髙橋 秀幸

To study the effect of the space environment on plant growth including the reproductive growth and genetic aberration for a long-term plant life cycle, we have initiated development of a new type of facility for growing plants under microgravity conditions. The facility is constructed with subsystems for controlling environmental elements. In this paper, the concept of the facility design is outlined. Subsystems controlling air temperature, humidity, CO2 concentration, light and air circulation around plants and delivering recycled water and nutrients to roots are the major concerns, Plant experiments for developing the facility and future plant experiments with the completed facility are also overviewed. We intend to install this facility in the Japan Experiment Facility (JEM) bearded on the International Space Station. (C) 2000 COSPAR. Published by Elsevier Science Ltd.

We examined the effect of microgravity on the peg formation of cucumber seedlings for clarifying the mechanism of gravimorphogenesis in cucurbitaceous plants. The spaceflight experiments verified that gravity controls the formation of peg, hypocotyl hook and growth orientation of cucumber seedlings. Space-grown cucumber developed a peg on each side of the transition zone of the hypocotyl and root, indicating that on the ground peg formation is regulated negatively by gravity (Takahashi et al. 2000). It was found that the auxin-regulated gene, CS-IAA1, was strongly expressed in the transition zone where peg develops (Fujii et al. 2000). In the seedlings grown horizontally on the ground, CS-IAA1 transcripts were much abundant on the lower side of the transition zone, but no such differential expression of CS-IAA1 was observed in the space-grown cucumber (Kamada et al. 2000). These results imply that gravity plays a role in peg formation through auxin redistribution. By the negative control, peg formation on the upper side of the transition zone in the horizontally growing seedlings might be suppressed due to a reduction in auxin concentration. The threshold theory of auxin concentration accounted for the new concept, negative control of morphogenesis by gravity (Kamada et al. 2000). Anatomical studies have shown that there exists the target cells destined to be a peg and distinguishable at the early stage of the growth. Ultra-structural analysis suggested that endoplasmic reticulum develops well in the cells of the future peg. Furthermore, it was found that re-organization of cortical microtubules is required for the change in cell growth polarity in the process of peg formation. The spaceflight experiment with cucumber seedlings also suggested that in microgravity positive hydrotropic response of roots occurred without interference by gravitropic response (Takahashi et al. 1999b). Thus, this spaceflight experiment together with the ground-based studies has shown that cucumber seedling is an ideal for the study of gravimorphogenesis, hydrotropism and their interaction. Although peg formation is seen specifically in cucurbitaceous seedlings, it involves graviperception, auxin transport and redistribution and cytoskeletal modification for controlling cell growth polarity. This system could be a useful model for studying important current issues in plant biology.

Seedlings of Cucurbitaceae plants form a protuberance, termed peg, on the transition zone between hypocotyl and root. Our spaceflight experiment verified that the lateral positioning of a peg in cucumber seedlings is modified by gravity. a has been suggested that auxin plays an important role in the gravity-controlled positioning of a peg on the ground. Furthermore, cucumber seedlings grown in microgravity developed a number of the lateral roots that grew towards the water-containing substrate in the culture vessel, whereas on the ground they oriented perpendicular to the primary root growing down. The response of the lateral roots in microgravity was successfully mimicked by clinorotation of cucumber seedlings on the three dimensional clinostat However, this bending response of the lateral roots was observed only in an aeroponic culture of the seedlings but not in solid medium. We considered the response of the lateral roots in microgravity and on clinostat as positive hydrotropism that could easily be interfered by gravitropism on the ground. This system with cucumber seedlings is thus a useful model of spaceflight experiment for the study of the gravimorphogenesis, root hydrotropism and their interaction.

Compared to wild type, the lazy mutant in Oryza sativa L. shows a reduced gravitropic response. In order to locate the lesion in the stimulus-response chain, coleoptile segments of lazy rice were investigated with respect to auxin transport. Gravity-induced lateral movement of radiolabelled indoleacetic acid (IAA) was strongly inhibited by the lazy mutation compared to wild type while uptake and longitudinal transport of IAA, as well as amyloplast sedimentation, were not significantly affected. These findings suggest that LAZY controls a step in the signalling chain between statoliths and auxin secretion.

