemble those found during spring (i.e., increasing light intensity with low temperatures) and autumn (i.e., decreasing light intensity with higher temperatures). Seasonality can be used as an aquaculture tool to better control the reproduction of delayed gametophytes.

Far-red light promotes Botrytis cinerea disease development in tomato leaves via jasmonate-dependent modulation of soluble sugars.

Plants experience a decrease in the red:far-red light ratio (R:FR) when grown at high planting density. In addition to eliciting the shade avoidance response, low R:FR also enhances plant susceptibility to pathogens via modulation of defense hormone-mediated responses. However, other mechanisms, also affected by low R:FR, have not been considered as potential components in FR-induced susceptibility. Here, we identify FR-induced accumulation of leaf soluble sugars as a novel component of FR-induced susceptibility. We observed that phytochrome inactivation by FR or phytochrome B mutation was associated with elevated leaf glucose and fructose levels and enhanced disease severity caused by Botrytis cinerea. By experimentally manipulating internal leaf sugar levels, we found that the FR-induced susceptibility in tomato was partly sugar-dependent. Further analysis revealed that the observed sugar accumulation in supplemental FR occurred in a jasmonic acid (JA)-dependent manner, and the JA biosynthesis mutant def1 also displayed elevated soluble sugar levels, which was rescued by exogenous methyl jasmonate (MeJA) application. We propose that the reduced JA responsiveness under low R:FR promotes disease symptoms not only via dampened induction of defense responses, but also via increased levels of soluble sugars that supports pathogen growth in tomato leaves.

Molecular Profiles of Contrasting Shade Response Strategies in Wild Plants: Differential Control of Immunity and Shoot Elongation.

Plants growing at high densities elongate their shoots to reach for light, a response known as the shade avoidance syndrome (SAS). Phytochrome-mediated detection of far-red light reflection from neighboring plants activates growth-promoting molecular pathways leading to SAS However, it is unknown how plants that complete their life cycle in the forest understory and are shade tolerant prevent SAS when exposed to shade. Here, we show how two wild Geranium species from different native light environments regulate contrasting responses to light quality cues. A comparative RNA sequencing approach unveiled the molecular underpinnings of their contrasting growth responses to far-red light enrichment. It also identified differential phytochrome control of plant immunity genes and confirmed that far-red enrichment indeed contrastingly affects resistance against Botrytis cinerea between the two species. Furthermore, we identify a number of candidate regulators of differential shade avoidance. Three of these, the receptor-like kinases FERONIA and THESEUS1 and the non-DNA binding bHLH protein KIDARI, are functionally validated in Arabidopsis thaliana through gene knockout and/or overexpression studies. We propose that these components may be associated with either showing or not showing shade avoidance responses.

Variation in plastic responses to light results from selection in different competitive environments-A game theoretical approach using virtual plants.

Phenotypic plasticity is a vital strategy for plants to deal with changing conditions by inducing phenotypes favourable in different environments. Understanding how natural selection acts on variation in phenotypic plasticity in plants is therefore a central question in ecology, but is often ignored in modelling studies. Here we present a new modelling approach that allows for the analysis of selection for variation in phenotypic plasticity as a response strategy. We assess selection for shade avoidance strategies of Arabidopsis thaliana in response to future neighbour shading signalled through a decrease in red:far-red (R:FR) ratio. For this, we used a spatially explicit 3D virtual plant model that simulates individual Arabidopsis plants competing for light in different planting densities. Plant structure and growth were determined by the organ-specific interactions with the light environment created by the vegetation structure itself. Shade avoidance plastic responses were defined by a plastic response curve relating petiole elongation and lamina growth to R:FR perceived locally. Different plasticity strategies were represented by different shapes of the response curve that expressed different levels of R:FR sensitivity. Our analyses show that the shape of the selected shade avoidance strategy varies with planting density. At higher planting densities, more sensitive response curves are selected for than at lower densities. In addition, the balance between lamina and petiole responses influences the sensitivity of the response curves selected for. Combining computational virtual plant modelling with a game theoretical analysis represents a new step towards analysing how natural selection could have acted upon variation in shade avoidance as a response strategy, which can be linked to genetic variation and underlying physiological processes.

The bHLH network underlying plant shade-avoidance.

Shade is a potential threat to many plant species. When shade-intolerant plants detect neighbours, they elongate their stems and leaves in an effort to maximise their light capture. This developmental programme, known as 'shade-avoidance' is tightly controlled by specialised photoreceptors and a suite of transcriptional regulators. The basic helix-loop-helix (bHLH) family of transcription factors are particularly important for shade-induced elongation. In recent years, it has become apparent that many members of this family heterodimerise and that together they form a complex regulatory network. This review summarises recent work into the structure of the bHLH network and how it regulates elongation growth. In addition to this, we highlight how photoreceptors modulate the function of the network via direct interaction with transcription factors. It is hoped that the information integrated in this review will provide a useful theoretical framework for future studies on the molecular basis of shade-avoidance in plants.

How light and biomass density influence the reproduction of delayed Saccharina latissima gametophytes (Phaeophyceae).

