Characterizing the Synergistic Response Induced by Warm Temperatures and Shade.

Plants exhibit an impressive capability to detect and respond to neighboring plants by closely monitoring changes in the light spectrum. They possess the ability to perceive adjustments in the ratio of red (R) to far-red (FR) light (R/FR) triggered by the presence of nearby plants, even before experiencing complete shading. When the R/FR ratio falls below 1, plants activate a shade avoidance response that manifests as hypocotyl elongation. Furthermore, elevated ambient temperatures can also stimulate hypocotyl elongation. As hypocotyl elongation is a visible characteristic, it is a valuable indicator for monitoring shade avoidance response, warm ambient temperature response, and the interplay between the two.

PIF7 is a master regulator of thermomorphogenesis in shade.

The size of plant organs is highly responsive to environmental conditions. The plant's embryonic stem, or hypocotyl, displays phenotypic plasticity, in response to light and temperature. The hypocotyl of shade avoiding species elongates to outcompete neighboring plants and secure access to sunlight. Similar elongation occurs in high temperature. However, it is poorly understood how environmental light and temperature cues interact to effect plant growth. We found that shade combined with warm temperature produces a synergistic hypocotyl growth response that dependent on PHYTOCHROME-INTERACTING FACTOR 7 (PIF7) and auxin. This unique but agriculturally relevant scenario was almost totally independent on PIF4 activity. We show that warm temperature is sufficient to promote PIF7 DNA binding but not transcriptional activation and we demonstrate that additional, unknown factor/s must be working downstream of the phyB-PIF-auxin module. Our findings will improve the predictions of how plants will respond to increased ambient temperatures when grown at high density.

Coordinating the morphogenesis-differentiation balance by tweaking the cytokinin-gibberellin equilibrium.

Morphogenesis and differentiation are important stages in organ development and shape determination. However, how they are balanced and tuned during development is not fully understood. In the compound leaved tomato, an extended morphogenesis phase allows for the initiation of leaflets, resulting in the compound form. Maintaining a prolonged morphogenetic phase in early stages of compound-leaf development in tomato is dependent on delayed activity of several factors that promote differentiation, including the CIN-TCP transcription factor (TF) LA, the MYB TF CLAU and the plant hormone Gibberellin (GA), as well as on the morphogenesis-promoting activity of the plant hormone cytokinin (CK). Here, we investigated the genetic regulation of the morphogenesis-differentiation balance by studying the relationship between LA, CLAU, TKN2, CK and GA. Our genetic and molecular examination suggest that LA is expressed earlier and more broadly than CLAU and determines the developmental context of CLAU activity. Genetic interaction analysis indicates that LA and CLAU likely promote differentiation in parallel genetic pathways. These pathways converge downstream on tuning the balance between CK and GA. Comprehensive transcriptomic analyses support the genetic data and provide insights into the broader molecular basis of differentiation and morphogenesis processes in plants.

Coordination of differentiation rate and local patterning in compound-leaf development.

The variability in leaf form in nature is immense. Leaf patterning occurs by differential growth, taking place during a limited window of morphogenetic activity at the leaf marginal meristem. While many regulators have been implicated in the designation of the morphogenetic window and in leaf patterning, how these effectors interact to generate a particular form is still not well understood. We investigated the interaction among different effectors of tomato (Solanum lycopersicum) compound-leaf development, using genetic and molecular analyses. Mutations in the tomato auxin response factor SlARF5/SlMP, which normally promotes leaflet formation, suppressed the increased leaf complexity of mutants with extended morphogenetic window. Impaired activity of the NAC/CUC transcription factor GOBLET (GOB), which specifies leaflet boundaries, also reduced leaf complexity in these backgrounds. Analysis of genetic interactions showed that the patterning factors SlMP, GOB and the MYB transcription factor LYRATE (LYR) coordinately regulate leaf patterning by modulating in parallel different aspects of leaflet formation and shaping. This work places an array of developmental regulators in a morphogenetic context. It reveals how organ-level differentiation rate and local growth are coordinated to sculpture an organ. These concepts are applicable to the coordination of pattering and differentiation in other species and developmental processes.

