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Plant Cell Physiol. 39(9): 914-921 (1998) JSPP © 1998 Cell Elongation and Revolving Movement in Phaseolus vulgaris L. Twining Shoots Anne-Frangoise Care, Leonid Nefed'ev, Bernard Bonnet, Bernard Millet and Pierre-Marie Badot' Sciences Vdgetales, Laboratoire de Biologie Ecophysiologie, University de Franche-Comte, Place Leclerc, F-25030 Besancon Cedex, France In Phaseolus vulgaris L., the shoot displays a revolving movement that occurs rhythmically in a highly regular manner. Previous data led to think that revolving movement is driven by turgor and volume changes in the epidermal cells of the bending zone. To document this hypothesis, the time course of in situ cell length variations in the bending zone was measured during the movement of the shoot and related to the phase of the revolving movement. Each ten minutes, a photograph of cells was taken and the revolving movement was simultaneously recorded using time-lapse microphotography and video-monitoring. In the moving part of the shoot, epidermal cells displayed partly reversible length variations during their growth. Data were processed by Fourier analysis to determine whether or not a periodicity exists. Rhythm in cell length variations was evidenced only when initial cell lengths were ranged between 60 and 120 fan. In this case, the period corresponds to that of the revolving movement. Thus, revolving movement is related to partly reversible length variations in the cells of the bending zone. These results agree with the hypothesis of an involvement of turgor mediated volume changes in the revolving movement. Key words: Cell elongation — Circumnutation — Phaseolus vulgaris L. — Revolving movement — Turgor changes. The French bean (Phaseolus vulgaris L.) twining shoots display a revolving movement, usually known as a circumnutation movement (Darwin 1865, Baillaud 1958, Johnsson 1979). This movement takes place in the terminal part of the shoot just below the apical bud. The moving zone consists of a linear horizontal apical part and a curved part. The trajectory of the apex is more or less helicoidal (Millet et al. 1984). The period of the movement is about 90 minutes at 25°C. The direction is constant and positive (counterclockwise). The revolving movement occurs as a result of the bending of the upper regions of the shoot. Each time the shoot 1 To whom correspondence should be adressed. (fax 33 3 81 66 56 98, email: [email protected]) 914 ceases to move, the curvature disappears. For a long time, it was thought that the curvature was the consequence of unequal growth on the opposite sides of the shoot (Baillaud 1958). The side with the highest growth rate becomes convex; the side with the lowest becomes concave. Thus, the revolving movement has usually been considered as a growth movement. However, Millet et al. (1988) noted a discrepancy between this theory and experimental facts. A calculation of the required growth rate was done using shoot geometry and the movement period. This theoretical growth rate was compared to the measured one showing that the daily enlargements of the bending zone are much lower than expected. It was concluded that the revolving movement mechanism could not be limited to rhythmic variations in cell elongation and might involve other processes such as turgor and volume changes. Scanning electron micrographs of the bending zone showed that epidermal cells appeared fully turgid with smooth walls when located in the convex side. On the other hand, when they were located in the concave side, epidermal cells showed transverse folds suggesting a lower turgor pressure. Other observations show that the relative water content was higher in the convex side than in the concave one (Badot 1987). Changes in osmotic potential and ion distribution were also evidenced during circumnutation (Badot et al. 1990). These previous data imply that partly reversible cell volume variations must be responsible for the revolving movement. The movement cannot be restricted to a growth phenomenon (irreversible cell volume variations), but must involve a reversible component such as turgor-driven changes. Other cells are known to undergo turgor changes that generate movement. Stomatal guard cells, for example, display large and reversible volume variations due to changes in cellular osmotic potential that are converted into turgor pressure modifications (Gorton 1990, Meidner and Bannister 1979). The leaf movement also originates from turgor-driven volume variations of the pulvinus parts usually denned as extensor and flexor (Mayer et al. 1985, Freudling et al. 1988). Estimations of cell volume changes in the bending zone would allow us to describe the revolving movement through the successive swelling and