Circumnutation of rice coleoptiles: its occurrence, regulation by phytochrome, and relationship with gravitropism

It has been found that coleoptiles of dark-grown rice (Oryza sativa L.) seedlings undergo regular circumnutation in circular orbits with periods of about 180 min. Both clockwise and counter-clockwise movements were observed, but individual coleoptiles continued to rotate only in one direction. Light-grown seedlings did not show circumnutation. In fact, dark-grown seedlings were found to cease circumnutating in response to a pulse of red light (R). This light-induced inhibition of circumnutation was demonstrated to involve both a FR-inducible very-low-fluence response, solely mediated by phytochrome A, and a FR-reversible low-fluence response, mediated by phytochrome B and/or C. The R-induced inhibition of circumnutation showed temporal agreement with the R-induced inhibition of coleoptile growth, suggesting that the former results from the latter. However, about 25% of growth activity remained after R treatment, indicating that circumnutation is more specifically regulated by phytochrome. The R-treated coleoptile showed gravitropism. Investigation of the growth differential for gravitropic curvature revealed that gravitropic responsiveness was rather enhanced by R. The results suggested that gravitropism is not a cause of circumnutation. It remained probable, however, that gravity perception is a part of the mechanism of circumnutation. It is speculated that the circumnutation investigated aids the seedling shoot in growing through the soil.     

Identification of the gravitropism-related rice gene LAZY1 and elucidation of LAZY1-dependent and -independent gravity signaling pathways

We identified the gene responsible for three allelic lazy1 mutations of Japonica rice (Oryza sativa L.) by map-based cloning, complementation and RNA interference. Sequence analysis and database searches indicated that the wild-type gene (LAZY1) encodes a novel and unique protein (LAZY1) and that rice has no homologous gene. Two lazy1 mutants were LAZY1 null. Confirming and advancing the previously reported results on lazy1 mutants, we found the following. (i) Gravitropism is impaired, but only partially, in lazy1 coleoptiles. (ii) Circumnutation, observed in dark-grown coleoptiles, is totally absent from lazy1 coleoptiles. (iii) Primary roots of lazy1 mutants show normal gravitropism and circumnutation. (iv) LAZY1 is expressed in a tissue-specific manner in gravity-sensitive shoot tissues (i.e. coleoptiles, leaf sheath pulvini and lamina joints) and is little expressed in roots. (v) The gravitropic response of lazy1 coleoptiles is kinetically separable from that absent from lazy1 coleoptiles. (vi) Gravity-induced lateral translocation of auxin, found in wild-type coleoptiles, does not occur in lazy1 coleoptiles. Based on the genetic and physiological evidence obtained, it is concluded that LAZY1 is specifically involved in shoot gravitropism and that LAZY1-dependent and -independent signaling pathways occur in coleoptiles. It is further concluded that, in coleoptiles, only the LAZY1-dependent gravity signaling involves asymmetric distribution of auxin between the two lateral halves and is required for circumnutation.     

Circumnutational movement in rice coleoptiles involves the gravitropic response: analysis of an agravitropic mutant and space‐grown seedlings

Plants exhibit helical growth movements known as circumnutation in growing organs. Some studies indicate that circumnutation involves the gravitropic response, but this notion is a matter of debate. Here, using the agravitropic rice mutant lazy1 and space‐grown rice seedlings, we found that circumnutation was reduced or lost during agravitropic growth in coleoptiles. Coleoptiles of wild‐type rice exhibited circumnutation in the dark, with vigorous oscillatory movements during their growth. The gravitropic responses in lazy1 coleoptiles differed depending on the growth stage, with gravitropic responses detected during early growth and agravitropism during later growth. The nutation‐like movements observed in lazy1 coleoptiles at the early stage of growth were no longer detected with the disappearance of the gravitropic response. To verify the relationship between circumnutation and gravitropic responses in rice coleoptiles, we conducted spaceflight experiments in plants under microgravity conditions on the International Space Station. Wild‐type rice seeds were germinated, and the resulting seedlings were grown under microgravity or a centrifuge‐generated 1 g environment in space. We began filming the seedlings 2 days after seed imbibition and obtained images of seedling growth every 15 min. The seed germination rate in space was 92–100% under both microgravity and 1 g conditions. LED‐synchronized flashlight photography induced an attenuation of coleoptile growth and circumnutational movement due to cumulative light exposure. Nevertheless, wild‐type rice coleoptiles still showed circumnutational oscillations under 1 g but not microgravity conditions. These results support the idea that the gravitropic response is involved in plant circumnutation.     

