TY - JOUR
AU - Boyer, John S.
AB - Abstract Profiles of water potential (Ψw) were measured from the soil to the tips of growing leaves of maize (Zea mays L.) when pressure (P) was applied to the soil/root system. At moderately low soil Ψw, leaf elongation was somewhat inhibited, large tensions existed in the xylem, and Ψw were slightly lower in the elongating leaf tissues than in the xylem, i.e. a growth‐induced Ψw was present but small. With P, the tension was relieved, enlarging the difference in Ψw between the xylem and the elongating tissues, i.e. enlarging the growth‐induced Ψw, which is critical for growth. Guttation occurred, confirming the high Ψw of the xylem, and the mature leaf tissue rehydrated. Water uptake increased and met the requirements of transpiration. Leaf elongation recovered to control rates. Under more severe conditions at lower soil Ψw, P induced only a brief elongation and the growth‐induced Ψw responded only slightly. Guttation did not occur, water flow did not meet the requirements of transpiration, and the mature leaf tissues did not rehydrate. A rewatering experiment indicated that a low conductance existed in the severely dehydrated soil, which limited water delivery to the root and shoot. Therefore, the initial growth inhibition appeared to be hydraulic because the enlargement of the growth‐induced Ψw by P together with rehydration of the mature leaf tissue were essential for growth recovery. In more severe conditions, P was ineffective because the soil could not supply water at the required rate, and metabolic factors began to contribute to the inhibition. Key words: Gradients, leaf elongation, osmotic potential, turgor, Zea mays L. Received 17 February 2003; Accepted 10 July 2003 Introduction This study was undertaken to determine whether growth‐induced water potentials (growth‐induced Ψw) respond to the pressurization of plant roots (P), with consequences for growth. Growth‐induced Ψw form when cell walls yield during growth (Boyer, 2001), preventing turgor pressure (Ψp) from reaching its maximum (Boyer, 1968; Westgate and Boyer, 1984, 1985). In multicellular plants, these potentials form gradients of declining Ψw extending downward and outward from the xylem into the growing tissues (Boyer, 1968; Fricke, 1997, 2002; Fricke et al., 1997; Fricke and Flowers, 1998; Martre et al., 1999; Molz and Boyer, 1978; Nonami and Boyer, 1993; Tang and Boyer, 2002; Westgate and Boyer, 1984, 1985). The gradient or field in three dimensions is the driving force moving water from the xylem into the growing tissues. If the xylem water potential (X) decreases, the field can reverse, reversing the force and flow, and preventing water uptake and growth. In theory, the reversal can be countered by P that raises X. A 1:1 correspondence was found between X and P in certain experiments (Hsiao et al., 1998; Wei et al., 1999). Moreover, leaves usually hydrate when P is applied (Janes and Gee, 1973; Munns et al., 2000; Nulsen et al., 1977; Passioura and Munns, 2000; Westgate et al., 1996). The P‐induced rehydration sometimes caused elongation to increase (Nonami et al., 1997; Tang and Boyer, 2002) but not always (Passioura, 1988; Tang and Boyer, 2002). If P was continuously applied, the growth inhibition normally caused by low Ψw still occurred (Passioura, 1988). These varied responses are of interest because applying P to the soil/root system should provide a critical test of whether the growth inhibition is controlled by hydraulic or non‐hydraulic factors (Nonami et al., 1997; Passioura, 1988; Tang and Boyer, 2002). When P is applied, only a small amount of water is required to increase Ψw in the xylem because its volume is small and its walls are quite rigid. Likewise, small amounts of water can change the Ψw in a few cells next to the xylem that will alter the field of growth‐induced Ψw and water flow into the entire growing region (Nonami et al., 1997; Tang and Boyer, 2002). But large amounts of water may be required to rehydrate the mature tissues. Therefore, conditions may exist where insufficient water is mobilized by P, and the expected rehydration does not occur. In the following work, these conditions and their consequences are explored. Materials and methods Plant materials Maize seed (Zea mays L. cv. B73×Mo17, Illinois Foundation Seeds, Inc.) was germinated between two sheets of paper towels wetted with nutrient solution at 29 °C in a dark and humid room. The nutrient solution was 12 mM Ca(NO3)2, 8 mM KNO3, 4 mM KH2PO4, 4 mM MgSO4, 50 µM H3BO3, 20 µM MnSO4, 4 µM ZnSO4, 1 µM CuSO4, 1 µM H2MoO4, and 250 µM Fe‐citrate (modified Hoagland’s solution). At 3 d, the seedlings were transferred individually to Plexiglas pots (6 cm inner diameter, 23 cm depth) with holes for drainage. Each pot was filled with a 1:1:1 by vol. mix of soil, peat moss and vermiculite at pH 6.5. For future exposure to P, the pot was covered with a pressure chamber top through which the coleoptile emerged. The seedling continued to grow in darkness until the mesocotyl passed through the opening in the top. After the mesocotyl had grown sufficiently, the plant and top were transferred to a controlled environment chamber (Environmental Growth Chambers, Ohio, USA) with a day/night temperature of 25/20 °C, relative humidity of 60/90%, and photosynthetically active radiation of 700–800 µmol m–2 s–1 for 14 h from daylight fluorescent bulbs. Because the pot was attached to the pressure chamber top, later‐developing