shrinking of cells. Changes in cell volume can be evaluated by measuring cell length variations. In this way, simultaneous recordings of Cell growth and revolving movement in bean 915 cell length variations and movement could help us to evidence that revolving movement originates in partly reversible volume variations of the cells in the bending zone. Thus, the aim of this paper is to investigate in situ cell growth in the moving part and to measure cell length variations in the bending zone of a revolving bean shoot. In other plant materials, estimations of variations in cell volume were often obtained indirectly from length measurements of shoot segments. Most of these previous results concerned the whole plant or part of it (Michelena and Boyer 1982, Luthen and Bottger 1992). In the same way, Baskin et al. (1985) analyzed the elongation pattern of small areas of maize coleoptile epidermis from time-lapse photomicrographic records. Other workers have used more direct ways to record cell volume changes. For example, Omasa et al. (1983) used an image instrumentation system to observe the stomatal response of plants to environmental changes. In this study, we intend to determine the in situ variations of cell elongation and to establish a clear relationship between it and the revolving movement of bean shoots. Materials and Methods Plant material and growth conditions—Phaseolus vulgaris L., cv. Blanc de Juillet seeds were provided by S.P.G. (Avignon, France). Plants were planted in 175-ml pots filled with vermiculite and watered with tap water. They were grown under constant conditions of light (40 W m~ 2 ), relative humidity (65±5%) and temperature (25 ± 1°C) as previously described by Millet et al. (1988). Fig. 1 A photo-micrograph of the surface of a Phaseolus vulgaris L. bean shoot. This is a printed example of one of the negatives from which the measurements were made. The four dye droplets delimit the three cells whose elongation was measured. These cells are located along the same generating line (arrowheads). The horizontal bar represents 100 nm. The plants were analyzed at the age of 12 d. At this stage of development, the shoots were 26-cm-long and the revolving movement Fig. 2 A schematic representation of the experimental setup built to measure in situ variations of the cell length in a Phaseolus vulgaris L. bean shoot, and to simultaneously record the apex revolving movement. 1, video camera; 2, French bean shoot; 3, green light (cold light source); 4, colored ball; 5, horizontal inverted microscope; 6, stand; 7, micro-manipulator; 8, camera; 9, video monitor. Cell growth and revolving movement in bean 916 had already started. Measurement of mean cell length along the shoot—Cell lengths were measured along the shoot of 6 different 12-day-old bean plants. Microphotographs of illuminated cells were taken every 1.5 cm along the 13.5 first cm of the shoot at a 250-fold magnification (Leica, Wild MPS 48). After calibration, 50 cells were measured at each level. Mean lengths, ± the standard deviation (SD) for p=0.05 were calculated. Measurement of cell elongation—The bean plant was put into a test tube so that the roots were placed in distilled water. Then, the tube was fixed to a holder and layed on a microscope stage. Epidermal cells in the beginning of the bending zone (6+1.5 cm from the apex) were marked with a micro-instrument made in the following way: a micropipette was pulled from a 1 mm outer diameter capillary with a Sutter Instruments Co. puller (Novato, Californie, U.S.A.). Then, the tip was closed by flame so that its final diameter was about 100 fim. It was coated with a viscous dye made up of a mix of honey and neutral red. The micropipette was fixed on a Leitz micromanipulator and observed with a microscope at a 100-fold magnification. A dye droplet of about in diameter was left at each end of an epidermal cell. One to three cells were marked on the same plant (Fig. 1). Afterwards, the tube was placed on a micromanipulator in the viewing field of a Leica inverted microscope (Fig. 2). A black and white photograph of the marked cells (Leica, Wild MPS 48) was taken every ten minutes at a 40-fold magnification (objective lens magnifying 10-fold, with a numerical aperture of 0.2 and a working distance of 2.3 cm). Time intervals were measured with a stopwatch. To increase illumination, a cold light source (Schott Glaswerke, KL 1500, Wiesbaden, Germany) with a green filter lit the shoot. The time course of cell length variations was then measured using an enlarger (Durst D 659, Bolzano-Bozen, Italy) to obtain an image that was magnified 280-fold. Data calibration was carried out as follows: at the beginning and at the end of each experiment, the distance between two droplets was measured with an ocular