Kelp life-cycle transitions are complex and susceptible to various (a)biotic controls. Understanding the microscopic part of the kelp's lifecycle is of key importance, as gametophytes form a critical phase influencing, among others, the distributional limits of the species. Many environmental controls have been identified that affect kelp gametogenesis, whose interactive effects can be subtle and counterintuitive. Here we performed a fully factorial experiment on the (interactive) influences of light intensity, light quality, and the Initial Gametophyte Density (IGD) on Saccharina latissima reproduction and vegetative growth of delayed gametophytes. A total of 144 cultures were followed over a period of 21 d. The IGD was a key determinant for reproductive success, with increased IGDs (≥0.04 mg DW · mL-1 ) practically halting reproduction. Interestingly, the effects of IGDs were not affected by nutrient availability, suggesting a resource-independent effect of density on reproduction. The Photosynthetically Usable Radiation (PUR), overarching the quantitative contribution of both light intensity and light quality, correlated with both reproduction and vegetative growth. The PUR furthermore specifies that the contribution of light quality, as a lifecycle control, is a matter of absorbed photon flux instead of color signaling. We hypothesize that (i) the number of photons absorbed, independent of their specific wavelength, and (ii) IGD interactions, independent of nutrient availability, are major determinants of reproduction in S. latissima gametophytes. These insights help understand kelp gametophyte development and dispersal under natural conditions, while also aiding the control of in vitro gametophyte cultures.

Neighbor detection at the leaf tip adaptively regulates upward leaf movement through spatial auxin dynamics.

Vegetation stands have a heterogeneous distribution of light quality, including the red/far-red light ratio (R/FR) that informs plants about proximity of neighbors. Adequate responses to changes in R/FR are important for competitive success. How the detection and response to R/FR are spatially linked and how this spatial coordination between detection and response affects plant performance remains unresolved. We show in Arabidopsis thaliana and Brassica nigra that localized FR enrichment at the lamina tip induces upward leaf movement (hyponasty) from the petiole base. Using a combination of organ-level transcriptome analysis, molecular reporters, and physiology, we show that PIF-dependent spatial auxin dynamics are key to this remote response to localized FR enrichment. Using computational 3D modeling, we show that remote signaling of R/FR for hyponasty has an adaptive advantage over local signaling in the petiole, because it optimizes the timing of leaf movement in response to neighbors and prevents hyponasty caused by self-shading.

Three Auxin Response Factors Promote Hypocotyl Elongation.

The hormone auxin regulates growth largely by affecting gene expression. By studying Arabidopsis (Arabidopsis thaliana) mutants deficient in AUXIN RESPONSE FACTORS (ARFs), we have identified three ARF proteins that are required for auxin-responsive hypocotyl elongation. Plants deficient in these factors have reduced responses to environmental conditions that increase auxin levels, including far-red-enriched light and high temperature. Despite having decreased auxin responses, the ARF-deficient plants responded to brassinosteroid and gibberellin, indicating that different hormones can act partially independently. Aux/IAA proteins, encoded by IAA genes, interact with ARF proteins to repress auxin response. Silencing expression of multiple IAA genes increased hypocotyl elongation, suggesting that Aux/IAA proteins modulate ARF activity in hypocotyls in a potential negative feedback loop.

Subtle variation in shade avoidance responses may have profound consequences for plant competitiveness.

Background and Aims: Although phenotypic plasticity has been shown to be beneficial for plant competitiveness for light, there is limited knowledge on how variation in these plastic responses plays a role in determining competitiveness. Methods: A combination of detailed plant experiments and functional-structural plant (FSP) modelling was used that captures the complex dynamic feedback between the changing plant phenotype and the within-canopy light environment in time and 3-D space. Leaf angle increase (hyponasty) and changes in petiole elongation rates in response to changes in the ratio between red and far-red light, two important shade avoidance responses in Arabidopsis thaliana growing in dense population stands, were chosen as a case study for plant plasticity. Measuring and implementing these responses into an FSP model allowed simulation of plant phenotype as an emergent property of the underlying growth and response mechanisms. Key Results: Both the experimental and model results showed that substantial differences in competitiveness may arise between genotypes with only marginally different hyponasty or petiole elongation responses, due to the amplification of plant growth differences by small changes in plant phenotype. In addition, this study illustrated that strong competitive responses do not necessarily have to result in a tragedy of the commons; success in competition at the expense of community performance. Conclusions: Together, these findings indicate that selection pressure could probably have played a role in fine-tuning the sensitive shade avoidance responses found in plants. The model approach presented here provides a novel tool to analyse further how natural selection could have acted on the evolution of plastic responses.

Ethylene- and Shade-Induced Hypocotyl Elongation Share Transcriptome Patterns and Functional Regulators.

Plants have evolved shoot elongation mechanisms to escape from diverse environmental stresses such as flooding and vegetative shade. The apparent similarity in growth responses suggests a possible convergence of the signaling pathways. Shoot elongation is mediated by passive ethylene accumulating to high concentrations in flooded plant organs and by changes in light quality and quantity under vegetation shade. Here, we study hypocotyl elongation as a proxy for shoot elongation and delineate Arabidopsis (Arabidopsis thaliana) hypocotyl length kinetics in response to ethylene and shade. Based on these kinetics, we further investigated ethylene- and shade-induced genome-wide gene expression changes in hypocotyls and cotyledons separately. Both treatments induced a more extensive transcriptome reconfiguration in the hypocotyls compared with the cotyledons. Bioinformatics analyses suggested contrasting regulation of growth promotion- and photosynthesis-related genes. These analyses also suggested an induction of auxin, brassinosteroid, and gibberellin signatures and the involvement of several candidate regulators in the elongating hypocotyls. Pharmacological and mutant analyses confirmed the functional involvement of several of these candidate genes and physiological control points in regulating stress-escape responses to different environmental stimuli. We discuss how these signaling networks might be integrated and conclude that plants, when facing different stresses, utilize a conserved set of transcriptionally regulated genes to modulate and fine-tune growth.