Does indoleacetic acid promote growth via cell wall acidification?

Growth of Triticum aestivum L. cv. Cappelle Desprez coleoptiles is promoted by 5.7×10^–5 M indole acetic acid (IAA) as effectively in pH 3.4 buffer as in water, but IAA is not effective in the presence of buffer at pH 3.0 or 3.2 A combination of 5.7×10^–5 M IAA and pH 3.4 buffer promotes growth to a greater extent than pH 3.2 buffer alone, which is optimal for acid-induced growth. IAA employed at 10^–7 M is still effective at promoting growth in the presence of pH 3.4 buffer, moreover, IAA at 10^–7 M interacts synergistically with the acidic buffer to promote growth. It is concluded that IAA and acid promote growth via separate mechanisms, and that IAA does not promote cell wall loosening by rendering the cell wall more acid.Abbreviation IAA Indoleacetic acid     

Inhibitory effect of ethanol on growth and solute accumulation by Saccharomyces cerevisiae as affected by plasma-membrane lipid composition

Incorporation of ethanol (1.0 or 1.25 M) into exponential-phase cultures of Saccharomyces cerevisiae NCYC 366 growing anaerobically in a medium supplemented with ergosterol and an unsaturated fatty acid caused a retardation in growth rate, which was greater when the medium contained oleic rather than linoleic acid. Ethanol incorporation led to an immediate drop in growth rate, and ethanol-containing cultures grew at the slower rate for at least 10 h. Incorporation of ethanol (0.5 M) into buffered (pH 4.5) cell suspensions containing d-[6-³H] glucose, d-[1-¹⁴C] glucosamine, l-[U-¹⁴C] lysine or arginine, or KH₂ ³²PO₄ lowered the rate of solute accumulation by cells. Rates of accumulation of glucose, lysine and arginine were retarded to a greater extent when cells had been grown in the presence of oleic rather than linoleic acid. This difference was not observed with accumulation of phosphate. Ethanol was extracted from exponential-phase cells by four different methods. Cells grown in the presence of linoleic acid contained a slightly, but consistently, lower concentration of ethanol than cells grown in oleic acid-containing medium. The ethanol concentration in cells was 5–7 times greater than that in the cell-free medium.     

Studies on the action of abscisic acid on IAA-induced rapid growth of Avena coleoptile segments

Summary A linear displacement transducer has been used to monitor the growth of a column of Avena coleoptile segments in flowing solution. IAA at 10^-5M in phosphate buffer of pH7 promotes growth after a latent period of 10.9 min, the initial maximum growth rate occurring after 25 min. Simultaneous treatment with 10^-5 M ABA does not affect either the latent period or the initial maximum growth rate in response to the IAA treatment, but subsequently gives rise to an inhibition of growth detectable after 30 min. In contrast, pretreatment with ABA for 100 min increases the duration of the latent period and reduces the initial maximum growth rate. Removal of the ABA rapidly relieves the inhibition of IAA-induced growth but a growth rate comparable to that of material treated only with IAA is never attained. Studies using 2-[¹⁴C]ABA and 1-[¹⁴C]IAA suggest that the latent period before ABA inhibition of growth is detectable is not due to a lag in ABA uptake, and that ABA is not acting by reducing IAA uptake.     

Rapid-growth responses of corn root segments: Effect of auxin on elongation

The effect of indole-3-acetic acid (IAA) on the elongation rates of 2 mm corn (Zea mays L.) root segments induced by citrate-phosphate buffer (or unbuffered) solutions of pH 4.0 and 7.0 was studied. At pH 7.0, auxin initially reduced the elongation rate in both buffered and unbuffered solutions. Only in buffer at pH 7.0 was auxin at a concentration of 0.1 M found to promote the elongation rate though briefly. THis promoted rate represented only ca. 20% of the rate achieved with only buffer at pH 4.0. Auxin in pH 4.0 buffered and unbuffered solutions only served to reduce the elongation rates of root segments. Some comparative experiments were done using 2 mm corn coleoptile segments. Auxin (pH 6.8) promoted the elongation rate of coleoptile segments to a level equal or greater than the maximal H ion-induced rate. The two responses of root segments to auxin are compared to auxin action in coleoptile growth.     