We isolated an endoxyloglucan transferase cDNA (Ps-EXGT1) from the roots of an agravitropic pea mutant, ageotropum. The putative product of the cDNA was 34.1 kDa and consisted of 293 amino acid residues. The predicted amino acid sequence was 75.1-88.6% identical to those of EXGT genes in other plants. The Ps-EXGT1 cDNA was strongly expressed in elongating roots and stems but not in either mature stems or young leaves. In roots, the transcription level of Ps-EXGT1 was most abundant in the rapidly growing region. When root elongation was inhibited by a water stress, Ps-EXGT1 transcription was repressed. The roots curved hydrotropically due to differential growth of the cortical cells in the elongation zone when the root cap was exposed to a gradient of water potential; the length of the cells on the side of lower water potential was much longer than those on the side of higher water potential. The expression pattern of Ps-EXGT1 in the hydrotropically responding roots fluctuated between the side of the higher water potential and that of the lower water potential in the elongation zone. In other words, the accumulation of Ps-EXGT1 mRNA was much greater on the side of lower water potential than on that of higher potential just prior to the commencement of positive hydrotropism. When the roots started to curve slightly away from the side of higher water potential causing a rhythmic oscillatory movement [Takano et al, (1995) Planta 197: 410], there was more transcription of Ps-EXGT1 on the side of higher water potential. These results suggest that the transcription of Ps-EXGT1 is involved in cell growth and that this regulation of transcription plays a role in the differential growth of hydrotropically responding roots.

Roots have been shown to respond to a moisture gradient by positive hydrotropism, Agravitropic mutant plants are useful for the study of the hydrotropism in roots because on Earth hydrotropism is obviously altered by the gravity response in the roots of normally gravitropic plants. The roots are able to sense water potential gradient as small as 0.5 MPa mm(-1). The soot cap includes the sensing apparatus that causes a differential growth at the elongation region of roots, A gradient in apoplastic calcium and calcium influx through plasmamembrane in the root cap is somehow involved in the signal transduction mechanism in hydrotropism, which may cause a differential change in cell wall extensibility at the elongation region. We have isolated an endoxy loglucan transferase (EXGT) gene that is strongly expressed in pea roots and appears to be involved in the differential growth in hydrotropically responding roots. Thus, it is now possible to study hydrotropism in roots by comparing with or separate from gravitropism, These results also imply that microgravity conditions in space are useful for the study of hydrotropism and its interaction with gravitropism, (C)1999 COSPAR. Published by Elsevier Science Ltd.

Seedlings of cucurbitaceous plants develop a protuberant tissue, or peg, on the lower side of the transition region between root and hypocotyl when germinated in a horizontal position. Peg develops due to a change in growth polarity of the cortical cells. We have examined the role of the cytoskeketal structure in peg formation of cucumber seedlings. We observed that in both peg and normal cortical cells of 36 h-old seedlings the microtubules (MTs) were arranged perpendicular to the longitudinal axis of the elongating cells. Application of colchicine perturbed the MTs structure and inhibited the formation of pegs. In 20 h-old seedlings, MTs in cortical cells destined to be a peg tissue had no preferential organization, whereas MTs in normal cortical cells were transversely oriented. After 24 h, the MTs in future peg cells were arranged similar to those of 36 h-old seedlings, although the initiation of peg tissue was not yet visible. These results suggest that reorganization of MTs is required for peg formation and causes the change in growth polarity of the cortical cells. (C) 1999 COSPAR. Published by Elsevier Science Ltd.

This paper presents the results of study on a prototype design and its set up to measure plant water uptake under the influence of microgravity (μG). The results are summarized as follows: 1) A proposed prototype system smoothly functioned and was controlled throughout the flight. 2) Leaf temperature increased by 0.8-1.6°C during μG condition. 3) Water uptake rate rapidly dropped when the gravity condition changed into μG from. 4) The water uptake recovered as the gravity increased to 1.5G which created by the ascending flight following to the μG flight. Uptake rate change took place at the time of gravity change (high-G to /from μG) but it did not sustain and its relaxation time varied.

Development of water recovery and recycling system in a plant growth chamber is most important challenge for plant cultivation under micro-gravity. In this paper, a water recycling system consisting of Peltier cooler unit and capillary fibrous mat for water transport is proposed as a suitable technology for small-scale closed plant growth chamber under micro-gravity. In the preliminary experiment, maximum water condensation rate by the fin of Peltier cooler unit was 100g/day. This value is not high enough to support water circulation in plant growth chamber when mature plants are densely grown. Water could be successfully transferred from the fin to the plant growth medium through the fibrous mat in the short-term experiment (144h). A long-term experiment is required to evaluate the system performance and stability. Key factors controlling water movement of the system, including hydraulic conductivity of the fibrous mat and medium and water condensation capacity of cooling fin, are discussed.

