Wall extensibility and cell hydraulic conductivity decrease in enlarging stem tissues at low water potentials

Measurements with a guillotine psychrometer (H Nonami, JS Boyer [1990] Plant Physiol 94: 1601-1609) indicate that the inhibition of stem growth at low water potentials (low ψ_w) is accompanied by decreases in cell wall extensibility and tissue hydraulic conductance to water that eventually limit growth rate in soybean (Glycine max L. Merr.). To check this conclusion, we measured cell wall properties and cell hydraulic conductivities with independent techniques in soybean seedlings grown and treated the same way, i.e. grown in the dark and exposed to low ψ_w by transplanting dark grown seedlings to vermiculite of low water content. Wall properties were measured with an extensiometer modified for intact plants, and conductances were measured with a cell pressure probe in intact plants. Theory was developed to relate the wall measurements to those with the psychrometer. In the elongation zone, the plastic deformability of the walls decreased when measured with the extensiometer while growth was inhibited at low ψ_w. It increased during a modest growth recovery. This behavior was the same as that for the wall extensibility observed previously with the psychrometer. Tissue that was killed before measurement with the extensiometer also showed a similar response, indicating that changes in wall extensibility represented changes in wall physical properties and not rates of wall biosynthesis. The elastic compliance (reciprocal of bulk elastic modulus) did not change in the elongating or mature tissue. The hydraulic conductivity of cortical cells decreased in the elongating tissue and increased slightly during growth recovery in a response similar to that observed with the psychrometer. We conclude that the plastic properties of the cell walls and the conductance of the cells to water were decreased at low ψ_w but that the elastic properties of the walls were of little consequence in this response.     

Effect of inhibition of abscisic Acid accumulation on the spatial distribution of elongation in the primary root and mesocotyl of maize at low water potentials

Previous work showed that accumulation of endogenous abscisic acid (ABA) acts both to maintain primary root growth and inhibit shoot growth in maize seedlings at low water potentials (ψ_w) (IN Saab, RE Sharp, J Pritchard, GS Voetberg [1990] Plant Physiol 93: 1329-1336). In this study, we have characterized the growth responses of the primary root and mesocotyl of maize (Zea mays L. cv FR27 × FRMo 17) to manipulation of ABA levels at low ψ_w with a high degree of spatial resolution to provide the basis for studies of the mechanism(s) of ABA action. In seedlings growing at low ψ_w and treated with fluridone to inhibit carotenoid (and ABA) biosynthesis, ABA levels were decreased in all locations of the root and mesocotyl growing zones compared with untreated seedlings growing at the same ψ_w. In the root, low ψ_w (−1.6 megapascals) caused a shortening of the growing zone, as reported previously. The fluridone treatment was associated with severe inhibition of root elongation rate, which resulted from further shortening of the growing zone. In the mesocotyl, low ψ_w (−0.3 megapascal) also resulted in a shortened growing zone. In contrast with the primary root, however, fluridone treatment prevented most of the inhibition of elongation and the shortening of the growing zone. Final cell length measurements indicated that the responses of both root and mesocotyl elongation to ABA manipulation at low ψ_w involve large effects on cell expansion. Measurements of the relative changes in root and shoot water contents and dry weights after transplanting to a ψ_w of −0.3 megapascal showed that the maintenance of shoot elongation in fluridone-treated seedlings was not attributable to increased water or seed-reserve availability resulting from inhibition of root growth. The results suggest a developmental gradient in tissue responsiveness to endogenous ABA in both the root and mesocotyl growing zones. In the root, the capacity for ABA to protect cell expansion at low ψ_w appears to decrease with increasing distance from the apex. In the mesocotyl, in contrast, the accumulation of ABA at low ψ_w appears to become increasingly inhibitory to expansion as cells are displaced away from the meristematic region.     