nodal roots were prevented from entering the soil. Nutrients were supplied by flooding the whole pot from the bottom upward, then draining away the excess. Dehydration was imposed by withholding the nutrient solution from the soil. Leaf elongation After leaf 5 (from the stem base) emerged around 11 d after planting (dap), the pot was placed inside a pressure chamber (7.6 cm inner diameter, 26.7 cm depth), and the top through which the mesocotyl had grown was sealed to the chamber as in Fig. 1A. A Kevlar thread was attached to the exposed mature blade where it emerged from the ensheathing leaves, using double‐sided mounting tabs (Ace Hardware Corp., Illinois, USA). The thread was wrapped around a wheel on an optical incremental encoder (25G, Sequential Information Systems Inc., New York, USA) that recorded motion by rotating as the leaf elongated. An advantage of the encoder was its lack of sensitivity to temperature (Fig. 1B) or supply voltage, which allowed it to be used in the controlled environment chamber without correction. As the wheel turned, the encoder registered a count (pulse of light) for each increment of rotation. At 2 s intervals, the counts were entered in a datalogger (CR7, Campbell Scientific, Inc., Utah, USA) which stored them as a 1 min average of the total accumulated counts during the minute and converted the average to the elongation rate. The rate was downloaded to a computer for plotting. As a reference, the leaf base was clamped to a rigid bar (Fig. 1A). Toward the end of each light period, the pressure chamber was pressurized for 2–2.5 h. The seal around the mesocotyl in the chamber top was tightened and compressed air was supplied to the root system while leaf elongation was continuously monitored with the encoder. P was increased until guttation was observed at the margin of leaf 5 or until 1.5 MPa was reached. A controller (DP25‐E Process Meter, Omega Engineering Ingc, Stamford, Connecticut, USA) connected to a pressure transducer (PG856–250, Statham Laboratories Inc., Puerto Rico) maintained the P constant for the remainder of the P treatment. At the end of the pressurization, the shoot was excised for the determination of the Ψw profile while the root system was still under P. The profile was measured by first excising the shoot inside the controlled environment chamber and immediately transferring it to a humidity‐saturated glove box. Four 2 cm segments (total area 3–4 cm2) were excised from leaf 5 from the mature tip toward the growing base (Mc, Mb, Ma, and G in Fig. 1A) and placed in psychrometer cups coated with melted and re‐solidified petrolatum. Each cup was immediately attached to its heat sink, sealed, and placed in an isopiestic psychrometer system (Isopiestics Co., Delaware, USA). The Ψw were determined isopiestically (Boyer, 1966, 1995). Thereafter, the leaf tissues were frozen and thawed, and the osmotic potential (Ψs) was measured by the same technique. The Ψp was calculated from Ψw–Ψs. Before P was applied, the Ψw profile was determined on separate plants grown identically at the same time and sampled as above. Immediately after sampling leaf 5 as above in these control plants, their pots were moved to a humidity‐saturated room where the soil was sampled at a depth of 4 cm inside the pot (S in Fig. 1A). The soil sample covered the psychrometer cup at least 2 mm deep. The whole root/soil medium was then carefully detached from the pot. One root of 10 cm with attached branches was sampled after removing adhering soil and root tips (R in Fig. 1A). The Ψw of the soil and the mature root were determined isopiestically as above. The Ψs of the mature root also was determined and the Ψp was calculated from Ψw–Ψs. Soil water content The soil water contents were determined after removing the shoot and the whole root system by immediately measuring the fresh weight of the soil and then the dry weight after 3 d in an oven at 80 °C. Transpiration and exudation Transpiration was measured by weighing the whole apparatus while leaf 5 elongation was simultaneously monitored with the encoder in the controlled environment chamber where the plants had been grown (Fig. 1). The weight change was recorded every 10 s and stored in a computer. Measurements were summed in groups of six to give the weight each minute. The running average was then calculated for the previous 10 min. For comparison with the rate of transpiration, the rate of water delivery by the soil/root system was measured. The shoot was removed from plants grown under the same conditions and having an elongation rate similar to the intact plants used for growth and transpiration studies. The shoot was excised near the top of the mesocotyl, the cut mesocotyl surface was rinsed with water and blotted dry, P was applied, and the exudate was collected every minute for the first 5 min, then every 5–10 min thereafter. No exudation occurred without P. Conductance of the soil/root system Because the Ψw of the soil (S) was known from measurements with the psychrometer, and P was known, the conductance of the soil/root system was determined from the rate of exudation measured as above. The exudate first appeared on the cut mesocotyl surface when P balanced S (i.e. P=–S). In this balanced condition with a flat exudate meniscus at atmospheric pressure, the solution in the xylem had Ψw near zero because xylem solute concentrations were near zero (Tang and Boyer, 2002). As P was raised further, exudate moved