We have compared shoot responses of agravitropic rice and barley plants to vertical inversion with those of normal ones. When rice plants were vertically inverted, the main stems of a japonica type of rice, cv. Kamenoo, showed negative gravitropism at nodes 2-15 of both elongated and non-elongated internodes. However, shoots of lazy line of rice, lazy-Kamenoo, bent gravitropically at nodes 11-15 only elongated internodes but not at nodes 2-10 of non-elongated ones, Thus, shoots of Kamenoo responded gravitropically at all stages of growth, whereas shoots of lazy-Kamenoo did not show gravitropic response before heading. In Kamenoo plants, lengths of both leaf-sheath and leaf-blade were shortened by vertical inversion, but those of the vertically inverted plants of lazy-Kamenoo were significantly longer than the plants in an upright position, When agravitropic and normal plants of barley were vertically inverted, the same results as in rice were obtained; elongation of both leaf-sheath and leaf-blade was inhibited in normal barley plants, Chikurin-Ibaragi No. 1, but significantly stimulated in agravitropic plants of serpentina barley. These results suggest that vertical inversion of rice and barley plants enhances the elongation growth of leaves in the absence of tropistic response.

It has been shown that deep-seeding and ethylene stimulate the elongation of the first internodes an wheat seedlings. In the present study, patterns of inheritance in the elongation of wheat first internodes and coleoptiles responding to deep-seeding and ethylene were examined using three crosses. Length of the coleoptile in Fz segregants showed a simple unimodal distribution resembling a normal distribution both in deep-seeded and ethylene-treated plants. Except in one cross, however, length of the first internode in Fa segregants showed a unimodal distributions with large transgressive segregation on the shorter length side in deep-seeded plants and the longer side exceeding the length of parents' first internodes in ethylene-treated plants. These results suggest that ability of plants to respond to ethylene, a regulatory factor for the elongation of the first internode in wheat, can be changed through genetic recombination.

A recA-like gene was identified in the Caenorhabditis elegans genome project database. The putative product of the gene, termed Ce-rdh-1 (C. elegans RAD51 and DMC1/LIM15 homolog 1), consists of 357 amino acid residues. The predicted amino acid sequence of Ce-rdh-1 showed 46-60% identity to both RAD51 type and DMC1/LIM15 type genes in several eukaryote species. The results of RNAi (RNA-mediated interference) indicated that repression of Ce-rdh-1 blocked chromosome condensation of six bivalents and dissociation of chiasmata in oocytes of F1 progeny. Oogenesis did not proceed to the diakinesis stage. Accordingly, all the eggs produced (F2) died in early stages. These results suggest that Ce-rdh-1 participates in meiotic recombination.

Using two varieties of wheat with genetically differing leaf lengths, several parameters concerning leaf growth were examined under controlled environmental conditions. Heritabilities for the parameters of leaf growth were calculated based on measurements of F-2 generation of the hybrid between these two varieties. The rate of leaf emergence (i.e., the speed of leaf emergence per unit time or per unit length) was almost the same between the two parent varieties, although the average length of leaf and average number of days required for the emergence of one leaf were different due to the difference in leaf length. Heritabilities for the average length of leaf and for the days required for the emergence of one leaf were 77.7 and 85.2%, respectively. In F,generation, however, no correlation was detected between the two characteristics (r=0.152). The heritabilities for the speed of leaf emergence per unit time and unit length were 55.5 and 45.2%, respectively, suggesting that the leaf growth rate (speed of leaf growth per unit time and per unit length) can be changed by selection.

Deep-seeding and ethylene were found to stimulate extension growth of the first internode of intact wheat (Triticum aestivum L.) seedlings in darkness. Seedlings of Hon Mang Mai emerged from much deeper in the soil than the seedlings of the other varieties used and their first internodes elongated to a much greater extent in response to ethylene. Carbon dioxide slowed elongation of the first internode and inhibited ethylene action. Elongation of the first internode due to deep-seeding and ethylene treatment showed high heritabilities, suggesting a genetic basis underlying those traits.

The response of roots to a moisture gradient has been reexamined, and positive hydrotropism has been demonstrated in recent years. Agravitropic roots of a pea mutant have contributed to the studies on hydrotropism. The kinetics of hydrotropic curvature, interactions between hydrotropism and gravitropism, moisture gradients required for the induction of hydrotropism, the sensing site for moisture gradients, characteristics of hydrotropic signal and differential growth, and calcium involvement in signal transduction have been subjects of these studies. This review summarizes the current state of our knowledge on hydrotropism in roots.

