Gravity-controlled asymmetrical transport of auxin regulates a gravitropic response in the early growth stage of etiolated pea (Pisum sativum) epicotyls: studies using simulated microgravity conditions on a three-dimensional clinostat and using an agravitropic mutant, ageotropum

Increased expression of the auxin-inducible gene PsIAA4/5 was observed in the elongated side of epicotyls in early growth stages of etiolated pea (Pisum sativum L. cv. Alaska) seedlings grown in a horizontal or an inclined position under 1 g conditions. Under simulated microgravity conditions on a 3D clinostat, accumulation of PsIAA4/5 mRNA was found throughout epicotyls showing automorphosis. Polar auxin transport in the proximal side of epicotyls changed when the seedlings were grown in a horizontal or an inclined position under 1 g conditions, but that under clinorotation did not, regardless of the direction of seed setting. Accumulation of PsPIN1 and PsPIN2 mRNAs in epicotyls was affected by gravistimulation, but not by clinorotation. Under 1 g conditions, auxin-transport inhibitors made epicotyls of seedlings grown in a horizontal or inclined position grow toward the proximal direction to cotyledons. These inhibitors led to epicotyl bending toward the cotyledons in seedlings grown in an inclined position under clinorotation. Polar auxin transport, as well as growth direction, of epicotyls of the agravitropic mutant ageotropum did not respond to various gravistimulation. These results suggest that alteration of polar auxin transport in the proximal side of epicotyls regulates the graviresponse of pea epicotyls.     

Control of gravimorphogenesis by auxin: accumulation pattern of CS-IAA1 mRNA in cucumber seedlings grown in space and on the ground

Cucumber (Cucumis sativus L.) seedlings grown in microgravity developed a peg on each side of the transition zone between hypocotyl and root, whereas seedlings grown in a horizontal position on the ground developed a peg on the concave side of the gravitropically bending transition zone. The morphological features of the space-grown seedlings were similar to those of seedlings grown in a vertical position on the ground with their radicles pointing down: both became two-pegged seedlings. Morphogenesis of cucumber seedlings is thus inhibited by gravity. Analysis by in-situ hybridization of an auxin-inducible gene, CS-IAA1, showed that its mRNA accumulated to a much greater extent on the lower side of the transition zone in the horizontally placed seedlings on the ground just prior to and during the initiation period of peg formation. On the other hand, when seedlings were grown in microgravity or in a vertical position on the ground, accumulation of CS-IAA1 mRNA occurred all around the transition zone. Accumulation of CS-IAA1 mRNA in horizontally grown seedlings appreciably decreased on the upper side of the transition zone and increased on the lower side upon gravistimulation, compared with the two-pegged seedlings. Application of IAA to seedlings in a horizontal position caused the development of a peg on each side of the transition zone, or a collar-like protuberance, depending on the concentration used. These results suggest that upon gravistimulation the auxin concentration on the upper side of the horizontally placed transition zone is reduced to a level below the threshold value necessary for peg formation. Space-grown seedlings of cucumber might develop two pegs symmetrically because the auxin level in the entire transition zone is maintained above the threshold. This spaceflight experiment verified for the first time that auxin does not redistribute in microgravity. Received: 10 February 2000 / Accepted: 15 March 2000     

Graviresponse and its regulation from the aspect of molecular levels in higher plants: growth and development, and auxin polar transport in etiolated pea seedlings under microgravity.

In STS-95 space experiments we have demonstrated that microgravity conditions resulted in automorphosis in etiolated pea (Pisum sativum L. cv. Alaska) seedlings (Ueda et al. 1999). Automorphosis-like growth and development in etiolated pea seedlings were also induced under simulated microgravity conditions on a 3-dimensional (3-D) clinostat, epicotyls being the most oriented toward the direction far from the cotyledons. Detail analysis of epicotyl bending revealed that within 36 h after watering, no significant difference in growth direction of epicotyls was observed in between seedlings grown on the 3-D clinostat and under 1 g conditions, differential growth near the cotyledonary node resulting in epicotyl bending of ca. 45 degrees toward the direction far from the cotyledons. Thereafter epicotyls continued to grow almost straightly keeping this orientation on the 3-D clinostat. On the other hand, the growth direction in etiolated seedlings changed to antigravity direction by negative gravitropic response under 1 g conditions. Automorphological epicotyl bending was also phenocopied by the application of auxin polar transport inhibitors such as 9-hydroxyfluorene-9-carboxylic acid, N-(1-naphtyl)phthalamic acid and 2,3,5-triiodobenzoic acid. These results together with the fact that auxin polar transport activity in etiolated pea epicotyls was substantially reduced in space suggested that reduced auxin polar transport is closely related to automorphosis. Strenuous efforts to learn how gravity contributes to the auxin polar transport in etiolated pea epicotyls in molecular bases resulted in successful identification of PsPIN2 and PsAUX1 encoding putative auxin-efflux and influx carrier proteins, respectively. Based on the results of these gene expression under simulated microgravity conditions, a possible role of PsPIN2 and PsAUX1 genes for auxin polar transport in etiolated pea seedlings will be discussed.     