Transpiration- and growth-induced water potentials in maize

Recent evidence from leaves and stems indicates that gradients in water potential (ψ_w) necessary for water movement through growing tissues are larger than previously assumed. Because growth is sensitive to tissue ψ_w and the behavior of these gradients has not been investigated in transpiring plants, we examined the water status of all the growing and mature vegetative tissues of maize (Zea mays L.) during high and low rates of transpiration. The ψ_w measured in the mature regions of the plant responded primarily to transpiration, while the ψ_w in the growing regions was affected both by transpiration and growth. The transpiration-induced potentials of the mature tissue formed a gradient of decreasing ψ_w along the transpiration stream while the growth-induced potentials formed a gradient of decreasing ψ_w from the transpiration stream to the expanding cells in the growing tissue. The growth-induced gradient in ψ_w within the leaf remained fairly constant as the xylem ψ_w decreased during the day and was associated with a decreased osmotic potential (ψ_s) of the growing region (osmotic adjustment). The growth-induced gradient in ψ_w was not caused by excision of the tissue because intact maize stems exhibited a similar ψ_w. These observations support the concept that large gradients in ψ_w are required to maintain water flow to expanding cells within all the vegetative tissues and suggest that the maintenance of a favorable gradient in ψ_w for cell enlargement may be an important role for osmotic adjustment.     

Osmotic adjustment, symplast volume, and nonstomatally mediated water stress inhibition of photosynthesis in wheat

At low water potential (ψ_w), dehydration reduces the symplast volume of leaf tissue. The effect of this reduction on photosynthetic capacity was investigated. The influence of osmotic adjustment on this relationship was also examined. To examine these relationships, comparative studies were undertaken on two wheat cultivars, one that osmotically adjusts in response to water deficits (`Condor'), and one that lacks this capacity (`Capelle Desprez'). During a 9-day stress cycle, when water was withheld from plants grown in a growth chamber, the relative water content of leaves declined by 30% in both cultivars. Leaf osmotic potential (ψ_s) declined to a greater degree in Condor plants. Measuring ψ_s at full turgor indicated that osmotic adjustment occurred in stressed Condor, but not in Capelle plants. Two methods were used to examine the degree of symplast (i.e. protoplast) volume reduction in tissue rapidly equilibrated to increasingly low ψ_w. Both techniques gave similar results. With well-watered plants, symplast volume reduction from the maximum (found at high ψ_w for each cultivar) was the same for Condor and Capelle. After a stress cycle, volume was maintained to a greater degree at low ψ_w in Condor leaf tissue than in Capelle. Nonstomatally controlled photosynthesis was inhibited to the same degree at low ψ_w in leaf tissue prepared from well-watered Condor and Capelle plants. However, photosynthetic capacity was maintained to a greater degree at low ψ_w in tissue prepared from stressed Condor plants than in tissue from stressed Capelle plants. Net CO₂ uptake in attached leaves was monitored using an infrared gas analyzer. These studies indicated that in water stressed plants, photosynthesis was 106.5% higher in Condor than Capelle at ambient [CO₂] and 21.8% higher at elevated external [CO₂]. The results presented in this report were interpreted as consistent with the hypothesis that there is a causal association between protoplast (and presumably chloroplast) volume reduction at low ψ_w and low ψ_w inhibition of photosynthesis. Also, the data indicate that osmotic adjustment allows for maintenance of relatively greater volume at low ψ_w, thus reducing low ψ_w inhibition of chloroplast photosynthetic potential.     

Proline accumulation in maize (Zea mays L.) primary roots at low water potentials. II. Metabolic source of increased proline deposition in the elongation zone

The proline (Pro) concentration increases greatly in the growing region of maize (Zea mays L.) primary roots at low water potentials (ψ_w), largely as a result of an increased net rate of Pro deposition. Labeled glutamate (Glu), ornithine (Orn), or Pro was supplied specifically to the root tip of intact seedlings in solution culture at high and low ψ_w to assess the relative importance of Pro synthesis, catabolism, utilization, and transport in root-tip Pro deposition. Labeling with [³H]Glu indicated that Pro synthesis from Glu did not increase substantially at low ψ_w and accounted for only a small fraction of the Pro deposition. Labeling with [¹⁴C]Orn showed that Pro synthesis from Orn also could not be a substantial contributor to Pro deposition. Labeling with [³H]Pro indicated that neither Pro catabolism nor utilization in the root tip was decreased at low ψ_w. Pro catabolism occurred at least as rapidly as Pro synthesis from Glu. There was, however, an increase in Pro uptake at low ψ_w, which suggests increased Pro transport. Taken together, the data indicate that increased transport of Pro to the root tip serves as the source of low-ψ_w-induced Pro accumulation. The possible significance of Pro catabolism in sustaining root growth at low ψ_w is also discussed.     

