Root-tip-mediated inhibition of hydrotropism is accompanied with the suppression of asymmetric expression of auxin-inducible genes in response to moisture gradients in cucumber roots.
ABSTRACT: In cucumber seedlings, gravitropism interferes with hydrotropism, which results in the nearly complete inhibition of hydrotropism under stationary conditions. However, hydrotropic responses are induced when the gravitropic response in the root is nullified by clinorotation. Columella cells in the root cap sense gravity, which induces the gravitropic response. In this study, we found that removing the root tip induced hydrotropism in cucumber roots under stationary conditions. The application of auxin transport inhibitors to cucumber seedlings under stationary conditions suppressed the hydrotropic response induced by the removal of the root tip. To investigate the expression of genes related to hydrotropism in de-tipped cucumber roots, we conducted transcriptome analysis of gene expression by RNA-Seq using seedlings exhibiting hydrotropic and gravitropic responses. Of the 21 and 45 genes asymmetrically expressed during hydrotropic and gravitropic responses, respectively, five genes were identical. Gene ontology (GO) analysis indicated that the category auxin-inducible genes was significantly enriched among genes that were more highly expressed in the concave side of the root than the convex side during hydrotropic or gravitropic responses. Reverse transcription followed by quantitative polymerase chain reaction (RT-qPCR) analysis revealed that root hydrotropism induced under stationary conditions (by removing the root tip) was accompanied by the asymmetric expression of several auxin-inducible genes. However, intact roots did not exhibit the asymmetric expression patterns of auxin-inducible genes under stationary conditions, even in the presence of a moisture gradient. These results suggest that the root tip inhibits hydrotropism by suppressing the induction of asymmetric auxin distribution. Auxin transport and distribution not mediated by the root tip might play a role in hydrotropism in cucumber roots.
Project description:Roots are highly plastic and can acclimate to heterogeneous and stressful conditions. However, there is little knowledge of the effect of moisture gradients on the mechanisms controlling root growth orientation and branching, and how this mechanism may help plants to avoid drought responses. The aim of this study was to isolate mutants of Arabidopsis thaliana with altered hydrotropic responses. Here, altered hydrotropic response 1 (ahr1), a semi-dominant allele segregating as a single gene mutation, was characterized. ahr1 directed the growth of its primary root towards the source of higher water availability and developed an extensive root system over time. This phenotype was intensified in the presence of abscisic acid and was not observed if ahr1 seedlings were grown in a water stress medium without a water potential gradient. In normal growth conditions, primary root growth and root branching of ahr1 were indistinguishable from those of the wild type (wt). The altered hydrotropic growth of ahr1 roots was confirmed when the water-rich source was placed at an angle of 45° from the gravity vector. In this system, roots of ahr1 seedlings grew downward and did not display hydrotropism; however, in the presence of cytokinins, they exhibited hydrotropism like those of the wt, indicating that cytokinins play a critical role in root hydrotropism. The ahr1 mutant represents a valuable genetic resource for the study of the effects of cytokinins in the differential growth of hydrotropism and control of lateral root formation during the hydrotropic response.
Project description:The <i>hab1-1abi1-2abi2-2pp2ca-1</i> quadruple mutant (<i>Qabi2-2</i>) seedlings lacking key negative regulators of ABA signaling, namely, clade A protein phosphatases type 2C (PP2Cs), show more apoplastic H<sup>+</sup> efflux in roots and display an enhanced root growth under normal medium or water stress medium compared to the wild type. The presence of low ABA concentration (0.1 micromolar), inhibiting PP2C activity via monomeric ABA receptors, enhances root apoplastic H<sup>+</sup> efflux and growth of the wild type, resembling the <i>Qabi2-2</i> phenotype in normal medium. <i>Qabi2-2</i> seedlings also demonstrate increased hydrotropism compared to the wild type in obliquely-oriented hydrotropic experimental system, and asymmetric H<sup>+</sup> efflux in root elongation zone is crucial for root hydrotropism. Moreover, we reveal that <i>Arabidopsis</i> ABA-insensitive 1, a key PP2C in ABA signaling, interacts directly with the C terminus of <i>Arabidopsis</i> plasma membrane H<sup>+</sup>-dependent adenosine triphosphatase 2 (AHA2) and dephosphorylates its penultimate threonine residue (Thr<sup>947</sup>), whose dephosphorylation negatively regulates AHA2.