micrometer. The measurements of cell elongation were carried out with 38 cells, which were from 40 to 150//m long. Movement monitoring—At the same time, the revolving movement of the bean shoot was monitored with a video camera (Burle, Lancaster, U.K.) placed above the shoot. The obtained im- Orientation of the plane containing shoot curvature before (a,b and d) and after (c) device rotation Schematic representation o( the shoot seen in cross section before (a.b and d) and after (c) device rotation Schematic representation the same shoot after device rotation Orientation of the plane containing shoot curvatun after device rotation <•") (b") (d") Measurement of the distance between two dye droplets Fig. 3 Bean shoot seen from above at the beginning of the experiment (t0), after ten (t o +10) and twenty minutes (t o +20). The shoot is drawn in the bending zone where some cells were marked. The black point on the shoot epidermis represents a dye droplet. The half part of the shoot seen from above and coloured in grey with a bold surface corresponds to the convex side (ex). The other is the concave one (cv). The shoot displays a counterclockwise revolving movement. The plant is placed in the viewing field of an inverted microscope. At to, in (a), the plane which contains the shoot curvature was perpendicular to the optical axis. In such a configuration, it was not necessary to rotate the tube-plant device (a'). At t o +10, in (b), the convexity moved around the shoot. In order to avoid any geometrical mistake of the distance measured between the two marks (see "Methodological precautions"), the tube-plant device was rotated (manual rotation) to restore the perpendicular between the curvature and the optical axis (b'). The new position of the shoot was represented in (c). At t o + 20, in (d), the curvature moved again around the shoot and the tube-plant device was rotated once again to take a photograph at this time (d'). (a"), (b") and (d") display the way to measure the distance between the two dye droplets (cell length). They show that the angle between the optical axis of the objective and the plane which contained shoot curvature was always 90°. Cell growth and revolving movement in bean age was observed on a monitor (Electrohome LTD., Kitchener, Ontario, Canada). The position of the apex was recorded on screen every time a photograph was taken using the orthogonal projections (Tronchet 1942). These data were digitized according to Millet et al. (1984). Methodological precautions—When several cells were marked on the same shoot, they were always located along the same longitudinal line. Before taking each photograph, the tubeplant device was rotated so that the plane, which contained the curvature, was perpendicular to the objective axis. Indeed, the bending of the plant moved the surface of interest out of the plane perpendicular to the optical axis, and this was a source of error. This movement caused a projection of the length to be recorded. Figure 3 explains the geometrical relations between optical axis and the plane containing the curvature during the measurement. The rotation of the device allowed us to avoid any geometric mistake of the distance measured between the two marks: this distance was always set perpendicular to the optical axis at the time of the measurement. This implied locating the exact position of the system at each data point before rotating. Four coloured balls were placed on the tube and their position was systematically noted each ten minutes. Thus, it was possible to determine the apex relative position in relation to the tube, and therefore, in relation to the plant base even if rotations were necessary for the cell length measurements. The recording of movement and cell elongation began after a stabilization period of ninety minutes to rule out the disturbances due to handlings. Mathematical data processing—The possible periodicity of cell length variations was checked by a mathematical processing of the data using a method adapted from Assaad (1985). First, trend (growth) was eliminated by the least squares method. Then, data were processed by the Fourier spectral analysis. A time form y = f(t), where t is the time passing, was converted into a frequential form y=f(v), where v is a time frequency. The Fourier theorem explains that a periodic function can be split up into a sine and cosine sum for successive harmonics of the basic frequency (Brillouin 1939): 917 different shoots are shown in Fig. 4. Signals digitized from the revolving movement of these shoots are given in Fig. 5a, 6a, and 7a after elimination of the trend by the least squares method. Fig. 5b, 6b, and 7b show the signals issued from the corresponding cell length variations after the same mathematical treatment. The