Automorphosis of etiolated pea seedlings in space is simulated by a three-dimensional clinostat and the application of inhibitors of auxin polar transport

Etiolated pea (Pisum sativum L. cv. Alaska) seedlings grown under microgravity conditions in space show automorphosis: bending of epicotyls, inhibition of hook formation and changes in root growth direction. In order to determine the mechanisms of microgravity conditions that induce automorphosis, we used a three-dimensional clinostat and obtained the successful induction of automorphosis-like growth of etiolated pea seedlings. Kinetic studies revealed that epicotyls bent at their basal region towards the clockwise direction far from the cotyledons from the vertical line (0 degrees) at approximately 40 degrees in seedlings grown both at 1 g and in the clinostat within 48 h after watering. Thereafter, epicotyls retained this orientation during growth in the clinostat, whereas those at 1 g changed their growth direction against the gravity vector and exhibited a negative gravitropic response. On the other hand, the plumular hook that had already formed in the embryo axis tended to open continuously by growth at the inner basal portion of the elbow; thus, the plumular hook angle initially increased; this was followed by equal growth on the convex and concave sides at 1 g, resulting in normal hook formation; in contrast, hook formation was inhibited on the clinostat. The automorphosis-like growth and development of etiolated pea seedlings was induced by auxin polar transport inhibitors (9-hydroxyfluorene-9-carboxylic acid, N-(1-naphthyl)phthalamic acid and 2,3,5-triiodobenzoic acid), but not by anti-auxin (p-chlorophenoxyisobutyric acid) at 1 g. An ethylene biosynthesis inhibitor, 1-aminooxyacetic acid, inhibited hook formation at 1 g, and ethylene production of etiolated seedlings was suppressed on the clinostat. Clinorotation on the clinostat strongly reduced the activity of auxin polar transport of epicotyls in etiolated pea seedlings, similar to that observed in space experiments (Ueda J, Miyamoto K, Yuda T, Hoshino T, Fujii S, Mukai C, Kamigaichi S, Aizawa S, Yoshizaki I, Shimazu T, Fukui K (1999) Growth and development, and auxin polar transport in higher plants under microgravity conditions in space: BRIC-AUX on STS-95 space experiment. J Plant Res 112: 487492). These results suggest that clinorotation on a three-dimensional clinostat is a valuable tool for simulating microgravity conditions, and that automorphosis of etiolated pea seedlings is induced by the inhibition of auxin polar transport and ethylene biosynthesis.     

Graviresponse in higher plants and its regulation in molecular bases: relevance to growth and development, and auxin polar transport in etiolated pea seedlings]

We review the graviresponse under true and simulated microgravity conditions on a clinostat in higher plants, and its regulation in molecular bases, especially on the aspect of auxin polar transport in etiolated pea (Pisum sativum L. cv. Alaska) seedlings which were the plant materials subjected to STS-95 space experiments. True and simulated microgravity conditions substantially affected growth and development in etiolated pea seedlings, especially the direction of growth of stems and roots, resulting in automorphosis. In etiolated pea seedlings grown in space, epicotyls were the most oriented toward the direction far from the cotyledons, and roots grew toward the aerial space of Plant Growth Chamber. Automorphosis observed in space were well simulated by a clinorotation on a 3-dimensional clinostat and also phenocopied by the application of auxin polar transport inhibitors of 2,3,5-triiodobenzoic acid, N-(1-naphtyl)phthalamic acid and 9-hydroxyfluorene-9-carboxylic acid. Judging from the results described above together with the fact that activities of auxin polar transport in epicotyls of etiolated pea seedlings grown in space substantially were reduced, auxin polar transport seems to be closely related to automorphosis. Strenuous efforts to learn in molecular levels how gravity contributes to the auxin polar transport in etiolated pea epicotyls resulted in successful identification of PsPIN2 and PsAUX1 genes located in plasma membrane which products are considered to be putative efflux and influx carriers of auxin, respectively. Based on the results of expression of PsPIN2 and PsAUX1 genes under various gravistimulations, a possible role of PsPIN2 and PsAUX1 genes for auxin polar transport in etiolated pea seedlings will be discussed.     