out continuously driven by the force (P–(–S)). The conductance was then: where e was the exudation rate from the cut surface of the soil/root system after the shoot was removed (nm3 s–1) and k was the conductance of the soil/root system (nm3 s–1 MPa–1). Rewatering In a rewatering experiment, nutrient solution was added to the soil/root system by flooding the pressure chamber and draining the excess away through the inlet for compressed air. During the treatment, the elongation of leaf 5 was continuously monitored, and the mature tissue of the same leaf 5 was repeatedly sampled around Mb for isopiestic measurements of Ψw as above. Results P and profiles of Ψw In plants supplied with adequate water, leaf 5 elongated most rapidly early in the day, 2–4 h after the light came on. The leaf guttated at night and for about 20–30 min after the light turned on, but otherwise no guttation occurred during the day. If water was withheld for several days, elongation decreased, especially by the end of the light period, and the natural guttation at night became less and was eventually absent. When P was applied towards the end of the light period, guttation reappeared during the early days of the dehydration. For example, after 4 d of soil dehydration, leaf 5 of plants at 15 dap elongated at about 40% of the maximum rate (max) measured in comparable hydrated plants (Fig. 2A). The leaf Ψw was –0.7 to –0.9 MPa (Fig. 2B), and the xylem Ψw was –0.7 MPa at X. This was mostly a tension, because Tang and Boyer (2002) found Ψs was –0.04 MPa in xylem solution, which is small enough to neglect. A small growth‐induced Ψw of 0.13 MPa was present in the elongating region (X–G in shaded area, Fig. 2B). When P of 1.3–1.4 MPa was applied, guttation appeared and continued as long as P remained (Fig. 2A). The guttation indicated that the xylem solution was near atmospheric pressure and, because xylem Ψs also was near zero, X was near zero. Compared to X, the Ψw of Ma, Mb, and Mc rose to –0.2 to –0.4 MPa. The lack of near zero potential probably reflected the water potential difference between X and the mesophyll necessary to move water to the evaporating sites inside the mature parts of the leaf (Fig. 2B). In the elongating region, the (X–G) enlarged to 0.24 MPa (Fig. 2B). The (X–G) enlarged because X increased more than G. Leaf elongation recovered to the maximum rate in the hydrated controls (Fig. 2A), where it remained until the shoot was removed after 2 h to sample for Ψw shown in Fig. 2B. On the other hand, withholding water one more day (16 dap, Fig. 2C) caused leaf 5 to grow more slowly (15% of maximum rate in fully hydrated controls). The entire Ψw profile was lower than on the previous day (–0.8 to –1.0 MPa in the leaf, tension of –0.8 MPa at X), and the (X–G) was only 0.07 MPa (Fig. 2D). Applying P of 1.5 MPa (Fig. 2C) enlarged (X–G) somewhat (0.15 MPa, Fig. 2D) and increased the elongation rate initially, but did not cause guttation and did not maintain the elongation rate beyond the first hour (Fig. 2C). The Ψw of the exposed mature leaf blade was not increased by P (Fig. 2D). Over‐pressuring with P of 3 MPa failed to cause guttation or improve elongation (data not shown). After one more day (17 dap, Fig. 2E), leaf elongation had nearly ceased, the Ψw profile was even lower than on 16 dap (–1.2 to –1.3 MPa in the leaf, tension of 1.2 MPa at X), and the (X–G) was negligible (0.04 MPa, Fig. 2F). P of 1.5 MPa (or 3 MPa) enlarged (X–G) slightly (0.13 MPa, Fig. 2F) and led to a slight but short‐lived increase in elongation rate (Fig. 2E). No guttation appeared, and the Ψw of the exposed mature leaf was unchanged (Fig. 2F). P and profiles of Ψs and Ψp In the above experiment, the osmotic potential (Ψs) of leaf 5 was –1.1 to –1.2 MPa at 15 dap (Fig. 3A) and shifted to more negative levels as dehydration progressed (Fig. 3C, E). P generally did not change the Ψs (Fig. 3A, C, E) except at G when elongation recovered completely (Fig. 2A) and Ψs became considerably higher (Fig. 3A). The rise in Ψs probably resulted from water entering the elongating cells as the rate of elongation resumed, and also from solute depleted during the growth process. Figure 3B shows that the Ψp increased markedly in the mature region when P was applied at 15 dap, confirming that the mature tissues rehydrated when guttation occurred (Fig. 3B). However, Ψp scarcely changed in the elongating region. As dehydration became more severe on succeeding days (Fig. 3D, F), P affected Ψp only slightly. P and transpiration and root exudation The inability of Ψw and Ψp to respond to P in severely dehydrated plants (Figs 2D, F, 3D, F) indicated that the shoot failed to rehydrate fully. In order to investigate the causes, water loss and gain were measured in the shoot during P application. Water loss was determined from the transpiration rate (T=rate of weight loss of the entire apparatus in Fig. 1A in the growth environment). Water gain was determined from the rate of water delivery to the shoot by the soil/root system (e=rate of exudation by the mesocotyl after the shoot was removed from a plant grown separately but treated identically and subjected to the same P). Figure 4B shows that T was 1.2 nm3 s–1 before P and increased to about 1.7 nm3 s–1 after P at 15 dap (when elongation responded fully to P in Fig. 4A, as in Fig. 2A). There was an initial pulse of e when P was applied, and e always exceeded or equalled T (Fig. 4B). However, after one or two more