Gravitropic Set-point Angle Research Articles

IGT/LAZY genes are differentially influenced by light and required for light-induced change to organ angle

BackgroundPlants adjust their growth orientations primarily in response to light and gravity signals. Considering that the gravity vector is fixed and the angle of light incidence is constantly changing, plants must somehow integrate these signals to establish organ orientation, commonly referred to as gravitropic set-point angle (GSA). The IGT gene family contains known regulators of GSA, including the gene clades LAZY, DEEPER ROOTING (DRO), and TILLER ANGLE CONTROL (TAC).ResultsHere, we investigated the influence of light on different aspects of GSA phenotypes in LAZY and DRO mutants, as well as the influence of known light signaling pathways on IGT gene expression. Phenotypic analysis revealed that LAZY and DRO genes are collectively required for changes in the angle of shoot branch tip and root growth in response to light. Single lazy1 mutant branch tips turn upward in the absence of light and in low light, similar to wild-type, and mimic triple and quadruple IGT mutants in constant light and high-light conditions, while triple and quadruple IGT/LAZY mutants show little to no response to changing light regimes. Further, the expression of IGT/LAZY genes is differentially influenced by daylength, circadian clock, and light signaling.ConclusionsCollectively, the data show that differential expression of LAZY and DRO genes are required to enable plants to alter organ angles in response to light-mediated signals.

Peach LAZY1 and DRO1 protein-protein interactions and co-expression with PRAF/RLD family support conserved gravity-related protein interactions across plants.

IGT/LAZY proteins play a central role in determining gravitropic set point angle and orientation of lateral organs across plant species. Recent work in model systems has demonstrated that interactions between IGT/LAZY proteins and BREVIS RADIX (BRX)-domain containing proteins, such as PH, RCC1, AND FYVE/RCC1-LIKE DOMAIN (PRAF/RLD), and BREVIS RADIX LIKE (BRXL) family members, are mechanistically important for setting gravitropic set point angle. Here, we identified peach PRAF/RLD proteins as interactors of the peach IGT/LAZY proteins PpeLAZY1 and DEEPER ROOTING 1 (PpeDRO1) from a yeast-two-hybrid screen. We also show that the BRX domains of these interacting proteins have high sequence similarity with PRAF/RLD and BRX family proteins from rice and Arabidopsis. Further, PpeLAZY1 and the peach PRAF/RLD interactors are all expressed at relatively high levels in leaf, meristem, and shoot tip tissues. Together, this evidence supports the importance and conservation of IGT/LAZY-BRX-domain interactions, which underlie setting gravitropic set point angle across angiosperms.

Transcriptional regulation of MdPIN7 by MdARF19 during gravity-induced formation of adventitious root GSA in self-rooted apple stock

Self-rooted apple stock is widely used for apple production. However, the shallowness of the adventitious roots in self-rooted apple stock leads to poor performance in the barren orchards of China. This is because of the considerable difference in the development of a gravitropic set-point angle (GSA) between self-rooted apple stock and seedling rootstock. Therefore, it is crucial to study the molecular mechanism of adventitious root GSA in self-rooted apple stock for breeding self-rooted and deep-rooted apple rootstock cultivars. An apple auxin response factor MdARF19 functioned to establish the adventitious root GSA of self-rooted apple stock in response to gravity and auxin signals. MdARF19 bound directly to the MdPIN7 promoter, activating its transcriptional expression and thus regulating the formation of the adventitious root GSA in 12-2 self-rooted apple stock. However, MdARF19 influenced the expression of auxin efflux carriers (MdPIN3 and MdPIN10) and the establishment of adventitious root GSA of self-rooted apple stock in response to gravity signals by direct activation of MdFLP. Our findings provide new information on the transcriptional regulation of MdPIN7 by auxin response factor MdARF19 in the regulation of the adventitious root GSA of self-rooted apple stock in response to gravity and auxin signals.