The auxin-resistant diageotropica mutant of tomato responds to gravity via an auxin-mediated pathway

 Hypocotyls of the diageotropica (dgt) mutant of tomato (Lycopersicon esculentum Mill.) do not elongate in response to exogenous auxin, but can respond to gravity. This appears paradoxical in light of the Cholodny-Went hypothesis, which states that shoot gravicurvature results from asymmetric stimulation of elongation by auxin. While light-grown dgt seedlings can achieve correct gravitropic reorientation, the response is slow compared to wild-type seedlings. The sensitivity of dgt seedlings to inhibition of gravicurvature by immersion in auxin or auxin-transport inhibitors is similar to that of wild-type plants, indicating that both an auxin gradient and auxin transport are required for the gravitropic response and that auxin uptake, efflux, and at least one auxin receptor are functional in dgt. Furthermore, dgt gravicurvature is the result of asymmetrically increased elongation as would be expected for an auxin-mediated response. Our results suggest differences between elongation in response to exogenous auxin (absent in dgt) and elongation in response to gravistimulation (present but attenuated in dgt) and confirm the presence of two phases during the gravitropic response, both of which are dependent on functional auxin transport. Received: 16 July 1999 / Accepted: 24 September 1999     

Differential accumulation of the mRNA of the auxin-repressed gene CsGRP1 and the auxin-induced peg formation during gravimorphogenesis of cucumber seedlings

When cucumber seeds are germinated horizontally, an outgrowth (peg) develops on the lower side of the transition zone between the hypocotyl and the root for pulling the cotyledons and plumule out of the seed coat. We previously suggested that gravistimulation suppresses peg formation on the upper side of the transition zone when placed in a horizontal position. In the gravistimulated transition zone, auxin and the mRNAs of auxin-inducible genes are more abundant in the lower side than in the upper side. Here, using fluorescent differential display, we identified Cucumis sativus glycine-rich protein1(CsGRP1) as a gene whose mRNA accumulated more abundantly on the upper side than on the lower side of the transition zone in response to gravistimulation. Auxin starvation increased CsGRP1 mRNA in segments of the transition zone, and inhibition of polar auxin transport with 2,3,5-triiodobenzoic acid (TIBA) prevented the asymmetric accumulation of CsGRP1 mRNA. These results suggest that gravistimulation increases not only the expression of auxin-inducible genes on the lower side of the transition zone, but also the expression of auxin-repressed genes, such as CsGRP1, on the upper side of cucumber seedlings. In the hypocotyls of 3-day-old seedlings, neither gravistimulation nor changes in auxin level influenced the accumulation of CsGRP1 mRNA. These results suggest that the transition zone responds to gravistimulation in a specific manner by an asymmetric expression of CsGRP1 gene during regulation of peg formation.     

STS-95 space experiment for plant growth and development, and auxin polar transport.

The principal objective of the space experiment, BRIC-AUX on STS-95, was the integrated analysis of the growth and development of etiolated pea and maize seedlings in space, and the effect of microgravity conditions in space on auxin polar transport in the segments. Microgravity conditions in space strongly affected the growth and development of etiolated pea and maize seedlings. Etiolated pea and maize seedlings were leaned and curved during space flight, respectively. Finally the growth inhibition of these seedlings was also observed. Roots of some pea seedlings grew toward the aerial space of Plant Growth Chamber. Extensibilities of cell walls of the third internode of etiolated pea epicotyls and the top region of etiolated maize coleoptiles which were germinated and grown under microgravity conditions in space were significantly low. Activities of auxin polar transport in the second internode segments of etiolated pea seedlings and coleoptile segments of etiolated maize seedlings were significantly inhibited and extremely promoted, respectively, under microgravity conditions in space. These results strongly suggest that auxin polar transport as well as the growth and development of plants is controlled under gravity on the earth.     