Positive hydrotropic curvature in the roots of the agravitropic pea (Pisum sativum L.) mutant, ageotropum, occurred when the root cap was exposed to a gradient of water potential by an asymmetric application of agar containing sorbitol [Takano et al. (1995) Planta 197: 410], As previously reported [Takahashi and Suge (1991) Physiol, Plant, 82: 24], in this study the hydrotropic response due to unilateral application of sorbitol to the root cap was totally inhibited by pretreatment with ethyleneglycol-bis-(beta-aminoethylether) N,N,N',N'-tetraacetic acid (EGTA). However, hydrotropic response of the EGTA-treated roots was recovered only when EGTA was replaced by a 10 mM calcium (CaCl2) solution prior to hydrostimulation. A calcium channel blocker, lanthanum (LaCl3), also inhibited hydrotropic curvature of ageotropum roots, whereas the hydrotropic response was affected by neither nifedipine nor verapamil, Application of calcium ionophore, A23187, resulted in a significant promotion of hydrotropic curvature. Furthermore, ageotropum roots curved away from a calcium source when an agar block containing 10 mM calcium was asymmetrically applied to the root cap, This calcium-induced curvature was found to be accelerated by water stress and significantly inhibited by LaCl3. While the calcium-induced curvature commenced within Ih after application, hydrotropic curvature became visible 3 to 4 h after an exposure to a gradient of water potential. These results indicate that apoplastic calcium and its influx through the plasmamembrane are involved in the induction of hydrotropism in roots. A gradient of water potential in the root cap may cause a physiological change that is mediated by calcium, which ultimately leads to the curvature in the elongation region associated with the hydrotropic response.

The hydrotropic bending of roots of an ageotropic pea mutant, ageotropum, was studied in humid air in a chamber with a steady humidity gradient. We examined the effects of atmospheric humidity around the root on the water status of root tissues, as well as the wall growth and the hydraulic properties of the elongating tissues. Atmospheric humidity at the surface of the root was clearly lower on the side orientated towards the air with lower humidity than on the side orientated towards the air with higher humidity. However, there were no differences in water potential and osmotic potential between the tissues that faced air with higher and lower humidities in the elongating and mature regions. Plastic extensibility was higher in the tissues that faced the air with lower humidity than in the tissues that faced the air with higher humidity. No differences in turgor pressure and yield threshold were observed between the tissues that faced air with higher and lower humidities. Therefore, the extensibility of the cell wall appeared to be responsible for the different growth rates of tissues in root hydrotropism. A further probable cause of the hydrotropical bending of roots is changes in the hydraulic conductance in the elongating tissues. Since the hydrotropic bending of roots occurred only when a root tip was exposed to a humidity gradient, hydrotropism might occur after perception of a difference in humidity by the root tip, with accompanying changes in cell wall extensibility and hydraulic conductance.

Formation and/or positioning of a protuberance, or peg, in cucumber (Cucumis sativus L.) seedlings is controlled by gravity. The gravisensing apparatus for this gravimorphogenesis is possibly shared with that of gravitropism. Sheath cells in the transition zone between stem and root contain sedimentable amyloplasts, but amyloplasts in cortical cells (peg cells) do not sediment uniformly. These putative statoliths appear before peg initiation becomes visible on the lower side of the transition zone. The increased level of auxin on the lower side of the transition zone, which may occur following graviperception, could be a factor responsible for peg development. This system may be a useful model in spaceflight experiments for understanding the mechanisms underlying gravity-regulated formation of the peg, and its interaction with gravitropism, auxin physiology and cell growth polarity.

Using an isogenic line of rice having lazy gene (la), we studied the correlation between the agravitropic response at the young seedling stage and the lazy habit (prostrate growth of tillers) at the more advanced stage of growth. In this study, it was found that both agravitropism and lazy habit were controlled by the single recessive la gene. That is, F-2 segregants of Kamenoo x lazy-Kamenoo, which had an agravitropic response at their young seedling stage, showed a lazy habit of growth in the more advanced stage of vegetative growth. On the other hand, seedlings that showed normal gravitropic curvature at their early stage of growth had an upright growth in the mature stage.

Roots of the agravitropic pea (Pisum sativum L.) mutant ageotropum show positive hydrotropism, whereas roots of Alaska peas are hydrotropically almost non-responsive. When the gravitropic response was nullified by rotation on clinostats, however, roots of Alaska peas showed unequivocal positive hydrotropism in response to a water potential gradient. These results suggest that roots of Alaska peas possess normal ability to respond hydrotropically and their weak hydrotropic response results from a counteracting effect of gravitropism.