Leaf magnesium alters photosynthetic response to low water potentials in sunflower

We grew sunflower (Helianthus annuus L.) plants in nutrient solutions having nutritionally adequate but low or high Mg²⁺ concentrations and determined whether photosynthesis was effected as leaf water potentials (ψ_w) decreased. Leaf Mg contents were 3- to 4-fold higher in the plants grown in high Mg²⁺ concentrations (10 millimolar) than in those grown in low concentrations (0.25 millimolar). These contents were sufficient to support maximum growth, plant dry weight, and photosynthesis, and the plants appeared normal. As low ψ_w developed, photosynthesis was inhibited but moreso in high Mg leaves than in low Mg leaves. The effect was particularly apparent under conditions of light- and CO₂-saturation, indicating that the chloroplast capacity to fix CO₂ was altered. The differential inhibition observed in leaves of differing Mg contents was not observed in leaves having differing K contents, suggesting that the effect may have been specific for Mg. Because Mg²⁺ inhibits photophosphorylation and coupling factor activities at concentrations likely to occur as leaves dehydrate, Mg may play a role in the inhibition of chloroplast reactions at low ψ_w, especially in leaves such as sunflower that markedly decrease in water content as ψ_w decreases.     

Internal CO(2) Measured Directly in Leaves : Abscisic Acid and Low Leaf Water Potential Cause Opposing Effects

Observations of nonuniform photosynthesis across leaves cast doubt on internal CO₂ partial pressures (p_i) calculated on the assumption of uniformity and can lead to incorrect conclusions about the stomatal control of photosynthesis. The problem can be avoided by measuring p_i directly because the assumptions of uniformity are not necessary. We therefore developed a method that allowed p_i to be measured continuously in situ for days at a time under growth conditions and used it to investigate intact leaves of sunflower (Helianthus annuus L.), soybean (Glycine max L. Merr.), and bush bean (Phaseolus vulgaris L.) subjected to high or low leaf water potentials (ψ_w) or high concentrations of abscisic acid (ABA). The leaves maintained a relatively constant differential (Δp) between ambient CO₂ and measured p_i throughout the light period when water was supplied. When water was withheld, ψ_w decreased and the stomata began to close, but measured p_i increased until the leaf reached a ψ_w of −1.76 (bush bean), −2.12 (sunflower) or −3.10 (soybean) megapascals, at which point Δp = 0. The increasing p_i indicated that stomata did not inhibit CO₂ uptake and a Δp of zero indicated that CO₂ uptake became zero despite the high availability of CO₂ inside the leaf. In contrast, when sunflower leaves at high ψ_w were treated with ABA, p_i did not increase and instead decreased rapidly and steadily for up to 8 hours even as ψ_w increased, as expected if ABA treatment primarily affected stomatal conductance. The accumulating CO₂ at low ψ_w and contrasting response to ABA indicates that photosynthetic biochemistry limited photosynthesis at low ψ_w but not at high ABA.     

Effect of water deficits on seed development in soybean : I. Tissue water status

Water deficits during seed filling often decrease seed size in soybean (Glycine max L.). The physiological basis for this response is not known but may result from direct effects of low seed water potential (Ψ_w) on the seed filling process. To determine whether low Ψ_w occurred in reproductive tissues of soybean, we monitored the water status (Ψ_w, Ψ_s, and Ψ_p) of leaf, pericarp, and seed (embryo and testa) tissue of greenhouse-grown plants subjected to a brief water deficit during the linear period of seed growth. Water deficits were imposed by withholding water and monitored in the reproductive tissues by thermocouple psychrometry. When water was abundant, leaf, pericarp, and seed Ψ_w were −0.5 to −0.7 megapascal at midday. When water was withheld, leaf Ψ_w decreased to −2.3 megapascals within 6 days. Pericarp Ψ_w also decreased to −1.9 megapascal during this time. Pericarp Ψ_s followed the decline in Ψ_w, but osmotic adjustment was not evident as the pericarp lost turgor completely by day 6. However, seed Ψ_w, Ψ_s, and Ψ_p were not significantly different from the controls. These results indicate that the water status of the developing seeds of soybean is not altered by short-term water deficits severe enough to inhibit the metabolic activity of the maternal plant. Maintenance of a favorable water status may be important for the conservation of seed growth rate exhibited by soybean under dry conditions.     