Project description:Lateral roots are initiated postembryonically in response to environmental cues, enabling plants to explore efficiently their underground environment. However, the mechanisms by which the environment determines the position of lateral root formation are unknown. In this study, we demonstrate that in Arabidopsis thaliana lateral root initiation can be induced mechanically by either gravitropic curvature or by the transient bending of a root by hand. The plant hormone auxin accumulates at the site of lateral root induction before a primordium starts to form. Here we describe a subcellular relocalization of PIN1, an auxin transport protein, in a single protoxylem cell in response to gravitropic curvature. This relocalization precedes auxin-dependent gene transcription at the site of a new primordium. Auxin-dependent nuclear signaling is necessary for lateral root formation; arf7/19 double knock-out mutants normally form no lateral roots but do so upon bending when the root tip is removed. Signaling through arf7/19 can therefore be bypassed by root bending. These data support a model in which a root-tip-derived signal acts on downstream signaling molecules that specify lateral root identity.
Project description:<h4>Background and aims</h4>Root hydrotropism is a response to water-potential gradients that makes roots bend towards areas of higher water potential. The gene MIZU-KUSSEI1 (MIZ1) that is essential for hydrotropism in Arabidopsis roots has previously been identified. However, the role of root hydrotropism in plant growth and survival under natural conditions has not yet been proven. This study assessed how hydrotropic response contributes to drought avoidance in nature.<h4>Methods</h4>An experimental system was established for the study of Arabidopsis hydrotropism in soil. Characteristics of hydrotropism were analysed by comparing the responses of the miz1 mutant, transgenic plants overexpressing MIZ1 (MIZ1OE) and wild-type plants.<h4>Key results</h4>Wild-type plants developed root systems in regions with higher water potential, whereas the roots of miz1 mutant plants did not show a similar response. This pattern of root distribution induced by hydrotropism was more pronounced in MIZ1OE plants than in wild-type plants. In addition, shoot biomass and the number of plants that survived under drought conditions were much greater in MIZ1OE plants.<h4>Conclusions</h4>These results show that hydrotropism plays an important role in root system development in soil and contributes to drought avoidance, which results in a greater yield and plant survival under water-limited conditions. The results also show that MIZ1 overexpression can be used for improving plant productivity in arid areas.
Project description:Branching by de novo formation of lateral roots along the primary root of Arabidopsis seedlings follows a complex longitudinal and transverse pattern. How this pattern is generated is presently under debate. The 'bending hypothesis' proposes that lateral root primordia are initiated by a local accumulation of auxin at the convex side of bends resulting from deflections through obstacles, gravitropic bending, or other means. In contrast, the 'oscillation hypothesis' proposes the existence of an endogenous clock-type oscillator mechanism producing periodic pulses of gene expression in the root tip that determine the future sites of primordium initiation. Here we report physiological experiments dissecting periodic priming signals, pre-disposing the root to rhythmic lateral root formation, from bending-mediated signals responsible for the subsequent positioning of their initiation along the growing root. While the frequency of lateral roots can be promoted by auxin in the mature root, their positioning follows a pre-formed pattern determined by previous bending. Both types of signals turn out to be necessary, complementary components in an integrating concept of lateral root patterning.
Project description:Factors that affect the direction of root growth in response to environmental signals influence crop productivity. We analyzed the root tropic responses of thioredoxin (trxs), thigmotropic (wav2-1), and hydrotropic (ahr1 and nhr1) Arabidopsis thaliana mutants treated with low concentrations of paraquat (PQ), which induces mild oxidative stress, and established a new method for evaluating root waviness (root bending effort, RBE). This method estimates root bending by measuring and summing local curvature over the whole length of the root, regardless of the asymmetry of the wavy pattern under thigmostimulation. In roots of the wav2-1 mutant, but not in those of the trxs and ahr1 mutants, RBE was significantly inhibited under mild oxidative stress. Thigmotropic stimulation of wav2-1 mutant roots, with or without PQ treatment, showed high levels of reactive oxygen species fluorescence, in contrast to roots of the ahr1 mutant. Furthermore, PQ inhibited root growth in all genotypes tested, except in the wav2-1 mutant. In a hydrotropism assay of the trxs and wav2-1 mutants, root growth behavior was similar to the wild type with and without PQ, while the root growth of ahr1 and nhr1 mutants was diminished with PQ. These results indicate that hydrotropic and thigmotropic mutants respond differently to exogenous PQ, depending on the tropic stimulus perceived. Therefore, the mechanisms underlying hydrotropism and thigmotropism may differ.