Gravitropic response plays an important role in the nutational movements of the shoots of Pharbitis nil and Arabidopsis thaliana

The shoots of a Japanese strain of morning glory ( Pharbitis nil  ) called 'Shidare-asagao' display agravitropic and weeping growth. It has been shown that this shoot agravitropism may be due to the defective differentiation of endodermal cells that contain statoliths. Roots of the weeping morning glory show normal responsiveness to gravity and the shoots are positively phototropic. Shoots of the morning glory cultivar Violet used as a wild type exhibited distinct circumnutation with circular movements that increase as the plants grow. In weeping morning glory, however, nutation was limited to slight back and forth or side to side movements. To determine whether endodermal cells participate in circumnutation through a function that is independent of their role in gravitropism, the nutational movements of various gravitropic mutants of Arabidopsis thaliana were compared. The inflorescences of wild-type Arabidopsis showed relatively large circular movements. Inflorescences of the pgm-1 mutant, which is defective in starch synthesis, showed reduced nutation. Even more seriously affected were the sgr1-1 / scr-3 and sgr7-1 / shr-2 mutants, which are defective in endodermal cell differentiation, and the auxin-resistant axr2-1 mutant showed no significant nutational movements at all. 1- N -naphthylphthalamic acid (NPA) could inhibit Violet circumnutation, supporting the notion that auxin participates in circumnutation. Thus, the gravitropic response is an essential component in plant shoot circumnutation. Endodermal cells are involved in such circumnutation possibly because of their role in inducing the gravitropic response.     

Transport of Indole-3-Acetic Acid during Gravitropism in Intact Maize Coleoptiles

We have investigated the transport of tritiated indole-3-acetic acid (IAA) in intact, red light-grown maize (Zea mays) coleoptiles during gravitropic induction and the subsequent development of curvature. This auxin is transported down the length of gravistimulated coleoptiles at a rate comparable to that in normal, upright plants. Transport is initially symmetrical across the coleoptile, but between 30 and 40 minutes after plants are turned horizontal a lateral redistribution of the IAA already present in the transport stream occurs. By 60 minutes after the beginning of the gravitropic stimulus, the ratio of tritiated tracer auxin in the lower half with respect to the upper half is approximately 2:1. The redistribution of growth that causes gravitropic curvature follows the IAA redistribution by 5 or 10 minutes at the minimum in most regions of the coleoptile. Immobilization of tracer auxin from the transport stream during gravitropism was not detectable in the most apical 10 millimeters. Previous reports have shown that in intact, red light-grown maize coleoptiles, endogenous auxin is limiting for growth, the tissue is linearly responsive to linearly increasing concentrations of small amounts of added auxin, and the lag time for the stimulation of straight growth by added IAA is approximately 8 or 9 minutes (TI Baskin, M Iino, PB Green, WR Briggs [1985] Plant Cell Environ 8: 595-603; TI Baskin, WR Briggs, M Iino [1986] Plant Physiol 81: 306-309). We conclude that redistribution of IAA in the transport stream occurs in maize coleoptiles during gravitropism, and is sufficient in degree and timing to be the immediate cause of gravitropic curvature.     