days when Ψw had decreased further(see Fig. 2D, F), T was unaffected by P and the pulse of e matched T only briefly (Fig. 4D, F). After this initial pulse, e was less than T. The initial burst of elongation during P (Fig. 4C, E) coincided approximately with the pulse in e (Fig. 4D, F). However, the inability of e to account for T after the pulse indicates that the shoot was dehydrating, despite the application of P on the latter two days. Causes of low e The inability of P to move enough water to the shoot at 16 and 17 dap (Fig. 4D, F) was explored first by determining the total amount of water available to the plants. The soil displayed a gradual but typical decrease in Ψw as the water content decreased (Fig. 5). The soil Ψw was about –0.1 MPa at 11 dap and decreased to –0.7 MPa by the end of 17 dap (S in Fig. 5A). Leaf Ψw in the mature tissues (Mb in Fig. 5A) were always lower than soil Ψw. At 11 dap with the soil at its field capacity, the pot contained about 305 g of water that was drawn down to 28 g when the soil Ψw was at –1.5 MPa, i.e. the soil contained about 277 g of water available to the plant (Fig. 5B). By 17 dap, about 25 g of this water remained available to the plant (shaded area, Fig. 5B). The plants required only about 3 g of water to rehydrate (measured from the weight of water absorbed by the shoot when its cut base was in water for 12 h at 4 °C). Moreover, T consumed no more than 7 g during the 2 h application of P. Therefore, Fig. 5 indicates that sufficient water was in the soil to rehydrate the shoot fully and meet the demands of T throughout these experiments. Because there was adequate water in the soil, the conductance of the soil/root system may have prevented the water from entering the shoot. S became as low as –0.7 MPa at 17 dap (Fig. 5), and applying P of 0.7 MPa to the soil/root system should have removed all tensions normally present in the soil. The P actually applied was greater than –S, causing water to exude from the mesocotyl with a driving force (P–(–S)). From the average rate of exudation during P under these conditions, the soil/root conductance was determined to be 2.55 nm3 s–1 MPa–1 at 15 dap but only about 0.44 nm3 s–1 MPa–1 at 17 dap (Fig. 6). The decline in conductance of the soil/root system raises the possibility that the root vascular supply contained emboli blocking water uptake. However, Fig. 7A shows that elongation resumed immediately when water was added to the soil at 17 dap, indicating that the vascular system was highly conductive for water. Water uptake clearly occurred because the Ψw of –1.38 MPa rose to –0.82 MPa within 30 min at Mb and gradually increased afterwards to –0.51 MPa, which was similar to that in hydrated plants. The immediate recovery of growth was substantial (to about 0.5 µm s–1) but not complete (maximum rate was 1.2–1.3 µm s–1). Adding water to the soil after P caused a more gradual resumption of elongation than if P had not been applied (Fig. 7B). This shows that P itself probably caused some emboli to form, slowing the recovery when water was re‐supplied. However, the resumption of elongation and recovery to high Ψw were quite rapid nonetheless, indicating that this amount of embolism could not account for the decreased e. Adding water to the soil simultaneously with P caused a more complete recovery of leaf elongation and Ψw (Fig. 7C) than rewatering alone (Fig. 7A) or rewatering after P (Fig. 7B). The leaf guttated. However, elongation was not sustained and, after initially increasing to about 1.5 µm s–1, it decreased below the maximum rate. Discussion The (X–G) is a tissue‐level manifestation of the water potential field responsible for water uptake by enlarging cells during growth. When (X–G) becomes zero, growth cannot occur. Evidence for this central role of (X–G) comes from the pressurization of growing tissues (Boyer, 1968, 2001), maturation of growing tissues (Cavalieri and Boyer, 1982; Westgate and Boyer, 1984; Tang and Boyer, 2002), removal of auxin supply (Maruyama and Boyer, 1994), low temperature (Boyer, 1993), and decreased X caused by low soil water potentials (Tang and Boyer, 2002). Therefore, changes in (X–G) are important. The data show that (X–G) was small in all conditions where growth was inhibited by low Ψw in the soil. When P was applied at moderate levels of soil dehydration, (X–G) increased and the field of growth‐induced Ψw became steeper. The steeper field drove more water into the elongating cells, and their growth rate increased. P increased (X–G) because X rose more than G. Applying pressure to the roots caused Ψw in the xylem solution to increase more rapidly and to a larger extent than in the surrounding tissues of the elongating zone. The effect was probably caused by the high conductance of the xylem vessels and their rigidity and small volume compared with the surrounding tissues. With these properties, only a small amount of water entering the vessels would cause a large, rapidly transmitted increase in xylem potential. By contrast, a sheath of small, undifferentiated cells surrounds the xylem in the elongating zone, and Nonami et al. (1997) reported low diffusivities in these cells in soybean stems. There is evidence for a similar low conductance sheath in maize leaves (Tang and Boyer, 2002). As a consequence, X could change rapidly but radial movement out of the xylem would probably be limited. G in the surrounding cells would change less than in the xylem and, because the