Polysomes, Messenger RNA, and Growth in Soybean Stems during Development and Water Deficit

The polysome status and populations of polysomal mRNA were examined in different regions of dark-grown soybean (Glycine max [L.] Merr.) stems that contained either dividing, elongating, or mature (nongrowing) cells. There was a developmental gradient of polysome content in which the dividing tissue had the highest levels and the mature tissue the lowest. A few hours after transplanting the seedlings to vermiculite having low water content (water potential Ψ_w = −0.29 megapascals), stem growth rate decreased to 30% of well-watered controls and the polysome content decreased most in the dividing and elongating tissues. After 24 to 36 hours, stem growth and polysome content recovered gradually. In vitro translation products of polysomal mRNA from dividing, elongating or mature tissue were examined on two-dimensional gels. In well-watered controls, each of the stem regions was enriched in a small subset of the polysomal mRNA population, probably because of developmentally regulated gene expression. Exposing plants to low Ψ_w for 24 hours induced a change in the relative abundance of a small number of polysomal mRNAs in the elongating and mature tissues, but not in the dividing tissue. After 24 to 72 hours at low Ψ_w, the changes in polysomal mRNA population were reversed in the elongating tissue. The data indicate that changes in stem growth at low water potential are associated with changes in polysome status and polysomal mRNA in the elongating tissue.     

Correlation between the Maintenance of Photosynthesis and in Situ Protoplast Volume at Low Water Potentials in Droughted Wheat

Studies were undertaken to examine the relationship between water deficit effects on photosynthesis and the extent of protoplast volume reduction which occurs in leaves at low water potential (Ψ_w). This relationship was monitored in two cultivars (`Condor' and `Capelle Desprez') of cultivated wheat (Triticum aestivum) that differed in sensitivity to drought, and in a wild relative of cultivated wheat (Triticum kotschyi) that has been previously found to be `drought resistant.' When subjected to periods of water stress, Condor and T. kotschyi plants underwent osmotic adjustment; Capelle plants did not. Photosynthetic capacity was maintained to different extents in the three genotypes as leaf Ψ_w declined during stress; Capelle plants were most severely affected. Calculations of internal leaf [CO₂] and stomatal conductance from gas exchange measurements indicated that differences in photosynthetic inhibition at low Ψ_w among the genotypes were primarily due to nonstomatal effects. The extent of protoplast volume reduction that occurred in leaves at low Ψ_w was also found to be different in the three genotypes; maintenance of protoplast volume and photosynthetic capacity in stressed plants of the genotypes appeared to be correlated. When the extent of water stress-induced inhibition of photosynthesis was plotted as a function of declining protoplast volume, this relationship appeared identical for the three genotypes. It was concluded that there is a correlative association between protoplast volume and photosynthetic capacity in leaves of wheat plants subjected to periods of water stress.     

Water deficits and reproduction in maize : response of the reproductive tissue to water deficits at anthesis and mid-grain fill

Reproductive development in maize (Zea mays L.) is vulnerable to plant water deficits during anthesis but becomes less sensitive as reproduction progresses. To determine whether changes in tissue water status correlated with the change in sensitivity, we examined the water potential (Ψ_w), osmotic potential (Ψ_s), and turgor of reproductive tissues during a short-term water deficit imposed at anthesis or mid-grain fill. Plants were grown in controlled environments in soil. At anthesis, leaf, husk, silk, and ovary Ψ_w of control plants was similar (−0.5 to −0.65 megapascal) at midday. When water was withheld, Ψ_w decreased to −1.75, −1.3, −1.2, and −1.0 megapascal in these tissues. Net water uptake by the ovaries was inhibited, but final dry weight, solute content, and total extractable carbohydrates were similar to the controls. At mid-grain fill, leaf, husk, grain, and embryo Ψ_w of control plants were −0.55, −0.35, −0.75, and −0.80 megapascal at midday. When water was withheld, leaf and husk Ψ_w decreased to −2.4 and −1.4 megapascal within 6 days. However, grain and embryo Ψ_w remained within 0.15 megapascal of control values. The grain continued to accumulate dry matter despite a net loss of water and a reduction in total solute content. These results indicate that the response of the reproductive tissues to plant water deficits varies with stage of grain development. The maintenance of a favorable water status only after grain filling is under way may explain, at least in part, the high sensitivity to plant water deficits early in reproductive development and the decrease in sensitivity as reproduction progresses.     