experimental conditions were in such a way that maxima in signal digitized from revolving movement were observed when the marked cells were located in the concavity (Fig. 5a, 6a, 7a); on the contrary, minima were noted when these cells were located on the convex side. Corresponding power spectra obtained after data processing by the Fourier analysis are shown in Fig. 5c, 6c, 7c, and 5d, 6d, 7d. For the entire experimental duration, the 55 jum long cell elongated in a linear way and measured 85 //m at the end of the experiment (Fig. 4). Moreover, reversible and transient changes in cell length were observed. They increased with time from 2fim for the four first hours and later increased to 8 ^m. No periodicity was shown after processing by the Fourier analysis (Fig. 5d). In that case, cell elongation displayed no rhythmicity. The period of the corresponding shoot revolving movement was 75 min (Fig. 5c). In Fig. 4, another cell elongated from HO^/m to 160 /im in 8 h. As in the case of the shorter cell, its elongation was not completely linear. The variations of cell length were only partly irreversible and displayed oscillations. Six cycles, and consequently 6 higher cell lengths, were recorded (lh, 2h30, 3hl0, 4h40, 6h, 7h20). Cell length was observed in relation to its position in the bending zone (con- 1 where vo=— F(t)=Ao+ Z (An l The Fourier coefficients—AQ, An and Bn—represent the successive sine and cosine amplitudes of the periodic function F(t) to the nlh value of a whole number. These coefficients are calculated as follows (Crawford 1972): F(t)dt An = 2v o l F(t)sin 27TVontdt for any to, value of t B n =2v 0 ( °F(t)cos 27tV(,ntdt K Results are expressed in terms of a frequency spectrum that decomposes into an amplitude spectrum a(n) and a phase spectrum (n). I o(n) | = x o si tfn)=Arctg( - | j Results Representative examples for the time course of length variations of three cells with different initial lengths (55 pm, WOfim and 140 fim) in the bending zone of three Fig. 4 Time course of length variations for three representative epidermal cells with different initial lengths (55 fim, 110/<m and 140/<m). 918 Cell growth and revolving movement in bean PERIODS (min) Ill-Ill. ••— irt-OHi^vomiTfl-'tnnc PERIODS (min) Fig. 5 (a) Successive orthogonal projections of the apex movement during 8 h. (b) Time course of elongation for an epidermal cell with an initial length of 55 //m. (a) and (b) were calculated after mathematical processing of the rough data by the least squares method (trend out). When minima were recorded on curve (a), the marked cell was located on the convex side (dotted line), (c) and (d) The power spectra obtained from (a) and (b), respectively, after processing them with the Fourier spectral analysis. vexity, concavity). The 110-/vm cell was longer when it was located in the convexity (dotted lines in Fig. 6a, b). On the other hand, in the concavity, the cell was shorter. The shoot revolving movement occurred with a 75-min period similar to that of cell elongation (Fig. 6c, d). During the recording, the longer cell (140 /an) elongated linearly for the most part (Fig. 4). It measured 205 fun after 8 h. Cell length varied with a low amplitude. In that case, the Fourier analysis showed no periodicity for cell length (Fig.7d), although the shoot revolving movement completed rhythmic cycle every 80 min (Fig. 7c). The elongation pattern of thirty-five cells on fourteen different shoots and their corresponding revolving movement were recorded and processed in the same way as the previous ones. The periods, when detected by the Fourier analysis, and initial length of the corresponding cells were reported in Fig. 8. The cells that were marked in the beginning of the bending zone measured from 45 to 150/an. However, no rhythmicity of cell elongation was shown for the cells between 45 and 65 fxm, and for the ones longer than 120 fim (white circles). Cell length variations were periodic for most of the cells from 65 to 120 fim. Moreover, the difference between the cell length variations period and the corresponding apex revolving movement period is no 15 PERIODS (min) PERIODS (min) Fig. 6 (a) Successive orthogonal projections of the apex movement during 8 h. (b) Time course of elongation for an epidermal cell with an initial length of 110 fim. (a) and (b) were calculated after the mathematical processing of the rough data by the least squares method (trend out). When minima were recorded on curve (a), the marked cell was located on the convex side (dotted line), (c) and (d) The power spectra obtained from (a) and (b), respectively, after processing them with the Fourier spectral analysis. Cell growth and revolving movement in bean §? 3 w 15- a q / / 32 -5-15- V1 i \ /^ F\ T\ /V / V \ /r V\ f\ 919 ?R ft \ // \\ // \ ' V w w \/\/ 1 ' 3 4 1 5 i r PERIODS (min) TTMEQi) 60 iL.