surrounding cells are large in number, G would reflect their properties and change less than X. It is noteworthy that P applied to roots had only a slight effect on turgor pressures in the growing leaf cells. While changes in turgor pressure undoubtedly cause growth rates to change, there are many instances where growth responds to other factors. For example, growth diminished in parallel with (X–G) when tissue matured, but turgor changed in opposition and increased substantially (Westgate and Boyer, 1984; Tang and Boyer, 2002). Similar parallel responses of (X–G), but opposing effects of turgor, were reported during auxin‐depletion (Maruyama and Boyer, 1994), low temperature treatment (Boyer, 1993), and exposure to low Ψw (Nonami et al., 1997; Tang and Boyer, 2002). In soybean seedlings exposed to low Ψw, turgor pressure did not decrease in most of the cells of the growing tissues, despite marked decreases in growth (Nonami and Boyer, 1989). Turgor was measured with four independent methods including direct cell measurements with a pressure probe. However, decreased turgor could be detected in the inner cells next to the xylem, suggesting that (X–G) had decreased (Nonami and Boyer, 1989). Consequently, changes in turgor pressure frequently cannot explain changes in growth rates, especially when significant (X–G) are present. In maize, the protoxylem runs right through the elongating region and into the exposed mature part of the leaf (Tang and Boyer, 2002). When P caused X to rise, the protoxylem fed water to the mature part. The sheath of undifferentiated cells is absent in the mature tissues (having differentiated into mature cells), and the mature metaxylem vessels directly contact the bundle sheath cells (Tang and Boyer, 2002). Water supplied by the xylem was quickly absorbed by the bundle sheath and mesophyll, and their Ψw (Ma, Mb, Mc) rose. Therefore, raising X with P played two roles. First, it enlarged (X–G). Second, it increased M. Both roles appeared essential if growth recovery was sustained after P was applied. The evidence for this dual role appears in the contrasting behaviour of the moderately and severely dehydrated plants. Leaf growth experienced a gradual decline when water was withheld from the soil. If the decline was only moderate, an application of P recovered growth fully. The field in (X–G) was steepened and water uptake by the soil/root system was more rapid than T in the mature tissues. The additional water hydrated the mature tissue, detected as guttation and increased Ψw and Ψp in the mature leaf tissues. Therefore, both roles of P were satisfied. By contrast, further dehydration prevented P from supplying enough water to compensate fully for T. Guttation did not occur and the mature tissues did not increase in Ψw or Ψp. The (X–G) improved only slightly, and leaf elongation improved briefly. The brief elongation probably resulted from the early P‐induced pulse of xylem water that could be detected in the detopped soil/root systems. Under these circumstances, the two roles of P were not satisfied, and elongation soon settled to the rate before P. P and root water absorption An important question is why severely dehydrated leaves did not rehydrate during P application when there was enough water in the soil to completely rehydrate the shoots and account for T in all of the experiments? There was a decrease in the conductance of the soil/root system, and a possible reason was a block by emboli in the root vascular system, caused by large tensions developing in the xylem (Boyer, 1971; Canny, 1997; Fuchs and Livingston, 1996; McCully, 1999; Neufeld et al., 1992; Tyree et al., 1986; Zwieniecki and Holbrook, 1998). However, the evidence argues against this explanation because emboli take time to repair. Leaf elongation improved immediately after rewatering, and Ψw rapidly rose to hydrated levels in the mature tissue. If P was applied before rewatering, the resumption in elongation was more gradual and suggested that some emboli could have been induced by the application of P. Salleo et al. (1992) reported emboli in plant organs exposed to pressurized air because some air entered the xylem. Despite this effect, elongation soon recovered to the rate observed without P application, and Ψw increased in the mature tissues. Therefore, P‐induced emboli were inadequate to account for the soil/root block. A similar conclusion applies when P was present during rewatering. Emboli could not have significantly blocked water transport, which resumed immediately, and leaf Ψw recovered completely indicating that the roots could support rapid water movement. The recovery of leaf elongation was not sustained probably because other growth‐limiting factors, perhaps metabolic, became limiting. These results suggest that the source of low conductance was in the soil. The flow path included both the soil and the root system acting in series. While the root system may have been slightly affected by P‐induced emboli, it is likely that the soil hydraulic conductivity also decreased when soil water contents decreased (for a brief review, see Kramer and Boyer, 1995). Lang and Gardner (1970) point out that these decreases in soil conductivity can compensate for increasing gradients in water potential between the root and soil, resulting in less rather than more water flow. This well‐known effect would be immediately reversed by rewatering. The reversal would allow flow to resume immediately