Root Growth and Oxygen Relations at Low Water Potentials. Impact of Oxygen Availability in Polyethylene Glycol Solutions

Polyethylene glycol (PEG), which is often used to impose low water potentials (ψ_w) in solution culture, decreases O₂ movement by increasing solution viscosity. We investigated whether this property causes O₂ deficiency that affects the elongation or metabolism of maize (Zea mays L.) primary roots. Seedlings grown in vigorously aerated PEG solutions at ambient solution O₂ partial pressure (pO₂) had decreased steady-state root elongation rates, increased root-tip alanine concentrations, and decreased root-tip proline concentrations relative to seedlings grown in PEG solutions of above-ambient pO₂ (alanine and proline accumulation are responses to hypoxia and low ψ_w, respectively). Measurements of root pO₂ were made using an O₂ microsensor to ensure that increased solution pO₂ did not increase root pO₂ above physiological levels. In oxygenated PEG solutions that gave maximal root elongation rates, root pO₂ was similar to or less than (depending on depth in the tissue) pO₂ of roots growing in vermiculite at the same ψ_w. Even without PEG, high solution pO₂ was necessary to raise root pO₂ to the levels found in vermiculite-grown roots. Vermiculite was used for comparison because it has large air spaces that allow free movement of O₂ to the root surface. The results show that supplemental oxygenation is required to avoid hypoxia in PEG solutions. Also, the data suggest that the O₂ demand of the root elongation zone may be greater at low relative to high ψ_w, compounding the effect of PEG on O₂ supply. Under O₂-sufficient conditions root elongation was substantially less sensitive to the low ψ_w imposed by PEG than that imposed by dry vermiculite.     

Identification of Low Molecular Mass GTP-Binding Proteins in Membranes of the Halotolerant Alga Dunaliella salina

A family of specific guanine nucleotide-binding proteins in Dunaliella salina was studied. Polypeptides of different subcellular fractions were separated by electrophoresis and transferred to nitrocellulose or Immobilon membranes. Incubation of the transfer blots with [³⁵S]GTPγS or [α-³²P]GTP showed no evidence for GTP-binding proteins in the chloroplast and cytosol fractions. However, two GTP-binding proteins with molecular masses of 28 and 30 kilodaltons were present in the plasma membrane and microsomal fractions. An additional 29 kilodalton GTP-binding protein was detected in the plasma membrane. The mitochondrial fraction contained significant amounts of only the 28 kilodalton GTP-binding protein. Binding of [³²P]GTP to the protein blots was completely prevented by 10 micromolar GTP or guanosine 5′-O-(2-thiodiphosphate) (added in 3 × 10⁴-fold excess), whereas ATP or CTP had no effect on the binding. The 28 kilodalton GTP-binding protein was recognized by polyclonal antibodies to the ras-related YPT1 protein of yeast but not by the anti-ras Y13-259 monoclonal antibody. GTP-binding proteins present in the microsomal fraction could not be solubilized by incubation of microsomes with 1 molar NaCl or 0.2 molar Na₂CO₃, but some GTP-binding activity was solubilized when microsomes were treated with 6 molar urea. These results indicate that D. salina GTP-binding proteins are tightly associated with the membranes. The covalent attachment of fatty acids to these proteins was also investigated. Electrophoresis followed by fluorography of delipidated microsomal proteins extracted from [³H]myristic acid-labeled cells showed an intense labeling of a 28 kilodalton protein. We conclude that D. salina contains proteins resembling the ras-related proteins found in animal cells and higher plants.     

Estimation of osmotic parameters accompanying zeatin-induced growth of detached cucumber cotyledons

Water potential (ψ), the osmotic potential (ψ_π), and the pressure potential (ψ_p) of detached cotyledons isolated from Cucumis sativus L. cv Marketer seedlings after 0, 1.5, and 3 days growth with and without zeatin were determined. From zero time to 3 days, cotyledons incubated without exogenous zeatin exhibited a slight decrease in ψ (from −0.4 to −1.0 bars), while those grown with zeatin developed even more negative values (about −4 bars). Both groups showed rising ψ_π values (decreases in solutes per unit volume), but this rise was more dramatic in those treated with zeatin. These data indicate that the capacity of zeatin-treated cotyledons to take up water more rapidly than controls and thus expand faster must be due to wall loosening, as reflected in ψ_p values which declined during 3 days from about +11 bars to about +1.4 bars.