Gravitropism of maize and rice coleoptiles: dependence on the stimulation angle

Gravitropism of maize and rice coleoptiles was investigated with respect to its dependence on the angle of displacement or the initial stimulation angle (ISA). Close examination of curvature kinetics and the response to a drop in stimulation angle (SA) indicated that the gravtropic response during an early but substantial part of the curvature development is directly related to the ISA, there being no effect of the reduction of SA resulting from the curvature response itself. On the basis of this finding, the relationship between the steady SA and the curvature rate was determined. In maize, the curvature rate increased linearly with the sines of SAs up to an SA of 90 degrees. Rice coleoptiles, however, showed a saturation curve in the same range of SAs. The saturation profile was nearly identical between coleoptiles grown in air and those submerged in water, although the latter elongated much faster. Rice coleoptiles appeared to be far more sensitive to gravity than maize coleoptiles. It is concluded that the sensitivity to gravity, assessed through dependence on ISA, is a property inherent to a given gravitropic organ. Long-term measurements of curvature indicated that the coleoptiles bend back past the vertical. This overshooting was marked in submerged rice coleoptiles.     

Effects of Chlorpromazine on Gravitropism in Avena Coleoptiles

Chlorpromazine (CPZ), an inhibitor of the calcium-activatedform of calmodulin, is readily taken up by the roots of intactoat seedlings but poorly translocated from the roots to thecoleoptile of these plants. However, plants repeatedly rotatedthrough solutions containing low concentrations of CPZ (10^–8–10^–5M)are infiltrated, and under these conditions, CPZ significantlyinhibits the negative gravitropic response of the coleoptilewithout retarding elongation growth. This effect is observablein ‘decapitated’ (apical 1–2 mm removed) coleoptilesections and in intact whole coleoptiles. If exogenous auxinis supplied to the decapitated sections, both their growth ratesand gravitropic responsiveness are increased and, under theseconditions, CPZ can reduce the gravitropic curvature withoutreducing the overall growth rate. These results are discussedin relation to the possible role of calmodulin-dependent calcium-ionpumps in gravitropism. chlorpromazine, gravitropism, calmodulin, calcium, oat, Avena sativa     

Gravitropic responses of wild-type and mutant strains of the moss Physcomitrella patens

The gravitropic responses of dark-grown caulonemata and gametophores of wild-type and mutant strains of the moss Physcomitrella patens have been investigated. In the wild-type both caulonemata and gametophores show negative orthogravitropism. No gravitropic response is observed when plants are rotated slowly on a clinostat and the inductive effect of gravity can be replaced by centrifugal force. The gravitropic response of caulonemanta is biphasic, consisting of an initial phase producing a bend of about 20 degrees within 12 h of 90 degrees reorientation and a subsequent slower phase leading to completion of the 90 degrees curvature. No obvious sedimentation of statoliths accompanies this response. Several mutants have been isolated that are either partially or completely impaired in caulonemal gravitropism and one mutant shows a positive gravitropic response. Complementation analysis using somatic hybrids obtained following protoplast fusion indicates that at least three genes can mutate to give an altered gravitropic phenotype. None of these mutants is altered in gametophore gravitropism, suggesting that the gravitropic response of caulonemal filaments may require at least some gene products that are not required for the response of the multicellular gametophores. One class of mutant with impaired caulonemal gravitropism shows a pleiotropic alteration in leaf shape.     

Nuclear magnetic resonance imaging in studies of gravitropism in soil mixtures

Gravitropic responses of oat coleoptiles were measured in different growth media; humid air, natural soil and artificial soil (glass beads). The oat coleoptiles in soil and glass beads were monitored by NMR imaging, while those in humid air were imaged in darkness with an infrared-sensitive charge-coupled device (CCD) camera. The present study shows for the first time that gravitropic experiments can be performed in artificial soil using NMR imaging as a convenient and suitable recording method. Not only was it possible to follow the gravitropic curvatures in natural soil, but the artificial soil allowed plant images of sufficient spatial and temporal resolution to be recorded. The advantages of using artificial soil in magnetic resonance imaging studies are that the iron content of glass beads is very low compared with natural soil, and that the artificial soil matrix can easily be standardized with regard to particle size distribution and nutrient content. Two types of glass beads were used, the diameter of the small and the large beads being 300–400 and 420–840 μm, respectively. The growth rate of the coleoptiles in soil and in big beads was roughly the same and only slightly lower than in humid air, whereas small beads reduced the growth rate by approx. 16%. The bending rate of the coleoptiles during the gravitropic response was reduced by c. 65% in soil and 75% in bead mixtures relative to bending in air. It should be noted, however, that the maximum curvature of the coleoptile tip was of the same order in all cases, about 35°. This value may represent the largest possible curvature of the organ. The potential of NMR imaging to study how plant organs penetrate the soil under the influence of gravitropism, mechanical impedance and thigmotropism is also discussed.     