-.i.-i.Ji-i.i. PERIODS (min) Fig. 7 (a) Successive orthogonal projections of the apex movement during 8 h. (b) Time course of elongation for an epidermal cell with an initial length of 140 ftm. (a) and (b) were calculated after the mathematical processing of the rough data by the least squares method (trend out). When minima were recorded on curve (a), the marked cell was located on the convex side (dotted line), (c) and (d) The power spectra obtained from (a) and (b), respectively, after processing them with the Fourier spectral analysis. greater than 15 min (black plots joined by a dotted line). A statistical correlation was evidenced between the two kinds of periods (correlation coefficient R 2 =0,921). Among these twenty-one cells, only four were found to elongate in a non-rhythmic way. Fig. 9 allows us to estimate the mean cell length versus the distance from the apex. Along the 11 first cms, growth appears regular. Then, it ceases and the maximum length statistically evaluated for the epidermal cells is 160//m. At 130 120 .= • * 110 - Q o 2 Discussion In the course of their growth, most of the cells in the bending zone of the bean shoot display partially reversible length variations (Fig. 8). These changes are rhythmic. All the cells that elongate in a partly reversible and rhythmic way had initial lengths from 65 fim to HO^m. Otherwise, our results show that some cells in the bending zone do not display reversible length variations or elongate in a partially reversible, but unrhythmical way. Among these cells, we • Cell elongation o • Revolving movement • the distance chosen for cell labeling, cells are found to measure statistically from 57±SD to 90±SD. An individual variability is observed in cell length for a particular distance from the apex. Thus, the cells randomly chosen for cell labeling are sometimes longer or shorter than the statistical means. IOO. o oooo# *§• * 90- E a 80- s 70 40 150- 60 80 100 120 140 160 INITIAL LENGTH OF CELLS dim) Fig. 8 Periods of cell elongation (•) and shoot revolving movement (o, •) versus the initial length of the cells. Periods of the movement represented with white circles correspond to records without periodicity for cell length variations. Cell length and revolving movement periods represented in black differ from less than fifteen minutes. The dotted lines join the coupled plots corresponding to the same record. I t 100- 50- DISTANCE FROM THE APEX (cm) Fig. 9 Variations of cell length (mean ± SD; n = 50; p=0.05) versus the distance from the apex in six different bean shoots. 920 Cell growth and revolving movement in bean find all the cells with an initial length lower than 65 #m and higher than 110 //m. On the other hand, all the cells with initial lengths from 65 to 110/rni do not inevitably undergo partly irreversible and/or periodic length changes. Some of them do not exhibit any reversibility of length variations. Others elongate in a partly reversible way but their length changes display no periodicity. The shortest cells are preferentially located in the upper part of the shoot (Fig. 9) and are the youngest ones. The longest cells are preferentially found below the curvature and are statistically the oldest ones. Thus, it seems that along the time, cells are first unable to display partly reversible length changes, then they become able to express this behaviour when located in the curvature. Afterwards, they lose this feature when they become older. However, individual variability of cell length explains that some of the shortest and the longest cells measured in these experiments may be found in the curvature. In this part of the shoot, the proportion of very short and long cells is low as observed in data given in Fig. 9. If we explain the twining movement by the alternative swelling and shrinking of the cells on opposite sides of the bending zone, it seems difficult to understand how some of these cells do not undergo any reversible nor rhythmic length variation. In that case, no revolving movement would occur. From a methodological point of view, we may attribute a part of these particular results to disturbances due to cell labeling. Indeed, touching cells and epidermal hairs with a micro-instrument, even without any visible damage, could have affected some of them and caused a mechanical stress. This one seems to have disturbed only the reversible part of cell length variations and not the growth. Previous experiments requiring growth measurements were usually performed with ink (Kutschera and Kohler 1994, Pfeiffer and Kutschera 1995) or with a black sheep hair placed directly on the coleoptile to provide a standard length (Baskin et al. 1985). These works did not intend to record one single cell