through a functioning root system, as observed in the rewatering experiments. Therefore, the rewatering experiments suggest that the severely dehydrated soil limited flow to the root and prevented a sustained delivery of water to the shoot with the result that leaf elongation could not be sustained. With the soil conductance limiting in this way, the pulse of exudate observed with P in severely dehydrated plants (Fig. 4D, F) may have come mostly from the roots. Application of P to the roots would release cell water just as it does from leaves during standard measurements with a pressure chamber (Scholander et al., 1965). The volume of water released from the roots would be finite, and only a pulse of water would appear at the cut surface of the root system. The P‐induced release of root water to the xylem would shrink the roots, reduce contact with the soil and exacerbate the low soil conductance especially at the soil/root interface. Therefore, soil water may not have contributed much to the water appearing at the cut surface of the root system in the severely dehydrated plants. P and root signalling Passioura (1988) investigated the signals affecting leaf elongation in dehydrating soil by applying P to maintain a xylem meniscus at a cut in a wheat leaf and thus a high leaf hydration despite the drying soil. Leaf elongation eventually was inhibited and provided strong evidence for chemical signalling of the shoot by the roots (Davies and Zhang, 1991). The major difference between the Passioura experiment and the present one is the continuous application of P by Passioura (1988), but brief application of P in the present experiment. With continuous application of P, the leaf was continuously in a hydrated state that would not allow early hydraulic signalling to be seen. With brief application, the leaf would dehydrate between applications, and early hydraulic signalling could be detected. Because leaf elongation recovered completely in the early stage of the inhibition in the present work, the early signals appeared to be entirely hydraulic. However, as the depletion became more severe, metabolic or chemical signalling from the root may have become important because rewatering did not fully recover elongation. These findings are consistent with the root signalling reported by Passioura (1988). Nonami et al. (1997) also found hydraulic root/shoot signalling in the early stages of soil dehydration in soybean, but abscisic acid concentrations increased and were followed by metabolic changes after 1–2 d (Bensen et al., 1988; Nonami and Boyer, 1990a, b). It is likely that the effects of P were specific to plants that were not fully hydrated. Exposing plants to an adequate supply of water in humid, non‐transpiring conditions caused P to have little effect on the growth of maize leaves or soybean stems (Hsiao et al., 1998; Nonami et al., 1997). No change in leaf Ψw or transpiration was reported in pepper grown in –0.05 MPa nutrient medium when P was applied (Janes and Gee, 1973). Additional P applied to the root/soil system had little effect on the xylem pressure of maize when the xylem pressure already reached the atmospheric level and guttation could occur (Wei et al., 1999). By contrast, Passioura and Munns (2000) observed transient surges or pauses in leaf expansion when P was changed in plants in low humidity when transpiration was occurring, suggesting that xylem under tension responds to P perhaps by changing water delivery to the xylem and thus (X–G). In effect, changing the tensions might cause local changes in a few cells around the xylem that would rapidly block or permit water entry for the entire growing region of the leaf. The effects on growth would be equally rapid and similar to those in moderately dehydrated maize plants in the present work. Growth‐induced potential fields Additional insight comes from considerations at the cellular level. Because growth‐induced Ψw originate from the yielding of the cell walls (Boyer, 1968; 2001), Ψp and Ψw are kept below the maximum that would occur if the cells were mature. Plant growth regulators such as auxin establish the ability of the walls to yield (Cleland, 1971; Taiz, 1984; Davies and Zhang, 1991) and thus underlie the formation of these potentials (Ikeda et al., 1999; Maruyama and Boyer, 1994). When water is available to keep X high, the low Ψw in the surrounding cells creates (X–G). Water moves out of the xylem and into the cells, and growth results. When water is less available and X decreases, wall yielding must create lower G in order for growth to continue. Eventually wall yielding is insufficient to lower G, (X–G) becomes zero, and growth ceases. (X–G) measured in the psychrometer is a tissue‐level potential that is the manifestation of the field in growth‐induced Ψw at the cellular level (Nonami and Boyer, 1993). Maize leaves have a rather steep field in the elongating region because the distance between adjacent xylem vessels is small, of the order of 1 mm, and the small cells ensheathing the xylem appear to act as a cellular barrier to flow (Tang and Boyer, 2002). The tissues between veins develop a sufficiently low Ψw to move water through the barrier to meet the demand for growth (Fricke, 1997; Fricke et al., 1997; Fricke and Flowers, 1998; Martre et al., 1999; Nonami and Boyer, 1993; Tang and Boyer, 2002). The barrier to flow caused by the small cells ensheathing the xylem causes the rapid Ψw changes in the xylem to be transmitted