Ethylene modulates flavonoid accumulation and gravitropic responses in roots of Arabidopsis

Plant organs change their growth direction in response to reorientation relative to the gravity vector. We explored the role of ethylene in Arabidopsis (Arabidopsis thaliana) root gravitropism. Treatment of wild-type Columbia seedlings with the ethylene precursor 1-aminocyclopropane carboxylic acid (ACC) reduced root elongation and gravitropic curvature. The ethylene-insensitive mutants ein2-5 and etr1-3 had wild-type root gravity responses, but lacked the growth and gravity inhibition by ACC found in the wild type. We examined the effect of ACC on tt4(2YY6) seedlings, which have a null mutation in the gene encoding chalcone synthase, the first enzyme in flavonoid synthesis. The tt4(2YY6) mutant makes no flavonoids, has elevated indole-3-acetic acid transport, and exhibits a delayed gravity response. Roots of tt4(2YY6), the backcrossed line tt4-2, and two other tt4 alleles had wild-type sensitivity to growth inhibition by ACC, whereas the root gravitropic curvature of these tt4 alleles was much less inhibited by ACC than wild-type roots, suggesting that ACC may reduce gravitropic curvature by altering flavonoid synthesis. ACC treatment induced flavonoid accumulation in root tips, as judged by a dye that becomes fluorescent upon binding flavonoids in wild type, but not in ein2-5 and etr1-3. ACC also prevented a transient peak in flavonoid synthesis in response to gravity. Together, these experiments suggest that elevated ethylene levels negatively regulate root gravitropism, using EIN2- and ETR1-dependent pathways, and that ACC inhibition of gravity response occurs through altering flavonoid synthesis.     

Lazy rice in space: gravity regulates helical movement in plants

Circumnutation, the helical movement of organs, has been observed in diverse species of land plants. Whether circumnutation arises purely from internal growth oscillations or as a response to exogenous forces such as gravity is a subject of active debate. By observing rice seedlings grown under microgravity at the International Space Station (ISS) and analyzing the agravitropic lazy1 mutant, Kobayashi et al. (2019) propose gravity as the causal force that regulates circumnutation of rice coleoptiles.     

Novel software for analysis of root gravitropism: comparative response patterns of Arabidopsis wild-type and axr1 seedlings

In an earlier study (Evans, Ishikawa & Estelle 1994, Planta 194, 215-222) we used a video digitizer system to compare the kinetics of auxin action on root elongation in wild-type seedlings and seedlings of auxin response mutants of Arabidopsis thaliana (L.) Heynh. We have since modified the system software to allow determination of elongation on opposite sides of vertical or gravistimulated roots and to allow continuous measurement of the angle of orientation of sequential subsections of the root during the response. We used this technology to compare the patterns of differential growth that generate curvature in roots of the Columbia ecotype and in the mutants axr1-3, axr1-12 and axr2, which show reduced gravitropic responsiveness and reduced sensitivity to inhibition by auxin. The pattern of differential growth during gravitropism differed in roots of wild-type and axr1 seedlings. In wild-type roots, initial curvature resulted from differential inhibition of elongation in the distal elongation zone (DEZ). This was followed by an acceleration of elongation along the top side of the DEZ. In roots of axr1-3, curvature resulted from differential stimulation of elongation whereas in roots of axr1-12 the response was variable. Roots of axr2 did not exhibit gravitropic curvature. The observation that the pattern of differential growth causing curvature is dramatically altered by a change in sensitivity to auxin is consistent with the classical Cholodny-Went theory of gravitropism which maintains that differential growth patterns induced by gravistimulation are mediated primarily by gravi-induced shifts in auxin distribution. The new technology introduced with this report allows automated determination of stimulus response patterns in the small but experimentally popular roots of Arabidopsis.     