In this study, ageotropum pea mutant was used to determine the threshold time for perception of an osmotic stimulation in the root cap and the time requirement for transduction and transmission of the hydrotropic signal from the root cap to the elongation region. The threshold time for the perception of an osmotic stimulation was compared to current estimates of threshold times for graviperception in roots. The time required for transduction and transmission in the hydrotropic response of ageotropum was compared to the time requirement in the gravity response of Alaska pea roots, We determined that threshold time for perception of an osmotic stimulation in the root cap is very rapid, occurring in less than 2 min following the application of sorbitol to the root cap. Furthermore, a single 5 min exposure of sorbitol to the root cap fully induced a hydrotropic response. We also found that transduction and transmission of an osmotic stimulus requires 90-120 min for movement from the root cap to more basal tissues involved in differential growth leading to root curvature. The very rapid threshold time for perception of root hydrotropism is similar to those times reported for root gravitropism. However, the time required for the transduction and transmission of an osmotic stimulation from the root cap is significantly longer than the time required in gravitropism. These results suggest that there must exist some differences between root hydrotropism and gravitropism in either the rate or mechanisms of transduction and transmission of the tropistic signal from the root cap.

Roots of the agravitropic pea (Pisum sativum L.) mutant, ageotropum, responded to a gradient in water potential as small as 0.5 MPa by growing toward the higher water potential. This positive response occurred when a sorbitol-containing agar block was unilaterally applied to the root cap but not when applied to the elongation region. Unilateral application of higher concentrations of sorbitol to the elongation region caused root curvature toward the sorbitol source, presumably because of growth reduction on the water-stressed side, The control blocks of plain agar applied to either the root cap or the elongation region did not cause significant curvature of the roots. These results demonstrate that hydrotropism in roots occurs following perception of a gradient in water potential by the root cap.

It has been proposed that hydrotropism interacts with gravitropism in seedling roots; that is, roots which are highly gravitropic show less hydrotropism (Takahashi and Suge, 1991 Physiologia Plantarum 82: 24-31; Takahashi and Scott, 1993 Plant, Cell and Environment 16: 99-103). Here, we examine varietal differences in the hydiotropic response and its interaction with gravitropism in wheat roots. Primary seminal roots of wheat (Triticum aestivum L.) were hydrotropically stimulated by different moisture gradients established by placing wet cheesecloth and saturated solutions of different salts in closed chambers. From equations obtained by relative humidity (RH) at different distances from the wet cheesecloth, moisture gradients at the root-lip level were estimated to be 0.03 to 1.84% RH mm(-1), depending upon the salt introduced into the chamber. The roots showed positive hydrotropism in response to 0.67% RH mm(-1), and the response apparently increased as the gradient was strengthened. When the primary seminal roots of 12 cultivars were exposed to a moisture gradient of 1.84% RH mm(-1), hydrotropic response significantly differed depending upon the cultivar tested. Among the cultivars, the roots of Norin 11, Norin 15, Norin 117, and Norin 125 responded hydrotropically more strongly than the others. These roots, with the exception of Norin II, showed a less vigorous response to gravity compared to the remaining cultivars. However, the roots of Norin 20, Norin 38, and Norin 107 were relatively unresponsive to both a moisture gradient and to gravity. Thus, the primary seminal roots of wheat respond hydrotropically, and the responsiveness differs among cultivars. However, the varietal difference in hydrotropic response cannot be explained solely by converse differences in responsiveness to gravity.

In pole bean plants, mechanical stress (MS) inhibited stem elongation and induced radial thickening of the stem. Application of uniconazole, an inhibitor of gibberellin biosynthesis, also reduced stem growth but had no effect on stem diameter. Both MS and uniconazole significantly reduced hollowing of the first internodes, but only the former increased ethylene evolution from the first internode. Application of GA(3) increased the length of the first internode and decreased its diameter in bush bean plants; this was accompanied by a significant promotion of stem hollowing. Aminooxyacetic acid (AOA) decreased ethylene evolution from the GA(3)-treated internodes, though it did not reduce the GA(3)-induced hollowing of the first internodes. Application of GA(3) affected neither ethylene evolution nor cellulase activity in the first internodes of bush bean plants. Application of GA(3) stimulated much greater cell elongation in the center of pith tissue than in the outer surrounding tissues, suggesting a possible physical breakage of the inner cells, which leads the hollowing of bean stems. These results suggest that gibberellin is a factor responsible for stem hollowing in bean plants. Because MS is known to reduce gibberellin content in bean plants [Suge (1978) Plant Cell Physiol. 21:303] MS may inhibit stem hollowing by reducing the amount of endogenous gibberellin.