HY5 and phytochrome activity modulate shoot-to-root coordination during thermomorphogenesis in Arabidopsis.

Temperature is one of the most impactful environmental factors to which plants adjust their growth and development. Although the regulation of temperature signaling has been extensively investigated for the aerial part of plants, much less is known and understood about how roots sense and modulate their growth in response to fluctuating temperatures. Here, we found that shoot and root growth responses to high ambient temperature are coordinated during early seedling development in Arabidopsis A shoot signaling module that includes HY5, the phytochromes and the PIFs exerts a central function in coupling these growth responses and maintaining auxin levels in the root. In addition to the HY5/PIF-dependent shoot module, a regulatory axis composed of auxin biosynthesis and auxin perception factors controls root responses to high ambient temperature. Taken together, our findings show that shoot and root developmental responses to temperature are tightly coupled during thermomorphogenesis and suggest that roots integrate energy signals with local hormonal inputs.

Local HY5 Activity Mediates Hypocotyl Growth and Shoot-to-Root Communication.

Plants optimize their growth in fluctuating environments using information acquired by different organs. This information is then transmitted through the rest of the plant using both short- and long-distance signals, including hormones and mobile proteins. Although a few of these signals have been characterized, long-distance signaling is not well understood in plants. Recently, the light-regulated transcription factor HY5 was reported to move from the shoot to the root to regulate root growth. We generated a cell-type specifically expressed HY5 fusion protein that could not be detected outside the tissue in which it was targeted. By expressing this DOF-HY5 protein in specific cell types of the hypocotyl, we showed that its local activity was sufficient to regulate hypocotyl growth. We also found that, although DOF-HY5 was expressed specifically in the shoot and not detected in the roots, it could rescue hy5 growth defects in primary roots but not in lateral roots. We therefore conclude that HY5 protein mobility is not required in the hypocotyl or for shoot-to-root communication. Our results indicate that a signal downstream of, or in parallel with, HY5 in the shoot is mobile and links shoot and root growth.

Chimeric Activators and Repressors Define HY5 Activity and Reveal a Light-Regulated Feedback Mechanism.

The first exposure to light marks a crucial transition in plant development. This transition relies on the transcription factor HY5 controlling a complex downstream growth program. Despite its importance, its function in transcription remains unclear. Previous studies have generated lists of thousands of potential target genes and competing models of HY5 transcription regulation. In this work, we carry out detailed phenotypic and molecular analysis of constitutive activator and repressor HY5 fusion proteins. Using this strategy, we were able to filter out large numbers of genes that are unlikely to be direct targets, allowing us to eliminate several proposed models of HY5's mechanism of action. We demonstrate that the primary activity of HY5 is promoting transcription and that this function relies on other, likely light-regulated, factors. In addition, this approach reveals a molecular feedback loop via the COP1/SPA E3 ubiquitin ligase complex, suggesting a mechanism that maintains low HY5 in the dark, primed for rapid accumulation to reprogram growth upon light exposure. Our strategy is broadly adaptable to the study of transcription factor activity. Lastly, we show that modulating this feedback loop can generate significant phenotypic diversity in both Arabidopsis (Arabidopsis thaliana) and tomato (Solanum lycopersicum).

Dancing in the dark: darkness as a signal in plants.

Daily cycles of light and dark provide an organizing principle and temporal constraints under which life on Earth evolved. While light is often the focus of plant studies, it is only half the story. Plants continuously adjust to their surroundings, taking both dawn and dusk as cues to organize their growth, development and metabolism to appropriate times of day. In this review, we examine the effects of darkness on plant physiology and growth. We describe the similarities and differences between seedlings grown in the dark versus those grown in light-dark cycles, and the evolution of etiolated growth. We discuss the integration of the circadian clock into other processes, looking carefully at the points of contact between clock genes and growth-promoting gene-regulatory networks in temporal gating of growth. We also examine daily starch accumulation and degradation, and the possible contribution of dark-specific metabolic controls in regulating energy and growth. Examining these studies together reveals a complex and continuous balancing act, with many signals, dark included, contributing information and guiding the plant through its life cycle. The extraordinary interconnection between light and dark is manifest during cycles of day and night and during seedling emergence above versus below the soil surface.