only slowly through the growing tissues (Passioura and Boyer, 2003). The slow transmission of the Ψw field to outlying tissues might account for some time‐scale effects observed by Munns et al. (2000) on leaf growth. Growth would respond rapidly to changes in xylem Ψw but the Ψw field might require several hours to re‐establish fully during which rates would probably adjust (Nonami et al., 1997; Passioura and Boyer, 2003). Acknowledgements We thank John B Passioura and Mark E Westgate for helpful suggestions. This study was supported by DOE grant DE‐FG02‐87ER13776 to JSB. View largeDownload slide Fig. 1. (A) Apparatus for the root pressurization experiments in the controlled environment chamber where the plants were grown. A: Supporting frame. B: Pressure chamber assembly. C: Plexiglas pot with maize root growing in a soil mix. D: Reference bar to hold the leaf base in a fixed position. E: Encoder to record the change in leaf length. F: Maize leaf 5. A Kevlar thread, shown as the dashed line, was attached to the mature part of leaf 5 on one end with a double sided tape, looped around the encoder wheel, and counter‐balanced by a weight (6 g) on the other end. Leaf elongation was continuously monitored with the encoder. The root/soil system was pressurized with compressed air. The transpiration was recorded from the weight change of the whole apparatus sitting on a balance. Samples for Ψw were taken from the soil (S), mature root (R), elongating region of leaf 5 (G), and mature regions of leaf 5 (Ma, Mb, and Mc). (B) Temperature response of the apparatus in the controlled environment. Instead of a plant, a metal tube was sealed in the pressure chamber, and the Kevlar thread was attached to the tip. Upper trace shows the air temperature and lower trace shows the elongation rate. View largeDownload slide Fig. 1. (A) Apparatus for the root pressurization experiments in the controlled environment chamber where the plants were grown. A: Supporting frame. B: Pressure chamber assembly. C: Plexiglas pot with maize root growing in a soil mix. D: Reference bar to hold the leaf base in a fixed position. E: Encoder to record the change in leaf length. F: Maize leaf 5. A Kevlar thread, shown as the dashed line, was attached to the mature part of leaf 5 on one end with a double sided tape, looped around the encoder wheel, and counter‐balanced by a weight (6 g) on the other end. Leaf elongation was continuously monitored with the encoder. The root/soil system was pressurized with compressed air. The transpiration was recorded from the weight change of the whole apparatus sitting on a balance. Samples for Ψw were taken from the soil (S), mature root (R), elongating region of leaf 5 (G), and mature regions of leaf 5 (Ma, Mb, and Mc). (B) Temperature response of the apparatus in the controlled environment. Instead of a plant, a metal tube was sealed in the pressure chamber, and the Kevlar thread was attached to the tip. Upper trace shows the air temperature and lower trace shows the elongation rate. View largeDownload slide Fig. 2. Root pressurization (P), elongation of leaf 5, and the corresponding Ψw profiles when soil was dehydrated around maize roots. Water was withheld at 11 dap. (A, C, E) Rate of leaf elongation at 15, 16, and 17 dap. P is shown by the shaded area. Guttation was observed during P in (A) but not (C) or (E). The maximum rate (max) was observed 2–4 h after illumination in plants grown identically at the same time and supplied with water each day except for (E) at 17 dap, where max is for plants at 16 dap that were comparable in size. (B, D, F) Profiles of Ψw at 15, 16, and 17 dap for plants in (A), (C), and (E) compared with profiles in unpressurized plants. Open circles are profiles from separate plants grown identically at the same time. Data are means ±1 SD for three unpressurized plants (sometimes SD was smaller than data point). Closed circles are profiles after 2 h of P application to roots in (A), (C), and (E). No root or soil data are available for the latter plants because P was present during the time of sampling. For all profiles, Ψw were single determinations at each position in a single plant. Positions S, R, G, Ma, Mb, Mc refer to those in Fig. 1. Dashed vertical line indicates Ψw in xylem (X), mostly a tension. Growth‐induced Ψw is in cells outside the xylem and is shown by the shaded area. The leaf growing region extended between 0 cm and +10 cm from the leaf base. The experiment was repeated twice with similar results. View largeDownload slide Fig. 2. Root pressurization (P), elongation of leaf 5, and the corresponding Ψw profiles when soil was dehydrated around maize roots. Water was withheld at 11 dap. (A, C, E) Rate of leaf elongation at 15, 16, and 17 dap. P is shown by the shaded area. Guttation was observed during P in (A) but not (C) or (E). The maximum rate (max) was observed 2–4 h after illumination in plants grown identically at the same time and supplied with water each day except for (E) at 17 dap, where max is for plants at 16 dap that were comparable in size. (B, D, F) Profiles of Ψw at 15, 16, and 17 dap for plants in (A), (C), and (E) compared with profiles in unpressurized plants. Open circles are profiles from separate plants grown identically at the same time. Data are means ±1 SD for three unpressurized plants (sometimes SD was smaller than data point). Closed circles are profiles after 2 h of P application to roots in (A), (C), and (E). No root or