Callose deposition during gravitropism ofZea mays andPisum sativum and its inhibition by 2-deoxy-D-glucose

In etiolated corn (Zea mays L.) and etiolated pea (Pisum sativum L.) seedlings, a gravitropic stimulation induces the deposition of callose. In the corn coleoptiles this occurs within 5 min of gravity stimulation, and prior to the beginning of curvature. Both gravitropic curvature and callose deposition reach their maxima by 12 h. Within the first 2 h more callose is deposited on the upper (concave) side, but after 2–3 h, this deposition pattern is reversed. An inhibitor of protein glycosylation, 2-deoxy-d-glucose (DDG), inhibits callose production and considerably retards gravitropic bending in both species of plants. Mannose can relieve the inhibition of gravitropic bending by DDG. The pea mutant Ageotropum, which does not respond to gravity when etiolated, also fails to produce callose in response to a gravitic stimulus. These correlations indicate that callose deposition may be a biochemical component of gravitropism in plant shoots.Abbreviation DDG 2-deoxy-d-glucose     

Tropisms of Avena coleoptiles: sine law for gravitropism,exponential law for photogravitropic equilibrium

The quantitative relation between gravitropism and phototropism was analyzed for light-grown coleoptiles of Avena sativa (L.). With respect to gravitropism the coleoptiles obeyed the sine law. To study the interaction between light and gravity, coleoptiles were inclined at variable angles and irradiated for 7 h with unilateral blue light (466 nm) impinging at right angles relative to the axis of the coleoptile. The phototropic stimulus was applied from the side opposite to the direction of gravitropic bending. The fluence rate that was required to counteract the negative gravitropism increased exponentially with the sine of the inclination angle. To achieve balance, a linear increase in the gravitropic stimulus required compensation by an exponential increase in the counteracting phototropic stimulus. The establishment of photogravitropic equilibrium during continuous unilateral irradiation is thus determined by two different laws: the well-known sine law for gravitropism and a novel exponential law for phototropism described in this work.     

Spatial organization of the gravitropic response in plants: applicability of the revised local curvature distribution model to Triticum aestivum coleoptiles

The revised local curvature distribution model, which provides accurate computer simulations of the gravitropic response of mushroom stems, was found to produce accurate simulations of the gravitropic reaction of wheat ( Triticum aestivum ) coleoptiles. The key feature of the mathematical model that enables it to approach universality of application is the assumption that the stem has an autonomic straightening reaction (curvature compensation or 'autotropism'). In the model, the local bending rate for any segment of the organ is determined by the difference between the 'bending signal' (generated by the gravitropic signal perception system) and a 'straightening signal' (which is proportional to the local curvature of the segment). The model reveals three major differences between the gravitropic reactions of wheat coleoptiles and Coprinus mushroom stems. First, in Coprinus , the capacity for autonomic straightening is much more concentrated in the apical region of the stem. Second, local perception of the gravitropic signal, which is necessary for exact simulation in Coprinus , is not needed in wheat coleoptiles (the corresponding constant in the model can be set to zero). Third, the transmission rate of the gravitropic signal is about seven times faster in wheat coleoptiles than in the mushroom stem. Thus, we demonstrate that a single model, depending on the values given to its parameters, is able to simulate the spatial organization of the gravitropic reaction of wheat coleoptiles and Coprinus mushroom stems. The model promises to be a valuable predictive tool in guiding future research into the gravitropic reaction of axial organs of all types.     