It was also found that freshly detached cotyledons or those grown without exogenous zeatin exhibited osmoregulation in polyethylene glycol (PEG) solutions. That is, while cotyledons initially lost H₂O into certain PEG solutions, their ψ values decreased over time and they began absorbing water after 1 to 4 hours. After 3 days growth, zeatin-treated cotyledons had lost most of this capacity of osmoregulate. It seems likely that osmoregulation in cotyledons not treated with zeatin is due to wall loosening rather than changes in ψ_π. Zeatin-treated cotyledons with already loosened walls may not have this option to deal with water stress and thus simply come to equilibrium with external PEG solutions.

     

Detection of endogenous gibberellins and their relationship to hypocotyl elongation in soybean seedlings

Four gibberellins, GA₅₃, GA₁₉, GA₂₀, and GA₁, were detected by bioassay, chromatography in two HPLC systems, and combined gas chromatography-mass spectroscopy-selected ion monitoring (GC-MS-SIM) in etiolated soybean (Glycine max [L.] Merr.) hypocotyls. GC-MS-SIM employed [²H₂]-labeled standards for each endogenous gibberellin detected, and quantities estimated from bioassays and GC-MS-SIM were similar. This result plus the tentative detection of GA₄₄ and GA₈ (standards not available) indicates that the early-C-13-hydroxylation pathway for gibberellin biosynthesis predominates in soybean hypocotyls. Other gibberellins were not detected. Growth rates decreased after transfer to low water potential (ψ_w) vermiculite and were completely arrested 24 hours after transfer. The GA₁ content in the elongating region of hypocotyls had declined to 38% of the 0 time value at 24 hours after transfer to low ψ_w vermiculite, a level which was only 13% of the GA₁ content in control seedlings at the same time (24 hours posttransfer). Rewatering seedlings following 24 hours growth in low ψ_w vermiculite resulted in a complete recovery in elongation rate, an increase in GA₁ (20% at 2 hours, two-fold at 8 hours, eightfold at 24 hours), and a decrease in ABA levels (tenfold at 2 hours). Treatment of well-watered seedlings with the GA-synthesis inhibitor tetcyclacis (TCY) resulted in lowered GA₁ levels and increased ABA levels. When seedlings grown 24 hours in low ψ_w vermiculite were rewatered with TCY, recovery of the elongation rate was delayed and reduced, and the decline in ABA levels was slowed. Addition of GA₃ restored the elongation rate inhibited by TCY. Seedlings were growth responsive to exogenous GA₃, and this GA₃-promoted growth was inhibited by exogenous ABA. The data are consistent with the hypothesis that changes in GA₁ and ABA levels play a role in adjusting hypocotyl elongation rates. However, the changes observed are not of sufficient magnitude nor do they occur rapidly enough to suggest they are the primary regulators of elongation rate responses to rapidly changing plant water status.     

Plant factors controlling seed set in maize : the influence of silk, pollen, and ear-leaf water status and tassel heat treatment at pollination

In maize (Zea mays L.) large decreases in kernel number result when low water potentials (Ψ_w) and high temperatures occur during pollination. To gain insight into the basis for the decreased seed set, silk, pollen, and ear-leaf Ψ_w, the capability for silk osmotic adjustment, and pollen appearance were measured to determine their relationship to seed set. A multiple-eared or prolific (high carbohydrate availability to the pistillate inflorescence) hybrid (B73 × FR25), a heat sensitive hybrid (WF9 × A632), and a commercial hybrid (B73 × Mo17) were studied. A cross-pollination experiment, with pollination limited by pollen amount, was conducted to determine the impact on seed set of water and heat stressing the tassel and water stressing the ear. At low Ψ_w, silk Ψ_w and seed set were decreased whereas pollen Ψ_w, appearance, and viability were unaffected. High temperature resulted in a 2 megapascal decrease in pollen Ψ_w, visually damaged pollen being shed, decreased pollen viability, and, in two of the hybrids, substantially decreased pollen shed. Prolificacy did not result in increased silk solute accumulation but did result in superior seed production by the pistillate inflorescence at low Ψ_w. The magnitude of the decrease in silk solute potential was small (0.2 megapascal) and similar for all genotypes. One hybrid maintained a relatively high silk turgor but this hybrid also decreased the most in seed production when the pistillate inflorescence was water deficient. These results indicated an adverse effect of high temperature on pollen development, a positive relationship between seed production and silk water status, and no advantage to high silk turgor after silk emergence in maintaining seed production. Additionally, there was no evidence of variation in silk solute regulation capability among hybrids which varied in prolificacy, a trait important in drought tolerance, but the seed production of the pistillate inflorescence of the prolific hybrid was least affected by water deficit.     