Inhibition of polar calcium movement and gravitropism in roots treated with auxin-transport inhibitors

Primary roots of maize (Zea mays L.) and pea (Pisum sativum L.) exhibit strong positive gravitropism. In both species, gravistimulation induces polar movement of calcium across the root tip from the upper side to the lower side. Roots of onion (Allium cepa L.) are not responsive to gravity and gravistimulation induces little or no polar movement of calcium across the root tip. Treatment of maize or pea roots with inhibitors of auxin transport (morphactin, naphthylphthalamic acid, 2,3,5-triiodobenzoic acid) prevents both gravitropism and gravity-induced polar movement of calcium across the root tip. The results indicate that calcium movement and auxin movement are closely linked in roots and that gravity-induced redistribution of calcium across the root cap may play an important role in the development of gravitropic curvature.Abbreviations 9-HFCA 9-hydroxyfluorenecarboxylic acid - NPA naphthylphthalamic acid - TIBA 2,3,5-triiodobenzoic acid - IAA indole-3-acetic acid     

Growth and development, and auxin polar transport in higher plants under microgravity conditions in space: BRIC-AUX on STS-95 space experiment.

The principal objectives of the space experiment, BRIC-AUX on STS 95, were the integrated analysis of the growth and development of etiolated pea and maize seedlings in space and a study of the effects of microgravity conditions in space on auxin polar transport in these segments. Microgravity significantly affected the growth and development of etiolated pea and maize seedlings. Epicotyls of etiolated pea seedlings were the most oriented toward about 40 to 60 degrees from the vertical. Mesocotyls of etiolated maize seedlings were curved at random during space flight but coleoptiles were almost straight. Finally the growth inhibition of these seedlings in space was also observed. Roots of some pea seedlings grew toward to the aerial space of Plant Growth Chamber. Extensibilities of cell walls of the third internode of etiolated pea epicotyls and the top region of etiolated maize coleoptiles, which were germinated and grown under microgravity conditions in space, were significantly low as compared with those grown on the ground of the earth. Activities of auxin polar transport in the second internode segments of etiolated pea seedlings and coleoptile segments of etiolated maize seedlings were significantly inhibited and promoted, respectively, under microgravity conditions in space. These results strongly suggest that auxin polar transport as well as the growth and development of plants is controlled under gravity on the earth.     

Gravistimulation changes the accumulation pattern of the CsPIN1 auxin efflux facilitator in the endodermis of the transition zone in cucumber seedlings

Cucumber (Cucumis sativus) seedlings grown in a horizontal position develop a specialized protuberance (or peg) on the lower side of the transition zone between the hypocotyl and the root. This occurs by suppressing peg formation on the upper side via a decrease in auxin resulting from a gravitational response. However, the gravity-stimulated mechanism of inducing asymmetric auxin distribution in the transition zone is poorly understood. The gravity-sensing tissue responsible for regulating auxin distribution in the transition zone is thought to be the endodermal cell. To characterize the gravity-stimulated mechanism, the auxin efflux facilitator PIN-FORMED1 (CsPIN1) in the endodermis was identified and the localization of CsPIN1 proteins during the gravimorphogenesis of cucumber seedlings was examined. Immunohistochemical analysis revealed that the accumulation pattern of CsPIN1 protein in the endodermal cells of the transition zone of cucumber seedlings grown horizontally differed from that of plants grown vertically. Gravistimulation for 30 min prompted changes in the accumulation pattern of CsPIN1 protein in the endodermis as well as the asymmetric distribution of auxin in the transition zone. Furthermore, 2,3,5-triiodobenzoic acid inhibited the differential distribution of auxin as well as changes in the accumulation pattern of CsPIN1 in the endodermis of the transition zone during gravistimulation. These results suggest that the altered pattern of CsPIN1 accumulation in the endodermis in response to gravistimulation influences lateral auxin transport through the endodermis, resulting in asymmetric auxin distribution in the transition zone.     