We have examined the graviresponding sites in the shoots of seedlings of rice (Oryza sativa L.) and their,relation to the agravitropic growth of a 'lazy' line. The graviresponding sites of the seedling shoots of a japonica type of rice, cv. Kamenoo, shifted from the mesocotyl/coleoptile region to the leaf-sheath base when the shoots grew in the dark. A lazy line of rice, lazy-Kamenoo, showed gravicurvature in the mesocotyl/coleoptile region at the early stage of growth, but eventually lost its graviresponse as the seedlings grew. The loss of graviresponsiveness of lazy-Kamenoo was attributed to a reduced response of the coleoptile and a diminished response of the leaf-sheath base to gravity. Later, the leaf-sheath base of lazy-Kamenoo became gravitropically incompetent, causing agravitropic growth of the shoots. Thus, shoots of lazy-Kamenoo lose graviresponsiveness in an organ-dependent fashion.

The interactions between vernalization and photoperiodic effects on the flowering of 12 turnip varieties were examined under controlled environment. Seedlings of all varieties bolted and flowered under a long-day condition of 24 h day-length (LD) when the germinated seeds had been pre-exposed to low temperature at 3-degrees-C (LT) for 30 days. Under the' LD condition, the ratio of flower formation of all the varieties except for 'Tennoji' significantly decreased as the duration of LT treatment was shortened to less than 7 days. With LT treatment of more than 14 days, 80 to 100% of plants in all varieties formed flower buds under LD. When the seedlings were subjected to either LT and subsequently grown under a short-day condition of 8 h day-length (SD) or 1,D without LT treatment, the number of plants that formed flower buds substantially decreased. Furthermore, the effect of seed vernalization was counteracted by subsequent SD conditions under which the vernalized seedlings were grown throughout the experiment. Namely, flower formation and bolting of the vernalized seedlings were significantly inhibited when the LT-treated seedlings were subsequently grown under SD. These flowering responses of turnip plants to temperature and photoperiod significantly differed among the varieties used. We have classified the turnip varieties into five groups. 'Tennoji', 'Shogoin', and 'Hakatasuwari' strongly responded to the single treatment of either LD or LT. resulting in a high ratio of flower formation (Type 1), whereas either treatment hardly induced the formation of flower bud in 'Yorii', 'Hinona', 'Ohnobeni'. and 'Kanamachi' (Type 11). 'Ohyabu', 'Ohmi', and 'Atsumi' showed an intermediate degree of flowering ratio between the Type I and II in repsonse to either LD or LT treatment (Type III). In 'Yamauchi', LD itself did not induce flower formation of non-vernalized plants, but the LT caused a high ratio of flower formation even under SD (Type IV). By contrast, 'Narusawana' showed a substantial ratio of flower formation due to LD without LT treatment, while the seedlings treated with LT did not flower under the following SD in this variety (Type V). Thus, the flowering of turnip plants is dramatically influenced by photoperiod as well as by temperature, and the responses to the two factors significantly differ among the varieties.

It has been proposed that peg formation in the vascular transition region (TR zone) between the hypocotyl and the root in Cucurbitaceae seedlings is a gravimorphogenetic phenomenon. Initiation of the peg became visible 36 h after imbibition when cucumber (Cucumis sativus L. cv. Burpee Hybrid II) seeds were germinated in a horizontal position at 24 degrees C in the dark. Simultaneously, sedimented amyloplasts (putative statoliths) were apparent in the sheath cells surrounding the vascular strands, and in the cortical cells immediately adjacent to them, in the TR zone. In contrast, the other cortical cells, some of which were destined to develop into the peg, contained amyloplasts which were not sedimented. These results suggest that the graviperception mechanism for peg formation may be like that of statoliths in shoot gravitropism. By 48 h following imbibition, the cells of the TR zone still had sedimented amyloplasts but had lost their sensitivity to gravity, possibly because of their maturation.