soil data are available for the latter plants because P was present during the time of sampling. For all profiles, Ψw were single determinations at each position in a single plant. Positions S, R, G, Ma, Mb, Mc refer to those in Fig. 1. Dashed vertical line indicates Ψw in xylem (X), mostly a tension. Growth‐induced Ψw is in cells outside the xylem and is shown by the shaded area. The leaf growing region extended between 0 cm and +10 cm from the leaf base. The experiment was repeated twice with similar results. View largeDownload slide Fig. 3. Osmotic potentials (A, C, E) and turgor pressures (B, D, F) in the plants of Fig. 2. Closed symbols are profiles after 2 h of P. Open symbols are profiles from separate plants grown identically at the same time but not pressurized. Data are means ±1 SD for three unpressurized plants. Sometimes SD was smaller than data point. View largeDownload slide Fig. 3. Osmotic potentials (A, C, E) and turgor pressures (B, D, F) in the plants of Fig. 2. Closed symbols are profiles after 2 h of P. Open symbols are profiles from separate plants grown identically at the same time but not pressurized. Data are means ±1 SD for three unpressurized plants. Sometimes SD was smaller than data point. View largeDownload slide Fig. 4. Elongation of leaf 5, and the corresponding transpiration and water delivery to the shoot when maize was subjected to dehydrated soil and P as in Figs 2 and 3. (A, C, E) Rate of leaf elongation at 15, 16, and 17 dap, with P shown by the shaded area. Guttation was observed during P in (A) but not (C) or (E). (B, D, F) Transpiration (solid line) and water delivery to the shoot (shaded area e) at 15, 16, and 17 dap. Water delivery was measured as exudation at the cut surface of the mesocotyl with the shoot removed at the beginning of P. The experiment was repeated once. View largeDownload slide Fig. 4. Elongation of leaf 5, and the corresponding transpiration and water delivery to the shoot when maize was subjected to dehydrated soil and P as in Figs 2 and 3. (A, C, E) Rate of leaf elongation at 15, 16, and 17 dap, with P shown by the shaded area. Guttation was observed during P in (A) but not (C) or (E). (B, D, F) Transpiration (solid line) and water delivery to the shoot (shaded area e) at 15, 16, and 17 dap. Water delivery was measured as exudation at the cut surface of the mesocotyl with the shoot removed at the beginning of P. The experiment was repeated once. View largeDownload slide Fig. 5. Ψw of soil (S) and mature leaf (Mb) at various soil water contents in these experiments. (A) Soil water content as a percentage of soil dry weight. (B) Total water content of soil in pot. Upward arrows show dap. Shaded area shows extractable water in the soil at 17 dap. Each data pair (S and Mb) is from the soil and plant in a single pot. View largeDownload slide Fig. 5. Ψw of soil (S) and mature leaf (Mb) at various soil water contents in these experiments. (A) Soil water content as a percentage of soil dry weight. (B) Total water content of soil in pot. Upward arrows show dap. Shaded area shows extractable water in the soil at 17 dap. Each data pair (S and Mb) is from the soil and plant in a single pot. View largeDownload slide Fig. 6. Soil/root conductances for water at various times when water was withheld from maize at 11 dap. Conductances were measured according to Eq. 1 in the soil/root system after the shoot was removed. e was averaged for 2 h while P was applied. Data are means ±1 SD for two or three soil/root systems. SD generally was smaller than the symbols. View largeDownload slide Fig. 6. Soil/root conductances for water at various times when water was withheld from maize at 11 dap. Conductances were measured according to Eq. 1 in the soil/root system after the shoot was removed. e was averaged for 2 h while P was applied. Data are means ±1 SD for two or three soil/root systems. SD generally was smaller than the symbols. View largeDownload slide Fig. 7. Recovery of elongation of leaf 5 upon rewatering the soil at 17 dap; (A) without P, (B) after P of 1.5 MPa for 2 h (shaded area), (C) during P for 2 h. Maximum rate (max) in (C) was measured at 16 dap in hydrated controls that were comparable in size. Soil Ψw was –0.70 MPa before rewatering and –0.13 MPa after rewatering. Numbers alongside trace show leaf Ψw measured in the same plants around Mb, as in Fig. 1. Data are typical for the experiment that was repeated four times. View largeDownload slide Fig. 7. Recovery of elongation of leaf 5 upon rewatering the soil at 17 dap; (A) without P, (B) after P of 1.5 MPa for 2 h (shaded area), (C) during P for 2 h. Maximum rate (max) in (C) was measured at 16 dap in hydrated controls that were comparable in size. Soil Ψw was –0.70 MPa before rewatering and –0.13 MPa after rewatering. Numbers alongside trace show leaf Ψw measured in the same plants around Mb, as in Fig. 1. Data are typical for the experiment that was repeated four times. References BensenRJ, Boyer JS, Mullet JE. 1988. 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Google Scholar Society for Experimental Biology
TI - Root pressurization affects growth‐induced water potentials and growth in dehydrated maize leaves
JF - Journal of Experimental Botany
DO - 10.1093/jxb/erg265
DA - 2003-11-01
UR - https://www.deepdyve.com/lp/oxford-university-press/root-pressurization-affects-growth-induced-water-potentials-and-growth-efgP0m4yud
SP - 2479
EP - 2488
VL - 54
IS - 392
DP - DeepDyve
ER -