Characterization of phosphate absorption in maize root cortex segments

Uptake of phosphate ions by 1 mm segments of isolated maize root cortex layers was studied. Cortex segments (from roots of 8 days old maize plants) absorb phosphate ions from 1 mM KH₂PO₄ in 0.2 mM CaSCO₄ at the average rate of 34.3 ±3.2 μg P_i g^?1 (fr. m.) h^?1,i.e. 0.35± 0.02 μmol P_i g^?1 (fr. m.) h^?1. Phosphate uptake considerably increases after a certain period of “augmentation”,i.e. washing in aerated 0.2 mM CaSO₄. This increase is completely blocked by the presence of 10 μg ml^?1 cycloheximide. The relation of uptake rate to phosphate concentration in the medium was shown to have 3 phases in the concentration range of 0.02 - 40 mM. Transition points were found between 0.8–1 mM and 10–20 mM. Following K_m and V_max values were found: K_m[mM] : 0.37 - 3.82 - 27.67 V_max[μg P_i g^?1 (fr. m.) h^?1] : 3.33 - 39.40 - 66.67 We have found no sharp pH optimum for phosphate uptake. It proceeds at almost constant rate till pH 6.0 and then the uptake rate drops with increasing pH. At low phosphate concentrations (1 mM) the lowest uptake rate was found at 5 and 13 °C, while the uptake is higher at 5 °C than at 13 °C at phosphate concentrations higher than 1 mM. At these concentrations uptake rate at 35 °C is lower than at 25 °C. Phosphate uptake considerably decreased in anaerobic conditions. DNP and iodoacetate (0.1 mM) completely blocked phosphate uptake from 1 mM KH₂PO₄, while uptake from 5 and 10 mM KH₂PO₄ was left unaffected by these substances. The inhibitors of active - SH groups NEM and PCMB inhibited phosphate uptake: 10^?3 M NEM by 81.6%, 10⁴ M NEM by 42% and 10^?4 M PCMB by 42%.     

Effect of medium acidity on growth and rooting of different plant species growing in vitro

Micropropagated Choisya, Daphne, Delphinium, Hemerocallis, Hosta, Iris and Photinia were found to adjust the pH of Murashige and Skoog's plant tissue culture medium (initial pH 5.6 or 3.5) to different values depending on the species. When plant growth and rooting rates were determined after plants had been grown on media initially adjusted or buffered to values between 2.6 and 5.7 the different plant species were also found to have distinct pH requirements for optimal growth and/or rooting rates.Abbreviations MS Murashige & Skoog's (1962) medium - MS19 MS with additionally 10 g l^–1 sucrose - 80 mg l^–1 adenine sulphate and 130.9 mg l^–1 NaH₂PO₄ - BA 6-benzyladenine - NAA 1-naphthyl-acetic acid - IBA 3-indole-butyric acid - IAA 3-indole-acetic acid - 2iP N₆(2-isopentyl) adenine     

Partitioning of xylanolitic complex from Penicillium janthinellum by an aqueous two-phase system

This work evaluates the influences of five parameters (pH, PEG molecular mass, PEG concentration, concentration of buffer K₂HPO₄–KH₂PO₄ and NaCl concentration) on xylanolitic complex partitioning produced by P. janthinellum in aqueous two-phase systems, using a 2⁵ factorial experimental design. A mathematical model to quantify the influence of these parameters was attained and statistically tested. The optimum point for total protein extraction was obtained under the following conditions: pH 7.0, PEG 10 000, 3.67% PEG, 10% potassium phosphate and 12.4% NaCl. The partition coefficient (K) value experimentally obtained was 5.25 and that predicted by the model was 5.89.     

Effect of temperature and pH on absorption of carbon dioxide by a free level of mixed solutions of some buffers

The rate of absorption of carbon dioxide by solutions of NaHCO₃, KH₂PO₄, hydrogencarbonate, phosphate and borate buffers at 20, 30 and 40°C was determined manometrically. The absorption rate increases for all buffers tested with increasing pH. The CO₂ absorption rate by KH₂PO₄ and by the phosphate buffer at low pH is lower than that of water. For other buffers tested it is equal to or higher than that of water, especially at higher temperatures.     

Synthesis of (R)-2-phenylpropanoic acid from its racemate through an isomerase-involving reaction by Nocardia diaphanozonaria

(R)-2-Phenylpropanoic acid was synthesized from the racemic acid through an isomerization reaction involving resting cells of Nocardia diaphanozonaria JCM3208. The isomerization activity of the cells was enhanced 25-fold by adding 5.5 mM racemic 2-phenylpropanoic acid to the culture medium. When 5 mM racemic 2-phenylpropanoic acid was included in the reaction mixture (4 ml) containing resting cells (100 mg dry cell wt) in 25 mM K₂HPO₄/KH₂PO₄ buffer (pH 7.0) at 30 °C for 8 h, 4.56 mM (R)-2-phenylpropanoic acid (95.8% e.e.) was formed with a 91% molar conversion yield.     