We have studied hydrotropism and its interaction with gravitropism in agravitropic roots of a pea mutant and normal roots of peas (Pisum sativum L.) and maize (Zea mays L.). The interaction between hydrotropism and gravitropism in normal roots of peas or maize were also examined by nullifying the gravitropic response on a clinostat and by changing the stimulus-angle for gravistimulation. Depending on the intensity of both hydrostimulation and gravistimulation, hydrotropism and gravitropism of seedling roots strongly interact with one another. When the gravitropic response was reduced, either genetically or physiologically, the hydrotropic response of roots became more unequivocal. Also, roots more sensitive to gravity appear to require a greater moisture gradient for the induction of hydrotropism. Positive hydrotropism of roots occurred due to a differential growth in the elongation zone; the elongation was much more inhibited on the moistened side than on the dry side of the roots. It was suggested that the site of sensory perception for hydrotropism resides in the root cap, as does the sensory site for gravitropism. Furthermore, an auxin inhibitor, 2,3,5-triiodobenzoic acid (TIBA), and a calcium chelator, ethyleneglycol-bis-(beta-aminoethylether) -N,N,N',N'-tetraacetic acid (EGTA), inhibited both hydro tropism and gravitropism in roots. These results suggest that the two tropisms share a common mechanism in the signal transduction step.

Roots of Pisum sativum L. and Zea mays L. were exposed to different moisture gradients established by placing both wet cheesecloth (hydrostimulant) and saturated aqueous solutions of various salts in a closed chamber. Atmospheric conditions with different relative humidity (RH) in a range between 98 and 86% RH were obtained at root level, 2 to 3 mm from the water-saturated hydrostimulant. Roots of Silver Queen corn placed vertically with the tips down curved sideways toward the hydrostimulant in response to approximately 94% RH but did not respond positively to RH higher than approximately 95%. The positive hydrotropic response increased linearly as RH was lowered from 95 to 90%. A maximum response was observed at RH between 90 and 86%. However, RH required for the induction of hydrotropism as well as the responsiveness differed among plant species used; gravitropically sensitive roots appeared to require a somewhat greater moisture gradient for the induction of hydrotropism. Decapped roots of corn failed to curve hydrotropically, suggesting the root cap as a major site of hydrosensing.

Orientation of root growth on earth and under microgravity conditions can possibly be controlled by hydrotropism-growth toward a moisture source in the absence of or reduced gravitropism. A porous-tube water delivery system being used for plant growth studies is appropriate for testing this hypothesis since roots can be grown aeroponically in this system. When the roots of the agravitropic mutant pea ageotropum (Pisum sativum L.) were placed vertically in air of 91% relative humidity and 2 to 3 mm from the water-saturated porous tube placed horizontally, the roots responded hydrotropically and grew in a continuous arch along the circular surface of the tube. By contrast, normal gravitropic roots of 'Alaska' pea initially showed a slight transient curvature toward the tube and then resumed vertical downward growth due to gravitropism. Thus, in microgravity, normal gravitropic roots could respond to a moisture gradient as strongly as the agravitropic roots used in this study. Hydrotropism should be considered a significant factor responsible for orientation of root growth in microgravity.

Ca2+ has been proposed to mediate inhibition of root elongation. However, exogenous Ca2+ at 10 or 20 millimolar, applied directly to the root cap, significantly stimulated root elongation in pea (Pisum sativum L.) and com (Zea mays L.) seedlings. Furthermore, Ca2+ at 1 to 20 millimolar, applied unilaterally to the caps of Alaska pea roots, caused root curvature away from the Ca2+ source, which was caused by an acceleration of elongation growth on the convex side (Ca2+ side) of the roots. Roots of an agravitropic pea mutant, ageotropum, responded to a greater extent. Roots of Merit and Silver Queen corn also responded to Ca2+ in similar ways but required a higher Ca2+ concentration than that of pea roots. Roots of all other cultivars tested (additional four cultivars of pea and one of corn) curved away from the unilateral Ca2+ source as well. The Ca2+-stimulated curvature was substantially enhanced by light. A Ca2+ ionophore, A23187, at 20 micromolar or abscisic acid at 0.1 to 100 micromolar partially substituted for the light effect and enhanced the Ca2+-stimulated curvature in the dark. Unilateral application of Ca2+ to the elongation zone of intact roots or to the cut end of detipped roots caused either no curvature or very slight curvature toward the Ca2+. Thus, Ca2+ action on root elongation differs depending on its site of application. The stimulatory action of Ca2+ may involve an elevation of cytoplasmic Ca2+ in root cap cells and may participate in root tropisms.

Vibration at 50 Hz significantly stimulated seed germination and root elongation in both rice and cucumber plants. The vibration barely affected the elongation of cucumber hypocotyls but stimulated the elongation of rice coleoptiles. Thus, plants' responses to vibration at a particular frequency differ from those to other mechanical stimuli.