elongation in situ, but the growth rate of shoot segments. Thus, it is not surprising that the disturbances due to labeling did not affect the results at the whole organ level. Moreover, the mathematical processing used in the present study is very discriminating. The Fourier analysis is strict, and therefore, does not allow for any irregularity. It implies a number of constraints such as a minimum and a whole number of periods. The signal has to be as regular as possible before processing. This could partly explain why some cells were not found to elongate in a periodic way. For the shortest ones, amplification of cell length variations could be too low to have been detected by the Fourier analysis. These methodological considerations tend to prove that the most significant results are the rhythmic pattern in cell elongation displayed by most of the studied cells. The cells in the bending zone of the bean shoot show rhythmic and partly reversible length variations (Fig. 6). Growth oscillations have already been observed in stems or other plant materials. Kristie and Joliffe (1986) used a high-resolution transducer system to obtain a continuous record of stem elongation rate in seedlings of different plant species. In most cases, they observed short period oscillations of growth and suggested that they were caused by synchronized variations in the rate of cell elongation. Other results deal with the so-called pulsatory growth of pollen tubes (Derksen 1996 and references therein). After short phases of rapid extension (10-20 s long), growth of the tubes is interrupted for long periods (up to minutes long). However, no reversible variation in cell length was reported by these authors. In bean shoot, the partly reversible variations of cell length could be a key component of the revolving movement mechanism. Cell elongation on the opposite sides of the bending zone runs in an opposite, but coordinated way. Cells exhibit a maximum length on the convex side, whereas they display a minimum on the concave side. Moreover, the period of the cell length variations corresponds to that of the apex revolving movement. If reversibility occurs in cell length variations, it means that cell elongation, i.e. growth is not the only phenomenon involved in the revolving movement mechanisms. To explain the reversibility component of the cell length variations pattern, we have to take into account reversible phenomena such as turgor variations. This fully agrees with the theoretical calculations from Millet et al. (1988) who showed that revolving movement could not be limited to a growth movement. Periodic volume—and therefore turgor—variations could successively affect the cells all around the shoot in the curvature and, thus could be partly responsible for the revolving movement of twining shoots. During one revolution, cells in the bending zone alternatively swell and shrink. Whether or not these volume changes are due to osmotic changes or to cell wall loosening is still unknown. Badot (1987) measured osmotic potential on bean epidermal strips taken from the concave and convex sides of the curvature zone and found that cell osmotic potential displayed periodical changes. The relationships between cell volume, turgor pressure, and osmotic potential variations are insufficiently investigated and remain unclear in our material. Changes in cell length evidenced in twining shoots could be correlated to concomitant variations in cell turgor as observed in sunflower hypocotyls (Kutschera and Kohler 1993). Turgor pressure is generally conceived as the driving force for wall yielding and extension (Cosgrove 1987). When turgor exceeds a yield threshold, cell wall extends (Passioura and Fry 1992) and this may result in cell growth. Previously, Millet et al. (1988) observed a stratification of the wall in the epidermal cells of twining shoots and hypothesized that this corresponds to intense polysaccharides deposits on microfibrils. Thus, previous and present results are consistent to the hypothesis that revolving movement is due to an alternative swelling and shrinking occuring in Cell growth and revolving movement in bean growing epidermal cells. The present observations demonstrate that the revolving movement could be an interesting model system to investigate the relationships between cell elongation and turgor changes. Biochemical and cytological studies have to be undertaken to explain how the wall materials of the cells are successively stretched then cleaved by wall-loosening enzymes. The involvement of expansins in the responses of walls and their regulation (McQueen-Mason et al. 1992, McQueen-Mason and Cosgrove 1995, Cosgrove 1997) have also to be investigated in bean cells. On the other hand, direct measurements of turgor pressure in the epidermal cells of the bending zone with a pressure probe would be the best way to confirm the present results. References Assaad, C.I. (1985) Growth rhythm in tomato. J. Interdiscipl. Cycle Res. 16: 118-119. Badot, P.M. (1987) Approche cellulaire du mecanisme du mouvement revolutif des tiges volubiles. Etude de quelques parametres physicochimiques. Ann. Sci. Univ. Besancon, Biol. Veg. 4, 7: 53-110. Badot, P.M., Melin, D. and Garrec, J.P. (1990) Circumnutation in Phaseolus vulgaris L. II. Potassium content in the free-moving part of the shoot. Plant Physiol. Biochem. 