The epidermis coordinates auxin-induced stem growth in response to shade.

Growth of a complex multicellular organism requires coordinated changes in diverse cell types. These cellular changes generate organs of the correct size, shape, and functionality. In plants, the growth hormone auxin induces stem elongation in response to shade; however, which cell types of the stem perceive the auxin signal and contribute to organ growth is poorly understood. Here, we blocked the transcriptional response to auxin within specific tissues to show that auxin signaling is required in many cell types for correct hypocotyl growth in shade, with a key role for the epidermis. Combining genetic manipulations in Arabidopsis thaliana with transcriptional profiling of the hypocotyl epidermis from Brassica rapa, we show that auxin acts in the epidermis in part by inducing activity of the locally acting, growth-promoting brassinosteroid pathway. Our findings clarify cell-specific auxin function in the hypocotyl and highlight the complexity of cell type interactions within a growing organ.

A role for APETALA1/fruitfull transcription factors in tomato leaf development.

Flexible maturation rates underlie part of the diversity of leaf shape, and tomato (Solanum lycopersicum) leaves are compound due to prolonged organogenic activity of the leaf margin. The CINCINNATA-teosinte branched1, cycloidea, PCF (CIN-TCP) transcription factor lanceolate (LA) restricts this organogenic activity and promotes maturation. Here, we show that tomato APETALA1/fruitfull (AP1/FUL) MADS box genes are involved in tomato leaf development and are repressed by LA. AP1/FUL expression is correlated negatively with LA activity and positively with the organogenic activity of the leaf margin. LA binds to the promoters of the AP1/FUL genes MBP20 and TM4. Overexpression of MBP20 suppressed the simple-leaf phenotype resulting from upregulation of LA activity or from downregulation of class I knotted like homeobox (KNOXI) activity. Overexpression of a dominant-negative form of MBP20 led to leaf simplification and partly suppressed the increased leaf complexity of plants with reduced LA activity or increased KNOXI activity. Tomato plants overexpressing miR319, a negative regulator of several CIN-TCP genes including LA, flower with fewer leaves via an SFT-dependent pathway, suggesting that miR319-sensitive CIN-TCPs delay flowering in tomato. These results identify a role for AP1/FUL genes in vegetative development and show that leaf and plant maturation are regulated via partially independent mechanisms.

The tomato leaf as a model system for organogenesis.

Compound tomato leaves are composed of multiple leaflets that are generated gradually during leaf development, and each resembles a simple leaf. The elaboration of a compound leaf form requires the maintenance of transient organogenic activity at the leaf margin. The developmental window of organogenic activity is defined by the antagonistic activities of factors that promote maturation, such as TCP transcription factors, SFT and gibberellin, and factors that delay maturation, such as KNOX transcription factors and cytokinin. Leaflet initiation sites are specified spatially and temporally by spaced and specific activities of CUCs, auxin and ENTIRE, as well as additional factors. The partially indeterminate growth of the compound tomato leaf makes it a useful model to understand the balance between determinate and indeterminate growth, and the mechanisms of organogenesis, some of which are common to many developmental processes in plants.

From organelle to organ: ZRIZI MATE-Type transporter is an organelle transporter that enhances organ initiation.