Antigravitropic PIN polarization maintains non-vertical growth in lateral roots

Lateral roots are typically maintained at non-vertical angles with respect to gravity. These gravitropic setpoint angles are intriguing because their maintenance requires that roots are able to effect growth response both with and against the gravity vector, a phenomenon previously attributed to gravitropism acting against an antigravitropic offset mechanism. Here we show how the components mediating gravitropism in the vertical primary root—PINs and phosphatases acting upon them—are reconfigured in their regulation such that lateral root growth at a range of angles can be maintained. We show that the ability of Arabidopsis lateral roots to bend both downward and upward requires the generation of auxin asymmetries and is driven by angle-dependent variation in downward gravitropic auxin flux acting against angle-independent upward, antigravitropic flux. Further, we demonstrate a symmetry in auxin distribution in lateral roots at gravitropic setpoint angle that can be traced back to a net, balanced polarization of PIN3 and PIN7 auxin transporters in the columella. These auxin fluxes are shifted by altering PIN protein phosphoregulation in the columella, either by introducing PIN3 phosphovariant versions or via manipulation of levels of the phosphatase subunit PP2A/RCN1. Finally, we show that auxin, in addition to driving lateral root directional growth, acts within the lateral root columella to induce more vertical growth by increasing RCN1 levels, causing a downward shift in PIN3 localization, thereby diminishing the magnitude of the upward, antigravitropic auxin flux.

Shoot gravitropism and organ straightening cooperate to arrive at a mechanically favorable shape in Arabidopsis

Gravitropism is the plant organ bending in response to gravity, while a straightening mechanism prevents bending beyond the gravitropic set-point angle. The promotion and prevention of bending occur simultaneously around the inflorescence stem tip. How these two opposing forces work together and what part of the stem they affect are unknown. To understand the mechanical forces involved, we rotated wild type and organ-straightening-deficient mutant (myosin xif xik) Arabidopsis plants to a horizontal position to initiate bending. The mutant stems started to bend before the wild-type stems, which led us to hypothesize that the force preventing bending was weaker in mutant. We modeled the wild-type and mutant stems as elastic rods, and evaluated two parameters: an organ-angle-dependent gravitropic-responsive parameter (β) and an organ-curvature-dependent proprioceptive-responsive parameter (γ). Our model showed that these two parameters were lower in mutant than in wild type, implying that, unexpectedly, both promotion and prevention of bending are weak in mutant. Subsequently, finite element method simulations revealed that the compressive stress in the middle of the stem was significantly lower in wild type than in mutant. The results of this study show that myosin-XIk-and-XIf-dependent organ straightening adjusts the stress distribution to achieve a mechanically favorable shape.

It takes more than being LAZY: regulation of shoot branching angle by BRXL4

It takes more than being <scp>LAZY</scp>: regulation of shoot branching angle by <scp>BRXL4</scp>

Gravity sensing and responses in the coordination of the shoot gravitropic setpoint angle.

Gravity is one of the fundamental environmental cues that affect plant development. Indeed, the plant architecture in the shoots and roots is modulated by gravity. Stems grow vertically upward, whereas lateral organs, such as the lateral branches in shoots, tend to grow at a specific angle according to a gravity vector known as the gravitropic setpoint angle (GSA). During this process, gravity is sensed in specialised gravity-sensing cells named statocytes, which convert gravity information into biochemical signals, leading to asymmetric auxin distribution and driving asymmetric cell division/expansion in the organs to achieve gravitropism. As a hypothetical offset mechanism against gravitropism to determine the GSA, the anti-gravitropic offset (AGO) has been proposed. According to this concept, the GSA is a balance of two antagonistic growth components, that is gravitropism and the AGO. Although the nature of the AGO has not been clarified, studies have suggested that gravitropism and the AGO share a common gravity-sensing mechanism in statocytes. This review discusses the molecular mechanisms underlying gravitropism as well as the hypothetical AGO in the control of the GSA.

Molecular Cloning and Functional Characterization of FOUR LIPS in Pomegranate, a Protein Involved in Regulating the Gravitropic Set-Point Angle of Adventitious Roots

Molecular Cloning and Functional Characterization of FOUR LIPS in Pomegranate, a Protein Involved in Regulating the Gravitropic Set-Point Angle of Adventitious Roots

CmRCC1 Gene From Pumpkin Confers Cold Tolerance in Tobacco by Modulating Root Architecture and Photosynthetic Activity.