28, 1: 123-130. Baillaud, L. (1958) La circumnutation des tiges volubiles et des vrilles. Annee Biol. 34: 17-28. Baskin, T.I., lino, M., Green, P.B. and Briggs, W.R. (1985) High-resolution measurement of growth during first positive phototropism in maize. Plant Cell Environ. 8: 595-603. Brillouin, L. (1939) Notions elementaires de Mathe'matiques pour les Sciences Expirimentales. pp. 100-103. Masson, Paris. Cleland, R. (1971) Cell wall extension. Annu. Rev. Plant Physiol. 22: 197222. Cosgrove, D.J. (1987) Wall relaxation and the driving forces for cell expansive growth. Plant Physiol. 84: 561-564. Cosgrove, D.J. (1997) Relaxation in a high-stress environment: the molecular bases of extensible cell walls and cell enlargement. Plant Cell 9: 10311041. Crawford, F.S. (1972) Cours de Physique Berkeley. Vol. 3, Ondes. pp. 6 1 63. Armand Colin, Paris. Darwin, C. (1865) On the movements and habits of climbing plants. / . Linn. Soc. Bot. 9: 1-118. Derksen, J. (1996) Pollen tubes: a model system for plant cell growth. Bot. Ada 109: 341-345. Freudling, C , Starrach, N., Flach, D., Gradmann, P. and Mayer, W.E. 921 (1988) Cell wall as reservoirs of K+ ions for reversible volume changes of pulvinar motor cells during rhythmic leaf movements. Planta 175: 193203. Gorton, H.L. (1990) Stomata and pulvini: a comparison of two rhythmic, turgor-mediated movement systems. In The Pulvinus: Motor Organ For Leaf Movement. Edited by Satter, R.L., Gorton, H.L. and Vogelmann, T.C. pp. 223-237. The American Society of Plant Physiologists. Johnsson, A. (1979) Growth movements not directed primarly by external stimuli. Circumnutation. In Encyclopedia of Plant Physiology. New series, 7. pp. 627-646. Springer Verlag, Berlin. Kristie, D.N. and Joliffe, P.A. (1986) High-resolution studies of growth oscillations during stem elongation. Can. J. Bot. 64: 2399-2405. Kutschera, U. and Kohler, K. (1993) Turgor pressure and elongation growth in developing Sunflower hypocotyls. J. Plant Physiol. 141: 757758. Kutschera, U. and Kohler, K. (1994) Cell elongation, turgor and osmotic pressure in developing sunflower hypocotyls. J. Exp. Bot. 45: 591-595. Liithen, H. and Bottger, M. (1992) A high-tech low-cost auxanometer for high-resolution determination of elongation rates in six simultaneous experimental setups. Mitt. Inst. Allg. Bot. Hamburg 24: 13-22. Mayer, W.E., Flach, D., Raju, M.V.S., Starrach, N. and Wiech, E. (1985) Mechanics of circadian pulvini movements in Phaseolus coccineus L. Shape and arrangement of motor cells, micellation of motor cells, and bulk moduli of extensibility. Planta 163: 381-390. McQueen-Mason, S J . and Cosgrove, D.J. (1995) Expansin mode of action on cell walls. Analyses of wall hydrolysis, stress relaxation, and binding. Plant Physiol. 107: 87-100. McQueen-Mason, S.J., Durachko D.M. and Cosgrove, D.J. (1992) Endogenous proteins that induce cell wall expansion in plants. Plant Cell 4: 1425-1433. Meidner, H. and Bannister, P. (1979) Pressures and solute potentials in stomatal cells of Tradescantia virginiana. J. Exp. Bot. 30: 255-265. Michelena, V.A. and Boyer, J.S. (1982) Complete turgor maintenance at low water potentials in the elongating region of maize leaves. Plant Physiol. 69: 1145-1149. Millet, B., Melin, D. and Badot, P.M. (1988) Circumnutation in Phaseolus vulgaris L. I. Growth, osmotic potential and cell ultrastructure in the free-moving part of the shoot. Physiol. Plant. 72: 133-138. Millet, B., Melin, D., Bonnet, B., Ibrahim, C.A. and Mercier, J. (1984) Rhythmic circumnutation movement of the shoots in Phaseolus vulgaris L. Chronobiol. Int. 1: 11-19. Omasa, K., Hashimoto, Y. and Aiga, I. (1983) Observation of stomatal movement of intact plants using an image instrumentation system with a light microscope. Plant Cell Physiol. 24: 281-288. Passioura, J.B. and Fry, S.C. (1992) Turgor and cell expansion: beyond the Lockhart equation. Aust. J. Plant Physiol. 19: 565-576. Pfeiffer, I. and Kutschera, U. (1995) Sucrose metabolism and cell elongation in developing sunflower hypocotyls. J. Exp. Bot. 46: 631-638. Tronchet, A. (1942) Techniques pour I'etude des mouvements des vrilles. Ann. Sc. Lyon Sc. Nat. 3: 27-44. (Received September 16, 1997; Accepted June 18, 1998)