Project description:Arabidopsis roots sense the moisture gradient in soils and grow toward an area with higher water potential - a process called hydrotropism. Our previous study has shown that the apoplastic proton extrusion in root tips is influenced by brassinosteroids (BRs) receptor BR-INSENSITIVE1 (BRI1) and is crucial for hydrotropic response in Arabidopsis. Here we show that BRI1 interacts directly not only with Arabidopsis plasma membrane H+-ATPase 2 (AHA2) but also with Arabidopsis plasma membrane H+-ATPase 7 (AHA7). Therefore, BRI1 may affect hydrotropic response via regulating the activities of AHA2 and AHA7.
Project description:Roots display hydrotropism in response to moisture gradients, which is thought to be important for controlling their growth orientation, obtaining water, and establishing their stand in the terrestrial environment. However, the molecular mechanism underlying hydrotropism remains unknown. Here, we report that roots of the Arabidopsis mutant mizu-kussei1 (miz1), which are impaired in hydrotropism, show normal gravitropism and elongation growth. The roots of miz1 plants showed reduced phototropism and a modified wavy growth response. There were no distinct differences in morphological features and root structure between miz1 and wild-type plants. These results suggest that the pathway inducing hydrotropism is independent of the pathways used in other tropic responses. The phenotype results from a single recessive mutation in MIZ1, which encodes a protein containing a domain (the MIZ domain) that is highly conserved among terrestrial plants such as rice and moss. The MIZ domain was not found in known genomes of organisms such as green algae, red algae, cyanobacteria, or animals. We hypothesize that MIZ1 has evolved to play an important role in adaptation to terrestrial life because hydrotropism could contribute to drought avoidance in higher plants. In addition, a pMIZ1::GUS fusion gene was expressed strongly in columella cells of the root cap but not in the elongation zone, suggesting that MIZ1 functions in the early phase of the hydrotropic response.
Project description:The Arabidopsis AtCRK5 protein kinase is involved in the establishment of the proper auxin gradient in many developmental processes. Among others, the At<i>crk5-1</i> mutant was reported to exhibit a delayed gravitropic response via compromised PIN2-mediated auxin transport at the root tip. Here, we report that this phenotype correlates with lower superoxide anion (O<sub>2</sub><sup>•-</sup>) and hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>) levels but a higher nitric oxide (NO) content in the mutant root tips in comparison to the wild type (AtCol-0). The oxidative stress inducer paraquat (PQ) triggering formation of O<sub>2</sub><sup>•-</sup> (and consequently, H<sub>2</sub>O<sub>2</sub>) was able to rescue the gravitropic response of At<i>crk5-1</i> roots. The direct application of H<sub>2</sub>O<sub>2</sub> had the same effect. Under gravistimulation, correct auxin distribution was restored (at least partially) by PQ or H<sub>2</sub>O<sub>2</sub> treatment in the mutant root tips. In agreement, the redistribution of the PIN2 auxin efflux carrier was similar in the gravistimulated PQ-treated mutant and untreated wild type roots. It was also found that PQ-treatment decreased the endogenous NO level at the root tip to normal levels. Furthermore, the mutant phenotype could be reverted by direct manipulation of the endogenous NO level using an NO scavenger (cPTIO). The potential involvement of AtCRK5 protein kinase in the control of auxin-ROS-NO-PIN2-auxin regulatory loop is discussed.
Project description:The phenomenon of plant root tips sensing moisture gradient in soil and growing towards higher water potential is designated as root hydrotropism, which is critical for plants to survive when water is a limited factor. Molecular mechanisms regulating such a fundamental process, however, are largely unknown. Here we report our identification that cytokinins are key signaling molecules directing root growth orientation in a hydrostimulation (moisture gradient) condition. Lower water potential side of the root tip shows more cytokinin response relative to the higher water potential side. Consequently, two cytokinin downstream type-A response regulators, ARR16 and ARR17, were found to be up-regulated at the lower water potential side, causing increased cell division in the meristem zone, which allows the root to bend towards higher water potential side. Genetic analyses indicated that various cytokinin biosynthesis and signaling mutants, including the arr16 arr17 double mutant, are significantly less responsive to hydrostimulation. Consistently, treatments with chemical inhibitors interfering with either cytokinin biosynthesis or cell division completely abolished root hydrotropic response. Asymmetrically induced expression of ARR16 or ARR17 effectively led to root bending in both wild-type and miz1, a previously known hydrotropism-defective mutant. These data demonstrate that asymmetric cytokinin distribution is a primary determinant governing root hydrotropism.