Low-temperature stress is the main limiting factor of cucurbit crop cultivation as it affects crop yield and quality. The identification of genes involved in cold tolerance is a crucial aspect of pumpkin rootstock breeding. Here, we examined the function of a pumpkin Regulator of Chromosome Condensation 1 (CmRCC1) gene in the root development and cold stress responses of tobacco (Nicotiana benthamiana). CmRCC1 expression was differentially induced in pumpkin root, stem, and leaf under cold stress. Transient transformation showed that CmRCC1 is located in the nucleus. CmRCC1 overexpression in tobacco increased the gravitropic set-point angle in lateral roots, as well as root diameter and volume. The expression of auxin polar transport factors, PIN1 and PIN3, decreased and increased in CmRCC1-overexpressed plants, respectively. Yeast two-hybrid verification and luciferase complementation imaging assay showed that CmRCC1 interacts with CmLAZY1. Furthermore, the decreases in maximum quantum yield of PS II, the effective quantum yield of PS II, and electron transfer rate and the increases in quantum yield of nonregulated energy dissipation and malondialdehyde content were compromised in transgenic plants compared with wild-type plants under cold stress. The results suggest that CmRCC1 plays an important role in the regulation of root architecture and positively modulates cold tolerance.

The Analysis of Gravitropic Setpoint Angle Control in Plants.

The history of research on gravitropism has been largely confined to the primary root-shoot axis and to understanding how the typically vertical orientation observed there is maintained. Many lateral organs are gravitropic too and are often held at specific non-vertical angles relative to gravity. These so-called gravitropic setpoint angles (GSAs) are intriguing because their maintenance requires that root and shoot lateral organs are able to effect tropic growth both with and against the gravity vector. This chapter describes methods and considerations relevant to the investigation of mechanisms underlying GSA control.

Staging of Emerged Lateral Roots in Arabidopsis thaliana.

The root system in plants plays a fundamental role in water and nutrient uptake. Lateral roots emerge from the primary root (PR) and its directional organ growth allows the plant to strategically explore the surrounding area. Compared to the main root, lateral roots initially display a distinct gravitropic set point angle, which is established shortly after emergence. Here, we describe a unifying protocol for the morphological description and classification of emerged, young lateral roots.

Auxin in Root Development.

Root system architecture is an important determinant of below-ground resource capture and hence overall plant fitness. The plant hormone auxin plays a central role in almost every facet of root development from the cellular to the whole-root-system level. Here, using Arabidopsis as a model, we review the multiple gene signaling networks regulated by auxin biosynthesis, conjugation, and transport that underpin primary and lateral root development. We describe the role of auxin in establishing the root apical meristem and discuss how the tight spatiotemporal regulation of auxin distribution controls transitions between cell division, cell growth, and differentiation. This includes the localized reestablishment of mitotic activity required to elaborate the root system via the production of lateral roots. We also summarize recent discoveries on the effects of auxin and auxin signaling and transport on the control of lateral root gravitropic setpoint angle (GSA), a critical determinant of the overall shape of the root system. Finally, we discuss how environmental conditions influence root developmental plasticity by modulation of auxin biosynthesis, transport, and the canonical auxin signaling pathway.

LAZY1-LIKE-mediated gravity signaling pathway in root gravitropic set-point angle control.

Gravity signaling components contribute to the control of root gravitropic set-point angle through protein polarization relay within columella.

Transcriptome analysis of apple rootstocks with different stem-root gravitropic setpoint angles

ABSTRACT Our objective was to study the differentially expressed genes (DEGs) of apple (Malus. sp.) rootstock with different gravitropic setpoint angles (GSAs), during the formation of stem-root GSAs. The annual shoots of eight years hybrid seedlings of Malus pumila‘Ralls’ and M. sp. were used as experimental materials, and softwood cutting was carried out in mid-May. Different strains of adventitious root GSA were screened and shoot cuttings were carried out again the following year. The cuttings were randomly selected at 24 day after cutting, and then the transcriptome analysis was carried out. The results show that: (1) according to the difference of stem-root GSA of cutting seedlings, the seedlings of apple rootstock were divided into 3 groups: deep root type (SC), intermediate root type (ZC) and shallow root type (QC). (2) Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that 2613 genes from shallow- and deep-rooted rootstock, 2091 from deep- and intermediate-rooted rootstock, and 3114 from intermediate- and shallow-rooted rootstock, were annotated into 125 pathways. Among these pathways, plant hormone signal transduction, biosynthesis of secondary metabolites, and phenylpropanoid biosynthesis pathways were significantly enriched (P < 0.01).