We have partially characterized root hydrotropism and its interaction with gravitropism in maize (Zea mays L.). Roots of Golden Cross Bantam 70, which require light for orthogravitropism, showed positive hydrotropism; bending upward when placed horizontally below a hydrostimulant (moist cheesecloth) in 85% relative humidity (RH) and in total darkness. However, the light-exposed roots of Golden Cross Bantam 70 or roots of a normal maize cultivar, Burpee Snow Cross, showed positive gravitropism under the same conditions; bending downward when placed horizontally below the hydrostimulant in 85% RH. Light-exposed roots of Golden Cross Bantam 70 placed at 70-degrees below the horizontal plane responded positively hydrotropically, but gravitropism overcame the hydrotropism when the roots were placed at 45-degrees below the horizontal. Roots placed vertically with the tip down in 85% RH bent to the side toward the hydrostimulant in both cultivars, and light conditions did not affect the response. Such vertical roots did not respond when the humidity was maintained near saturation. These results suggest that hydrotropic and gravitropic responses interact with one another depending on the intensity of one or both factors. Removal of the approximately 1.5 millimeter root tip blocked both hydrotropic and gravitropic responses in the two cultivars. However, removal of visible root tip mucilage did not affect hydrotropism or gravitropism in either cultivar.

Shoots of a pea mutant, ageotropum (Pisum sativum L.) exhibited agravitropic growth and less ability to form a plumular hook in the dark. Decapitated epicotyls of dark-grown ageotropum pea bent away from external IAA when applied to the cut-end asymmetrically. But symmetrical application of IAA to the entire cut-surface of the horizontally placed epicotyls could not induce gravitropic curvature. In horizontally placed epicotyls of dark-grown ageotropum pea, [H-3] label applied as [H-3]IAA moved basipetally but did not apparently move laterally. Seedlings of ageotropum pea formed the plumular hook to some extent in response to externally applied ethylene. Alaska (normal) epicotyls evolved 2 to 5 times more ethylene than ageotropum epicotyls. Gravistimulation of the shoots increased ethylene evolution in Alaska epicotyls but not in ageotropum epicotyls. These results suggest a positive correlation between auxin asymmetry and ethylene production, which may influence both shoot gravitropism and formation of the plumular hook.

We have partially characterized root hydrotropism of an agravitropic pea mutant, ageotropum (from Pisum sativum L. cv. Weibull's Weitor), without interference of gravitropism. Lowering the atmospheric air humidity inhibited root elongation and caused root curvature toward the moisture-saturated substrate in ageotropum pea. Removal of root tips approximately 1.5 mm in length blocked the hydrotropic response. A computer-assisted image analysis showed that the hydrotropic curvature in the roots of ageotropum pea was chiefly due to a greater inhibition of elongation on the humid side than the dry side of the roots. Similarly, gravitropic curvature of Alaska pea roots resulted from inhibition of elongation on the lower side of the horizontally placed roots, while the upper side of the roots maintained a normal growth rate. Gravitropic bending of Alaska pea roots was apparent 30 min after stimulation, whereas differential growth as well as curvature in positive root hydrotropism of ageotropum pea became visible 4-5 h after the continuous hydrostimulation. Application of 2,3,5-triidobenzoic acid or ethyleneglycol-bis-(beta-aminoethylether)-N,N,N',N'-tetraacetic acid was inhibitory to both root hydrotropism of ageotropum pea and root gravitropism of Alaska pea. Some mutual response mechanism for both hydrotropism and gravitropism may exist in roots, although the stimulusperception mechanisms differ from one another.

Mechanical perturbation (MP) applied unilaterally to cucumber (Cucumis sativus L.) hypocotyls induced thigmotropic curvature toward the stimulus. Gravitropic or phototropic curvature of the hypocotyl was inhibited by symmetrical application of MP to both sides of the hypocotyl. When both MP and IAA were unilaterally applied simultaneously to the same side, the hypocotyls always bent toward the MP stimulus, as in thigmotropism alone. Thus, the exogenous IAA did not control the direction of curvature. Aminoethoxyvinyl glycine (AVG) blocked thigmotropism as well as gravitropism and phototropism but promoted IAA-induced curvature. MP-stimulated ethylene evolution peaked about 4 h after MP, followed by a peak of thigmotropic curvature. For all tropisms, more ethylene evolved from the stimulated side than from the other side of the hypocotyls. MP-induced ethylene acting as a growth inhibitor, auxin-transport inhibitor, and/or modulator of tissue sensitivity to auxin, may be involved in thigmotropism and MP-induced inhibition of various tropisms. Ethylene produced as a result of MP was not affected by the removal of cotyledons. This MP-induced ethylene was additive to that of phototropically or gravitropically stimulated ethylene.