Plant architecture is a predictable but flexible trait. The timing and position of organ initiation from the shoot apical meristem (SAM) contribute to the final plant form. While much progress has been made recently in understanding how the site of leaf initiation is determined, the mechanism underlying the temporal interval between leaf primordia is still largely unknown. The Arabidopsis ZRIZI (ZRZ) gene belongs to a large gene family encoding multidrug and toxic compound extrusion (MATE) transporters. Unique among plant MATE transporters identified so far, ZRZ is localized to the membrane of a small organelle, possibly the mitochondria. Plants overexpressing ZRZ in initiating leaves are short, produce leaves much faster than wild-type plants and show enhanced growth of axillary buds. These results suggest that ZRZ is involved in communicating a leaf-borne signal that determines the rate of organ initiation.

Dynamic growth program regulated by LANCEOLATE enables flexible leaf patterning.

During their development, leaves progress through a highly controlled yet flexible developmental program. Transcription factors from the CIN-TCP family affect leaf shape by regulating the timing of leaf maturation. Characterization of mutants in the tomato (Solanum lycopersicum) CIN-TCP gene LANCEOLATE (LA) led us to hypothesize that a threshold LA-like activity promotes leaf differentiation. Here, we examined the relationship between LA activity, leaf maturation, and final leaf size and shape. Leaves of diverse shapes from various Solanaceae species or from different positions on the tomato plant differed in the timing of growth and maturation, and these were often associated with altered LA expression dynamics. Accordingly, genetic manipulations of LA activity in tomato altered leaf growth and maturation, leading to changes in leaf size and shape. LA expression sustained until late stages of tomato leaf development, and stage-specific overexpression of miR319, a negative regulator of CIN-TCP genes, confirmed that LA-like proteins affect leaf development through these late stages. Together, our results imply that dynamic spatial and temporal leaf maturation, coordinated by LA-like genes, enables the formation of variable leaf forms.

Cytokinin regulates compound leaf development in tomato.

Leaf shape diversity relies on transient morphogenetic activity in leaf margins. However, how this morphogenetic capacity is maintained is still poorly understood. Here, we uncover a role for the hormone cytokinin (CK) in the regulation of morphogenetic activity of compound leaves in tomato (Solanum lycopersicum). Manipulation of CK levels led to alterations in leaf complexity and revealed a unique potential for prolonged growth and morphogenesis in tomato leaves. We further demonstrate that the effect of CK on leaf complexity depends on proper localization of auxin signaling. Genetic analysis showed that reduction of CK levels suppresses the effect of Knotted1 like homeobox (KNOXI) proteins on leaf shape and that CK can substitute for KNOXI activity at the leaf margin, suggesting that CK mediates the activity of KNOXI proteins in the regulation of leaf shape. These results imply that CK regulates flexible leaf patterning by dynamic interaction with additional hormones and transcription factors.

Stage-specific regulation of Solanum lycopersicum leaf maturation by class 1 KNOTTED1-LIKE HOMEOBOX proteins.

Class 1 KNOTTED1-LIKE HOMEOBOX (KNOXI) genes encode transcription factors that are expressed in the shoot apical meristem (SAM) and are essential for SAM maintenance. In some species with compound leaves, including tomato (Solanum lycopersicum), KNOXI genes are also expressed during leaf development and affect leaf morphology. To dissect the role of KNOXI proteins in leaf patterning, we expressed in tomato leaves a fusion of the tomato KNOXI gene Tkn2 with a sequence encoding a repressor domain, expected to repress common targets of tomato KNOXI proteins. This resulted in the formation of small, narrow, and simple leaves due to accelerated differentiation. Overexpression of the wild-type form of Tkn1 or Tkn2 in young leaves also resulted in narrow and simple leaves, but in this case, leaf development was blocked at the initiation stage. Expression of Tkn1 or Tkn2 during a series of spatial and temporal windows in leaf development identified leaf initiation and primary morphogenesis as specific developmental contexts at which the tomato leaf is responsive to KNOXI activity. Arabidopsis thaliana leaves responded to overexpression of Arabidopsis or tomato KNOXI genes during the morphogenetic stage but were largely insensitive to their overexpression during leaf initiation. These results imply that KNOXI proteins act at specific stages within the compound-leaf development program to delay maturation and enable leaflet formation, rather than set the compound leaf route.