A gravitropic stimulation-induced growth inhibitor, β-(isoxazolin-5-on-2yl)-alanine,is a possible mediator of negative gravitropic bending of epicotyls in etiolated <Emphasis Type="Italic">Pisum sativum</Emphasis> seedlings

Negative gravitropic bending and its possible mediator in etiolated Alaska pea seedlings were intensively studied in comparison with seedlings of an agravitropic mutant, ageotropum. When 3.5-day-old etiolated Alaska seedlings were horizontally placed, the growth suppression at the upper side of the epicotyls began 10 min after the onset of the gravitropic stimulation, whereas the growth acceleration at the lower side began at 30 min, resulting in negative gravitropic bending. In contrast, no gravitropic bending was observed in the etiolated ageotropum seedlings, for which the epicotyls show an automorphogenesis-like growth. Strenuous efforts to identify a possible mediator that induces the gravitropic bending resulted in successfully identifying β-(isoxazolin-5-on-2yl)-alanine (βIA). The unilateral application of βIA to the etiolated Alaska epicotyls substantially induced epicotyl bending toward the application site, indicating that βIA could act as a growth inhibitor. Analyses of the distribution of βIA in the upper and lower flanks of the etiolated Alaska epicotyls revealed that its content rapidly increased twice in the upper flanks compared with that in the lower ones in response to gravitropic stimulation, whereas its content in the lower flanks was almost equal to that in the vertical control. In etiolated ageotropum epicotyls, an almost equal amount of βIA was distributed in the upper and lower flanks of epicotyls. These results suggest that a gravitropic stimulation increases βIA in the upper flank, resulting in the negative gravitropic bending of epicotyls via the suppression of the growth rate at the upper side of epicotyls in the etiolated Alaska pea seedlings.     

Mechano-sensitive orientation of cortical microtubules during gravitropism in azuki bean epicotyls

The orientation of cortical microtubules (cMT) during gravitropism was studied in epidermal cells of azuki epicotyls. The relative proportion of cells with longitudinal cMT increased in the upper epidermis, and those with transverse cMT increased in the lower epidermis. When epicotyls were kept straight during gravistimulation, no change in cMT orientation occurred in either the upper and lower epidermis. When epicotyls were forced to bend downward, cells with transverse cMT increased in the upper epidermis, and those with longitudinal cMT increased in the lower epidermis. When epicotyls were loaded with naphthylphthalamic acid, an inhibitor of auxin transport, both gravitropic bending and change in cMT orientation were inhibited. However, when a change in cMT orientation was induced by forced downward bending, cells with longitudinal cMT increased in the compressed (lower) side and those with transverse cMT increased in the extended (upper) side. It was suggested that cMT orientation was controlled by the bending of the epicotyl and not by a gravity signal per se. Loading with Gd³⁺, an inhibitor of the stretch-activated channel, did not inhibit gravitropic bending. However, it inhibited cMT reorientation induced by gravitropic bending and by forced bending. Involvement of the stretch-activated channel in mechano-sensitive orientation of cMT was suggested.     

Characterization of OsPID, the rice ortholog of PINOID, and its possible involvement in the control of polar auxin transport

PINOID, a serine threonine protein kinase in Arabidopsis, controls auxin distribution through a positive control of subcellular localization of PIN auxin efflux carriers. Compared with the rapid progress in understanding mechanisms of auxin action in dicot species, little is known about auxin action in monocot species. Here, we describe the identification and characterization of OsPID, the PINOID ortholog of rice. Phylogenetic analysis showed that the rice genome contains a single PID ortholog, OsPID. Constitutive overexpression of OsPID caused a variety of abnormalities, such as delay of adventitious root development, curled growth of shoots and agravitropism. Abnormalities observed in the plants that overexpress OsPID could be phenocopied by treatment with an inhibitor of active polar transport of auxin, indicating that OsPID could be involved in the control of polar auxin transport in rice. Analysis of OsPID mRNA distribution showed a complex pattern in shoot meristems, indicating that it probably plays a role in the pattern formation and organogenesis in the rice shoot.     

Expression of PIN and AUX1 genes encoding putative carrier proteins for auxin polar transport in etiolated pea epicotyls [correction of epicotyles] under simulated microgravity conditions on a three-dimensional clinostat.

Etiolated pea (Pisum sativum L. cv. Alaska) seedlings grown under simulated microgravity conditions on a 3-dimensional clinostat showed automorphosis-like growth and development similar to that observed in true microgravity conditions in space. Application of inhibitors of auxin polar transport phenocopied automorphosis-like growth on 1 g conditions, suggesting that automorophosis is closely related to auxin polar transport. Strenuous efforts to know the relationships between automorphosis and auxin polar transport in pea seedlings at molecular bases resulted in successful identification of PsPIN2 and PsAUX1 encoding putative auxin efflux and influx carrier protein, respectively. Significantly high levels in homology were found on nucleotide and deduced amino acid sequences among PsPIN2, PsPIN1 and AtPINs, and between PsAUX1 and AtAUX1. Expression of PsPIN1 and PsAUX1 genes in etiolated pea seedlings grown on the clinostat were substantially affected, but that of PsPIN2 was not. Roles of these genes in auxin polar transport and automorphosis of etiolated pea seedlings are also described.     