Gravity-Sensing Tissues for Gravitropism Are Required for "Anti-Gravitropic" Phenotypes of Lzy Multiple Mutants in Arabidopsis.

Plant posture is controlled by various environmental cues, such as light, temperature, and gravity. The overall architecture is determined by the growth angles of lateral organs, such as roots and branches. The branch growth angle affected by gravity is known as the gravitropic setpoint angle (GSA), and it has been proposed that the GSA is determined by balancing two opposing growth components: gravitropism and anti-gravitropic offset (AGO). The molecular mechanisms underlying gravitropism have been studied extensively, but little is known about the nature of the AGO. Recent studies reported the importance of LAZY1-LIKE (LZY) family genes in the signaling process for gravitropism, such that loss-of-function mutants of LZY family genes resulted in reversed gravitropism, which we term it here as the “anti-gravitropic” phenotype. We assume that this peculiar phenotype manifests as the AGO due to the loss of gravitropism, we characterized the “anti-gravitropic” phenotype of Arabidopsis lzy multiple mutant genetically and physiologically. Our genetic interaction analyses strongly suggested that gravity-sensing cells are required for the “anti-gravitropic” phenotype in roots and lateral branches. We also show that starch-filled amyloplasts play a significant role in the “anti-gravitropic” phenotype, especially in the root of the lzy multiple mutant.

Asymmetric cytokinin signaling opposes gravitropism in roots.

Plants depend on gravity to provide the constant landmark for downward root growth and upward shoot growth. The phytohormone auxin and its cell‐to‐cell transport machinery are central determinants ensuring gravitropic growth. Statolith sedimentation toward gravity is sensed in specialized cells. This positional cue is translated into the polar distribution of PIN auxin efflux carriers at the plasma membrane, leading to asymmetric auxin distribution and consequently, differential growth and organ bending. While we have started to understand the general principles of how primary organs execute gravitropism, we currently lack basic understanding of how lateral plant organs can defy gravitropic responses. Here we briefly review the establishment of the oblique gravitropic set point angle in lateral roots and particularly discuss the emerging role of asymmetric cytokinin signaling as a central anti‐gravitropic signal. Differential cytokinin signaling is co‐opted in gravitropic lateral and hydrotropic primary roots to counterbalance gravitropic root growth.

CEP receptor signalling controls root system architecture in Arabidopsis and Medicago.

Root system architecture (RSA) influences the effectiveness of resources acquisition from soils but the genetic networks that control RSA remain largely unclear. We used rhizoboxes, X-ray computed tomography, grafting, auxin transport measurements and hormone quantification to demonstrate that Arabidopsis and Medicago CEP (C-TERMINALLY ENCODED PEPTIDE)-CEP RECEPTOR signalling controls RSA, the gravitropic set-point angle (GSA) of lateral roots (LRs), auxin levels and auxin transport. We showed that soil-grown Arabidopsis and Medicago CEP receptor mutants have a narrower RSA, which results from a steeper LR GSA. Grafting showed that CEPR1 in the shoot controls GSA. CEP receptor mutants exhibited an increase in rootward auxin transport and elevated shoot auxin levels. Consistently, the application of auxin to wild-type shoots induced a steeper GSA and auxin transport inhibitors counteracted the CEP receptor mutant's steep GSA phenotype. Concordantly, CEP peptides increased GSA and inhibited rootward auxin transport in wild-type but not in CEP receptor mutants. The results indicated that CEP-CEP receptor-dependent signalling outputs in Arabidopsis and Medicago control overall RSA, LR GSA, shoot auxin levels and rootward auxin transport. We propose that manipulating CEP signalling strength or CEP receptor downstream targets may provide means to alter RSA.