Water Relations of Seed Development and Germination in Muskmelon (Cucumis melo L.) : V. Water Relations of Imbibition and Germination

The initiation of radicle growth during seed germination may be driven by solute accumulation and increased turgor pressure, by cell wall relaxation, or by weakening of tissues surrounding the embryo. To investigate these possibilities, imbibition kinetics, water contents, and water (Ψ) and solute (ψ_s) potentials of intact muskmelon (Cucumis melo L.) seeds, decoated seeds (testa removed, but a thin perisperm/endosperm envelope remains around the embryo), and isolated cotyledons and embryonic axes were measured. Cotyledons and embryonic axes excised and imbibed as isolated tissues attained water contents 25 and 50% greater, respectively, than the same tissues hydrated within intact seeds. The effect of the testa and perisperm on embryo water content was due to mechanical restriction of embryo swelling and not to impermeability to water. The Ψ and ψ_s of embryo tissues were measured by psychrometry after excision from imbibed intact seeds. For intact or decoated seeds and excised cotyledons, Ψ values were >−0.2 MPa just prior to radicle emergence. The Ψ of excised embryonic axes, however, averaged only −0.6 MPa over the same period. The embryonic axis apparently is mechanically constrained within the testa/perisperm, increasing its total pressure potential until axis Ψ is in equilibrium with cotyledon Ψ, but reducing its water content and resulting in a low Ψ when the constraint is removed. There was no evidence of decreasing ψ_s or increasing turgor pressure (Ψ-ψ_s) prior to radicle growth for either intact seeds or excised tissues. Given the low relative water content of the axes within intact seeds, cell wall relaxation would be ineffective in creating a Ψ gradient for water uptake. Rather, axis growth may be initiated by weakening of the perisperm, thus releasing the external pressure and creating a Ψ gradient for water uptake into the axis. The perisperm envelope contains a cap of small, thin-walled endosperm cells adjacent to the radicle tip. We hypothesize that weakening or separation of cells in this region could initiate radicle expansion.     

Protein Synthesis in Bromegrass (Bromus inermis Leyss) Cultured Cells during the Induction of Frost Tolerance by Abscisic Acid or Low Temperature

Bromus inermis Leyss cell cultures treated with 75 micromolar abscisic acid (ABA) at both 23 and 3°C developed more freezing resistance than cells cultured at 3°C. Protein synthesis in cells induced to become freezing tolerant by ABA and low temperature was monitored by [¹⁴C]leucine incorporation. Protein synthesis continued at 3°C, but net cell growth was stopped. Most of the major proteins detected at 23°C were synthesized at 3°C. However, some proteins were synthesized only at low temperatures, whereas others were inhibited. ABA showed similar effects on protein synthesis at both 23 and 3°C. Comparative electrophoretic analysis of [¹⁴C]leucine labeled protein detected the synthesis of 19, 21 and 47 kilodalton proteins in less than 8 hours after exposure to exogenous ABA. Proteins in the 20 kilodalton range were also synthesized at 3°C. In addition, a 31 kilodalton protein band showed increased expression in freezing resistant ABA treated cultures after 36 hours growth at both 3 and 23°C. Quantitative analysis of [¹⁴C]leucine labeled polypeptides in two-dimensional gels confirmed the increased expression of the 31 kilodalton protein. Two-dimensional analysis also resolved a 72 kilodalton protein enriched in ABA treated cultures and identified three proteins (24.5, 47, and 48 kilodaltons) induced by low temperature growth.     

Nitrate Reductase Activity in Maize (Zea mays L.) Leaves: II. Regulation by Nitrate Flux at Low Leaf Water Potential

Experiments were conducted to determine whether the nitrate flux to the leaves or the nitrate content of the leaves regulated the nitrate reductase activity (NRA) in leaves of intact maize (Zea mays L.) seedlings having low water potentials (ψ_w) when other environmental and endogenous factors were constant. In seedlings that were desiccated slowly, the nitrate flux, leaf nitrate content, and NRA decreased as ψ_w decreased. The decrease in nitrate flux was caused by a decrease in both the rate of transpiration and the rate of nitrate delivery to the transpiration stream. Upon rewatering, the recovery in NRA was correlated with the nitrate flux but not the leaf nitrate content.