Gravity-induced modification of auxin transport and distribution for peg formation in cucumber seedlings: possible roles for CS-AUX1 and CS-PIN1

Cucurbit seedlings potentially develop a peg on each side of the transition zone between the hypocotyl and root. Seedlings grown in a horizontal position suppress the development of the peg on the upper side of the transition zone in response to gravity. It is suggested that this suppression occurs due to a reduction in auxin levels to below the threshold value. We show in this study that the free indole-3-acetic acid (IAA) content is low, while IAA conjugates are significantly more abundant in the upper side of the transition zone of gravistimulated seedlings, compared to the lower side. A transient increase in mRNA of the auxin-inducible gene, CS-IAA1, was observed in the excised transition zone. The result suggests that the transition zone is a source of auxin. Cucumber seedlings treated with auxin-transport inhibitors exhibited agravitropic growth and developed a peg on each side of the transition zone. Auxin-transport inhibitors additionally caused an increase in CS-IAA1 mRNA accumulation at the transition zone, indicating a rise in intracellular auxin concentrations due to a block of auxin efflux. To study the involvement of the auxin transport system in peg formation, we isolated the cDNAs of a putative auxin influx carrier, CS-AUX1, and putative efflux carrier, CS-PIN1, from cucumber (Cucumis sativus L.) plants. Both genes (CS-AUX1 in particular) were auxin-inducible. Accumulation of CS-AUX1 and CS-PIN1 mRNAs was observed in vascular tissue, cortex and epidermis of the transition zone. A reduced level of CS-AUX1 mRNA was observed in the upper side of the gravistimulated transition zone, compared with the lower side. It is therefore possible that a balance in the activities of auxin influx and efflux carriers controls intracellular auxin concentration at the transition zone, which results in lateral placement of a peg in cucumber seedlings.Abbreviations HFCA 9-hydroxyfluorene-9-carboxylic acid - IAA indole-3-acetic acid - NPA 1-N-naphthylphthalamic acid - TIBA 2,3,5-triiodobenzoic acid     

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.     

Auxin-mediated response of cucumber seedlings to gravity]

Gravity regulates peg formation because cucumber seedlings grown in a horizontal position develop a peg on the lower side of the transition zone (TR zone) but not on the upper side. Studies on peg formation have suggested the regulation of peg formation by gravity as follows. Cucumber seedlings potentially develop a peg on both the lower and upper sides of the TR zone. The development of the peg on upper side of the TR zone is suppressed in response to gravity. A phytohormone, auxin, induces peg formation. Upon gravistimulation the auxin concentration on the upper side of the TR zone is reduced to a level below the threshold value necessary for peg formation. The unequally distributed auxin across TR zone is caused by a change in accumulation of auxin influx carrier (CsAUX1) protein and auxin efflux carrier (CsPIN1) protein in response to gravity. In addition, TR zone before peg initiation expresses both CsARF2 (putative activator of auxin response factor) and CsIAA1 (putative repressor of auxin-inducible gene expression), by which TR zone could respond the auxin gradient regulated by gravity.     

Negative gravitropic response of roots directs auxin flow to control root gravitropism

Root tip is capable of sensing and adjusting its growth direction in response to gravity, a phenomenon known as root gravitropism. Previously, we have shown that negative gravitropic response of roots (NGR) is essential for the positive gravitropic response of roots. Here, we show that NGR, a plasma membrane protein specifically expressed in root columella and lateral root cap cells, controls the positive root gravitropic response by regulating auxin efflux carrier localization in columella cells and the direction of lateral auxin flow in response to gravity. Pharmacological and genetic studies show that the negative root gravitropic response of the ngr mutants depends on polar auxin transport in the root elongation zone. Cell biology studies further demonstrate that polar localization of the auxin efflux carrier PIN3 in root columella cells and asymmetric lateral auxin flow in the root tip in response to gravistimulation is reversed in the atngr1;2;3 triple mutant. Furthermore, simultaneous mutations of three PIN genes expressed in root columella cells impaired the negative root gravitropic response of the atngr1;2;3 triple mutant. Our work revealed a critical role of NGR in root gravitropic response and provided an insight of the early events and molecular basis of the positive root gravitropism.