Optimization of submerged culture conditions for exopolysaccharide production in <Emphasis Type="Italic">Sarcodon aspratus</Emphasis> (Berk) S.lto TG-3

The effect of medium components (carbon, nitrogen, and mineral sources) and environmental factors (initial pH and temperature) for mycelial growth and exopolysaccharide (EPS) production in Sarcodon aspratus(Berk) S.lto TG-3 was investigated. The optimal temperature (25°C) and initial pH (5.0) for the EPS production in shake flask cultures of S. aspratus were determined using the two-dimensional contour plot. The most suitable carbon, nitrogen, and mineral sources for EPS production were glucose, yeast extract, CaCl₂ and KH₂PO₄, respectively. Notably, the EPS production was significantly enhanced by supplementation of calcium ion. Subsequently, the optimum concentration of glucose (30gl^–1), yeast extract (15gl^–1), CaCl₂ (1.1gl^–1), and KH₂PO₄ (1.2gl^–1) were determined using the orthogonal matrix method. The effects of nutritional requirement on the mycelial growth of S.aspratuswere in regular sequence of glucose>KH₂PO₄>yeast extract>CaCl₂, and those on EPS production were in the order of glucose>yeast extract>CaCl₂>KH₂PO₄. Under the optimal culture conditions, the maximum EPS concentration in a 5-l stirred-tank reactor was 2.68gl^–1 after 4days of fermentation, which was 6-fold higher than that at a basal medium. The two-dimensional contour plot and orthogonal matrix method allowed us to find the relationship between environmental factors and nutritional requirement by determining optimal operating conditions for maximum EPS production in S.asparatus. The statistical experiments used in this work can be useful strategies for optimization of submerged culture processes for other mushrooms.     

The effect of auxins (IAA and 4-Cl-IAA) on the redox activity and medium pH of Zea mays L. root segments

Indole-3-acetic acid (IAA) and 4-chloroindole-3-acetic acid (4-Cl-IAA) were tested at different concentrations and times for their capacity to change the redox activity and medium pH of maize root segments. The dose-response surfaces (dose-response curves as a function of time) plotted for redox activity and changes in medium pH (expressed as ΔpH) had a similar shape for both auxins, but differed significantly at the optimal concentrations. With 4-Cl-IAA, the maximal values of redox activity and medium pH changes were observed at 10⁻¹⁰ M, which was a 100-fold lower concentration than with IAA. Correlations were observed between redox activity and medium pH changes at the optimal concentrations of both IAA and 4-Cl-IAA. The results are discussed herein, taking into account both the concentration of the auxins and the effects produced by them.     

Isolation of Myrosinase Producing Microorganism

Screening tests were undertaken to obtain microorganisms which produce myrosinase. One strain of microorganism indicated a strong capasity for producing myrosinase. The morphological and physiological characteristics of this strain were studied. The organism was identified as Enterobacter cloacae, no. 506.

Enzyme production was induced by the addition of 0.01% sinigrin and 6% mustard extract*² to the culture medium. The highest production was obtained after 36~40 hr cultivation when the strain was cultivated at 28°C in a medium containing 0.01% sinigrin, 6% Mustard ext., 0.1% KH₂PO₄, 0.1% NH₄Cl, 0.1% NaCl and 0.1% MgSO₄·7aq, (pH 7.0).     

Energetics of the response of maize coleoptile tissue to indoleacetic acid

Indole-3-acetic acid (IAA) promotes an increase in steady-state heat production by corn (Zea mays L.) coleoptile tissue; this increase is associated with an elevation in aerobic respiration rates. A detailed time dependence of the exothermic response to IAA was obtained using flow calorimetry. The latent period and magnitude of response were evaluated as a function of IAA concentration and pH. The data indicate that more than one response may occur. The optimal change in heat production was produced by an IAA concentration of 3·10^-5 M. It was initiated within 5 min after the start of the IAA treatment, and reached a magnitude in excess of 25% of the tissue's basal heat production. Concentrations of IAA greater than 1·10^-4 M resulted in diminished response(s), but the effect was strongly pH dependent. Several possibilities for the increased heat production triggered by IAA are discussed.Abbreviation IAA indole-3-acetic acid     