AtDRO1 is nuclear localized in root tips under native conditions and impacts auxin localization

DEEPER ROOTING 1 (DRO1) contributes to the downward gravitropic growth trajectory of roots upstream of lateral auxin transport in monocots and dicots. Loss of DRO1 function leads to horizontally oriented lateral roots and altered gravitropic set point angle, while loss of all three DRO family members results in upward, vertical root growth. Here, we attempt to dissect the roles of AtDRO1 by analyzing expression, protein localization, auxin gradient formation, and auxin responsiveness in the atdro1 mutant. Current evidence suggests AtDRO1 is predominantly a membrane-localized protein. Here we show that VENUS-tagged AtDRO1 driven by the native AtDRO1 promoter complemented an atdro1 Arabidopsis mutant and the protein was localized in root tips and detectable in nuclei. atdro1 primary and lateral roots showed impairment in establishing an auxin gradient upon gravistimulation as visualized with DII-VENUS, a sensor for auxin signaling and proxy for relative auxin distribution. Additionally, PIN3 domain localization was not significantly altered upon gravistimulation in atdro1 primary and lateral roots. RNA-sequencing revealed differential expression of known root development-related genes in atdro1 mutants. atdro1 lateral roots were able to respond to exogenous auxin and AtDRO1 gene expression levels in root tips were unaffected by the addition of auxin. Collectively, the data suggest that nuclear localization may be important for AtDRO1 function and suggests a more nuanced role for DRO1 in regulating auxin-mediated changes in lateral branch angle.Key messageDEEPER ROOTING 1 (DRO1) when expressed from its native promoter is predominately localized in Arabidopsis root tips, detectable in nuclei, and impacts auxin gradient formation.

Polar recruitment of RLD by LAZY1-like protein during gravity signaling in root branch angle control

In many plant species, roots maintain specific growth angles relative to the direction of gravity, known as gravitropic set point angles (GSAs). These contribute to the efficient acquisition of water and nutrients. AtLAZY1/LAZY1-LIKE (LZY) genes are involved in GSA control by regulating auxin flow toward the direction of gravity in Arabidopsis. Here, we demonstrate that RCC1-like domain (RLD) proteins, identified as LZY interactors, are essential regulators of polar auxin transport. We show that interaction of the CCL domain of LZY with the BRX domain of RLD is important for the recruitment of RLD from the cytoplasm to the plasma membrane by LZY. A structural analysis reveals the mode of the interaction as an intermolecular β-sheet in addition to the structure of the BRX domain. Our results offer a molecular framework in which gravity signal first emerges as polarized LZY3 localization in gravity-sensing cells, followed by polar RLD1 localization and PIN3 relocalization to modulate auxin flow.

Transcriptional regulation of MdPIN3 and MdPIN10 by MdFLP during apple self-rooted stock adventitious root gravitropism

BackgroundThe close planting of dwarfing self-rooted rootstocks is currently a widely used method for apple production; however, self-rooted rootstocks are weak with shallow roots and poor grounding. Therefore, understanding the molecular mechanisms that establish the gravitropic set-point angles (GSAs) of the adventitious roots of self-rooted apple stocks is important for developing self-rooted apple rootstock cultivars with deep roots.ResultsWe report that the apple FOUR LIPS (MdFLP), an R2R3-MYB transcription factor (TF), functions in establishing the GSA of the adventitious roots of self-rooted apple stocks in response to gravity. Biochemical analyses demonstrate that MdFLP directly binds to the promoters of two auxin efflux carriers, MdPIN3 and MdPIN10, that are involved in auxin transport, activates their transcriptional expression, and thereby promotes the development of adventitious roots in self-rooted apple stocks. Additionally, the apple auxin response factor MdARF19 influences the expression of those auxin efflux carriers and the establishment of the GSA of adventitious roots of apple in response to gravity by directly activating the expression of MdFLP.ConclusionOur findings provide new insights into the transcriptional regulation of MdFLP by the auxin response factor MdARF19 in the regulation of the GSA of adventitious roots of self-rooted apple stocks in response to gravity.