Ethylene Interacts with Auxin in Regulating Developmental Attenuation of Gravitropism in Flax Root

Previous research shows that gravity-sensing in flax (Linum usitatissimum) root is initiated during seed imbibition and precedes root emergence. In this study we investigated the developmental attenuation of flax root gravitropism post-germination and the involvement of ethylene. Gravity response deteriorated significantly from 3 to 11?h after root emergence, which occurred at around 19?h after imbibition (that is, from “age” 22 to 30?h). Although the root elongation rate increased from 22 to 30?h, the gravitropic curving rate declined steadily. Older roots were able to tolerate higher levels of exogenous IAA before inhibition of elongation and gravitropism occurred. The age-dependent effect of IAA on root growth and gravitropism suggests that young roots are more sensitive to auxin and respond to a smaller vertical auxin gradient than older roots upon horizontal gravistimulation. The ethylene synthesis inhibitor AVG (2-aminoethoxyvinyl glycine, 10?μM) or ethylene action inhibitor Ag⁺ (10?μM) stimulated gravitropic curvature of 30?h roots by 24 and 32%, respectively, but had no effect on 22?h roots, suggesting that as roots age, ethylene begins to play a role in root gravitropism. The auxin transport inhibitor NPA (N-naphthylphthalamic acid, 50?μM) reduced gravitropic curvature of 30?h roots by 24% but had no effect on 22?h roots. On the other hand, treating roots simultaneously with the auxin transport inhibitor and ethylene synthesis or action inhibitor stimulated gravitropic curvature of 30?h roots but not 22?h roots. Taken together, these data indicate that as roots develop, their weakened gravity response is due to decreased auxin sensitivity and possibly auxin transport regulated by ethylene.     

Gravitropic response of inflorescence stems in Arabidopsis thaliana.

We have characterized the gravitropic response of inflorescence stems in Arabidopsis thaliana. When the inflorescence stems were placed horizontally, they curved upward about 90 degrees within 90 min in darkness at 23 degrees C, exhibiting strong negative gravitropism. Decapitated stem segments (without all flowers, flower buds, and apical apices) also showed gravitropic responses when they included the elongation zone. This result indicates that the minimum elements needed for the gravitropic response exist in the decapitated inflorescence stem segments. At least the 3-min gravistimulation time was sufficient to induce the initial curvature at 23 degrees C after a lag time of about 30 min. In the gravitropic response of inflorescence stems, (a) the gravity perception site exists through the elongating zone, (b) auxin is involved in this response, (c) the gravitropic curvature was inhibited at 4 degrees C but at least the gravity perception step could occur, and (d) two curvatures could be induced in sequence at 23 degrees C by two opposite directional horizontal gravistimulations at 4 degrees C.     

Physiology of Movements in Stems of Seedling Pisum sativum L. cv. Alaska : I. Experimental Separation of Nutation from Gravitropism

Gravitropism and nutation in the stems of dark-grown, seedling peas (Pisum sativum L. cv. Alaska) were recorded on time-lapse photographs made with photomorphogenetically inactive light. Although gravitropism and nutation have been connected by several different theories in the past, our experiments indicate that the two processes are in fact dissociable. The evidence is as follows: (a) Nutational patterns are asymmetric. There is much greater amplitude of oscillation in the plane parallel () to the plane of the apical hook than in the plane perpendicular (), yet the average gravitropic response is equal in these two planes. (b) Brief red light irradiation given 16 to 24 hours before observation greatly increases the amplitude of nutation in the -plane, but has no influence on the kinetics of gravitropic response. (c) An inhibitor of auxin transport, α-naphthylphthalamic acid, strongly inhibits nutation at 5 micromolar but affects gravitropism only at higher concentrations. (d) Nutation is also strongly inhibited by removal of the apical bud, but gravitropism is unaffected. (e) The period of nutation does not exhibit a constant relationship to the response time of gravitropism. The above evidence is inconsistent with theories that gravitropism is an asymmetrically modified nutation or, alternatively, that nutational oscillations result in a simple fashion from gravitropic overshoots. The evidence is consistent with, although not proof of, autonomous factors such as an endogenous rhythm of growth as the cause of nutation in pea stems. However, gravity and nutation do interact. Nutation in a population of seedlings can be synchronized and brought into phase by a single gravitropic induction. Furthermore, the response time and initial rate of gravitropic curvature depend to some extent on the phase of nutational curvature at which gravitropic induction is begun.