Promotion of elongation and acid invertase activity in Phaseolus vulgaris L. internode segments by phenylacetic acid

Phenylacetic acid (PAA) significantly stimulated the elongation of isolated Phaseolus vulgaris internodal segments and prevented the decline in acid invertase specific activity observed in segments incubated in the absence of growth substances. Unlike IAA, which stimulated both elongation and invertase activity over a very wide range of concentrations (<10^-4 - 1 mol.m^-3; optimum 10^-2 mol.m^-3), the response to PAA was restricted to a much narrower range of concentrations (3 × 10^-2 - 1 mol.m^-3; optimum ca. 1–2 × 10^-1mol.m^-3). At the optimum concentration of PAA, the stimulation of both responses was about 63–75% of that induced by the optimum concentration of IAA. The differences in the concentration range and magnitude of the responses to IAA and PAA were not due to differences in uptake of the two compounds. The stimulation of elongation by both compounds was prevented by 3.6 × 10^-2mol.m^-3 cycloheximide (CH), and acid invertase activites were greatly reduced compared with samples treated with growth substances alone. A saturating concentration of the specific auxin efflux carrier inhibitor N-1-naphthylphthalamic acid (NPA) slightly promoted the growth of control segments, probably by reducing the loss of residual endogenous auxin to the incubation medium. The elongation induced by PAA at its optimum concentration was considerably greater than the elongation induced by NPA, indicating that PAA did not cause growth by preventing the loss of endogenous auxin from the segments. Elongation responses to combinations of IAA and PAA suggested that the compounds were acting additively and that they were affecting growth by the same mechanism.     

Nature of the calcium effect in phosphate uptake by barley roots

Summary The rates of phosphate uptake of young intact plants were measured in solution at concentrations between 3.2×10^–7 M and 3.2×10^–5 M KH₂PO₄. The addition of 0.005 to 0.01M CaCl₂ increased the uptake rates to values of up to 4 times those of controls. The increases in uptake relative to the controls were most marked at the lowest phosphate concentrations. The addition of KCl also increased the uptake rates, but to a much smaller extent. The results are analyzed by the enzyme kinetic theory, and it is concluded that the uptake of phosphate at low ionic strengths is impeded by negative potentials at the root surface.The experimental results in this letter have appeared in a thesis.     

Role of electrolytes and starvation in altering apparent radiosensitivity of baker's yeast

1. Respiration and fermentation of yeast receiving 90,000 r of 250 kv. x-rays were studied under a variety of conditions. This dose will nearly completely inhibit growth or colony formation. 2. The apparent effects of irradiation are quite dependent on the K⁺ and H⁺ of the suspending medium. At pH 4.5 stimulatory effects were observed in KH₂PO₂ buffer and inhibition in potassium-free (T-S-T) buffer. At pH 6.5 the situation was reversed and the effects were very small (about 10 per cent). Addition of K⁺ to irradiated yeast in T-S-T buffer at pH 4.5 can completely reverse the inhibition seen. 3. Starving increases the apparent radiosensitivity of respiration and fermentation, probably by depletion of metabolite and/or electrolyte reserves. 4. Treatment with a cation exchange resin (dowex 50) results in marked inhibition of these processes in irradiated yeast, either fresh or starved. This was most effective if given after irradiation. Almost complete inhibition of anaerobic CO₂ production occurs with starvation, irradiation, and dowex treatment combined. 5. The effects of starvation and cation exchange resin treatment can be reversed, though not completely, by adding K⁺ to the medium.     

Role of protein and RNA synthesis in the initiation of auxin-mediated growth in coleoptiles of Zea mays L.

The kinetics of inhibition by protein- and RNA-synthesis inhibitors (cycloheximide and cordycepin, respectively) of indole-3-acetic acid (IAA)-induced elongation growth were investigated using abraded coleoptile segments of Zea mays L. Removal of the cuticle — a diffusion barrier for solutes — by mechanical abrasion of the outer epidermal cell wall increased the effectiveness of inhibitors tremendously. In an attempt to elucidate the role of growth-limiting protein(s) (GLP) in the growth mechanism the following results were obtained. The elongation induced by IAA was completely inhibited when cycloheximide (10 mol·l^-1) was applied to abraded coleoptile segments as shortly as 10 min before the onset of the growth response (=5 min after administration of IAA). However, when cycloheximide was applied after 60 min of IAA treatment (when a steady-state growth rate is reached), the time required for complete cessation of growth was much longer (about 40 min). Cycloheximide inhibited the incorporation of [³H]leucine into protein within about 5 min. Cordycepin (400 mol·l^-1) prevented IAA-induced growth when applied as shortly as 25 min before the onset of the growth response (=10 min before administration of IAA) but required more than 60 min for a full inhibition of steady-state growth. The incorporation of [³H]adenosine into RNA was inhibited by cordycepin within 10 min. It is concluded that, contrary to previous investigations with nonabraded organ segments, the initiation of growth by IAA depends directly on the synthesis of GLP. Moreover, the apparent lifetime of GLP is at least four times longer than the time required by cycloheximide to inhibit the initiation of growth by IAA. This is interpreted to mean that GLP is not present before IAA starts to act but is synthesized as a consequence of IAA action starting a few minutes before the initiation of growth. Interpreting the kinetics of growth inhibition by cordycepin in a similar way, we further conclude that GLP synthesis is mediated by IAA-induced synthesis of the corresponding mRNA which starts about 10 min before the onset of GLP synthesis. Inhibition by cycloheximide and cordycepin of IAA-induced growth cannot be alleviated by acidifying the cell wall to pH 4-5, indicating that these inhibitors do not act on growth via an inhibition of auxin-mediated proton excretion.Abbreviations CHI cycloheximide - COR cordycepin - GLP growth-dimiting protein(s) - IAA indole-3-acetic acid - mRNA_GLP mRNA coding for GLP     

Zum Einfluss von anorganischem Phosphat auf die Phosphohydrolasen von Weizenembryonen (Triticum aestivum L.)

Phosphate is an effective inhibitor of the phosphohydrolases of wheat embryos (Triticum aestivum L.).In vitro the inhibition was competitive with K₁ = 4.48 × 10^-3 M (1.6 × 10^-3 M P₁, applied as KH₂PO₄).In vivo the inhibition with KH₂PO₄ was relatively low during germination (48 h), but inhibition increased when other P₁-compounds were used. In ungerminated embryos the phosphatase activity comprised three isozymes; the pattern of iaozymes changes during germination, but there was no influence of P₁ in comparison with the control. Concluding we may state that the activity of phosphohydrolases is regulated by an inhibition mechanism caused by P_i-concentration.      

Effect of IAA and 4-Cl-IAA on growth rate in maize coleoptile segments

The dose-response curves for IAA and 4-Cl-IAA-induced growth of Zea mays L. coleoptile segments were studied as a function of time. Moreover, some characteristic growth parameters for both auxins were compared. The dose-response curve of growth rate measured after IAA or 4-Cl-IAA application was bell-shaped in all experiments. The optimum concentration was 10⁻⁶ M for 4-Cl-IAA and was found not to depend on the time of the growth measurement. However, in the case of IAA the optimum shifted from 10⁻⁶ M at the time of maximal growth rate to 10⁻⁵ M or even 10⁻⁴ M, when growth measured 3–4 hours after auxin application was analysed. The relative activity of 4-Cl-IAA-induced growth rate (as compared to IAA) increased significantly with increasing time from addition of this auxin to the medium. For both auxins the time needed to reach the maximal growth rate was clearly related to their concentrations. These data provided further evidence that 4-Cl-IAA is much more active auxin than IAA and can also suggest that IAA is more rapidly metabolized in comparison to 4-Cl-IAA.     

Proline accumulation induced by weak acids and IAA in coleoptiles of wheat seedlings

Proline accumulation in coleoptiles of wheat seedlings or in excised coleoptile segments incubated under shaking for a 24 h period was studied. There was no increase of proline content of coleoptiles after incubation of the seedlings in 5 mM citric acid (a relatively strong and slowly penetrating organic acid) in a pH range from 4.5 to 7.0 and only a slight increase of proline content after incubation in phosphate buffer at pH 7.0 to 7.5 duo to the higher osmotic concentration of phosphate buffer in this pH range. Quite different results were obtained with seedlings incubated in 10 mM acetic acid, a weak and easily penetrating organic acid. With increasing proton concentrations, proline accumulation increased. Application of 400 mM mannitol or higher concentrations of IAA (more than 10⁻⁵M) additionally increased proline accumulation in the presence of 10 mM acetic acid in the pH range from 6.0 to 7.5 in which acetic acid alone was loss effective. It is suggested that a decrease of cytosolic pH causes stress—induced proline accumulation.