Auxin Balance in the Avena Coleoptile

Coleoptiles of Avena possessed the capacity to degrade infiltrated indole-3-acetic acid (IAA). This activity decreased along the length of the coleoptile from apex to base on the bases of fresh weight, dry weight and protein; the apical 1 cm segment degraded more IAA than segments from other parts of the coleoptile. The naturally occurring inhibitor of the IAA oxidase activity increased in concentration up to 20 mm from the coleoptile apex; beyond, it decreased gradually towards the base. The spatial distribution of this inhibitor does not explain the gradient in IAA oxidase activity. Growth in length of the coleoptile and the IAA inactivating capacity of the apical 1 cm segment, increased 5- and 4,4-fold, respectively, between the ages of 70 and 130 h; but auxin secretion into agar platelets by the apical 2 mm of the coleoptile registered only a 2.7-fold increase. Deseeding and derooting the seedlings reduced the subsequent growth, diffusible auxin content and the IAA oxidase activity of the coleoptiles; derooting proved to be more deleterious than deseeding. A parallel reduction was evident in auxin content and IAA degrading activity following these treatments. Application of the cytokinin 6-benzylaminopurine (BAP) to coleoptiles of derooted seedlings failed to influence their capacity to degrade IAA. Nor was the activity of the aldehyde oxidase, which converts indole-3-acetaldehyde (IAAld) to IAA, affected by such treatment.     

The Effect of High Concentrations of Auxin on the Extension of Avena Coleoptile Sections

The response of Avena coleoptile sections to high concentrationsof auxin has been determined in the absence of all additivesexcept sucrose. In most experiments the growth-time curves with75 p.p.m. IAA showed two linear phases. In the first phase,which lasted for only 2–4 hours, extension was as rapidwith 75 p.p.m. IAA as with 5 p.p.m. IAA. This rapid initialexpansion phase was then succeeded by a second phase which persistedfor at least 20 hours. During this second linear phase the growth-ratewith 75 p.p.m. IAA was lower than with an auxin concentrationof 5 p.p.m. In some experiments the first phase was absent andonly the second phase was present. The response of sections to high concentrations of auxin wasnot influenced by the presence of buffers or absorbable cations.Omission of sucrose or the presence of moderate amounts of ethanolcaused the resulting growth curves to be non-linear. The rate of uptake of auxin into the tissues was dependent onthe auxin concentration and was constant for at least 24 hours.     

Osmoregulation by Oat Coleoptile Protoplasts (Effect of Auxin)

The effect of auxin on the physiology of protoplasts from growing oat (Avena sativa L.) coleoptiles was investigated. Protoplasts, isolated iso-osmotically from peeled oat coleoptile segments, were found to swell steadily over many hours. Incubated in 1 mM CaCl2, 10 mM KCl, 10 mM 2-(morpholino)ethanesulfonic acid/1,3-bis-[tris(hydroxymethyl)methylamino]propane, pH 6.5, and mannitol to 300 milliosmolal, protoplasts swelled 28.9% [plus or minus] 2.0 (standard error) after 6 h. Addition of 10 [mu]M indoleacetic acid (IAA) increased swelling to 41.1% [plus or minus] 2.1 (standard error) after 6 h. Swelling (in the absence of IAA) was partially dependent on K+ in the bath medium, whereas auxin-induced swelling was entirely dependent on K+. Replacement of mannitol in the bath by Glc increased swelling (in the absence of IAA) and eliminated auxin-induced swelling. Swelling with or without IAA was inhibited by osmotic shock and was completely reversed by 0.1 mM NaN3. Sodium orthovanadate, applied at 0.5 mM, only gradually inhibited swelling under various conditions but was most effective with protoplasts prepared from tissue preincubated in vanadate. Our data are interpreted to suggest that IAA increases the conductance of the plasma membrane to K+.     

The Effect of Gibberellin on Tryptophan Conversion and Elongation of the Avena Coleoptile

Gibberellic acid (GA₃) enhanced directly the release of ¹⁴CO₂ from tryptophan-1-^l4C by cell free preparations of Avena coleoptile tips. The rate of tryptophan metabolism in the presence of GA₃ was increased by approximately 100 per cent. The addition of auxin synthesis inhibitors to incubation flasks nullified the enhancement effect of GA₃ on elongation of the coleoptile tips. These studies implicate tryptamine as an intermediate in the formation of auxin from tryptophan. The possibility of GA₃-IAA interaction in the elongation processes was also investigated. Combination treatments of these growth-promoting substances did not induce a synergistic growth response by the coleoptile tissue.     

Inhibition of the Coleoptile Growth in Avena sativa by Endosperm Removal--Changes in Auxin, Osmotica and Cell Wall

Removal of the endosperm from 84-h-old etiolated oat seedlingsstrongly retarded the subsequent growth of coleoptiles. Thecontribution of the endosperm to coleoptile growth was studied.Endosperm removal was found to: (1) decrease the endogenouslevel of indole-3-acetic acid (IAA) in the coleoptile tip. IAAapplied to the coleoptile tip stimulated coleoptile growth inseedlings with and without the endosperm. The sensitivity ofthe coleoptile to a suboptimal concentration of IAA was higherin seedlings without the endosperm than in intact ones. At theoptimal concentration of IAA, however, the final length of thecoleoptile was larger in intact seedlings than in those withoutthe endosperm. (2) decrease the concentration of the solublesugars and amino acids in the cell sap. (3) retard the increasein the amount of polysaccharides in the cell wall of the coleoptile,particularly noncellulosic ones. (4) make the cell wall mechanicallyrigid according to stress-relaxation analysis of the cell wall.(5) induce an increase in the osmotic potential of the coleoptilecell sap. From these results, it was concluded that the endosperm suppliesthe coleoptile with IAA, sugars and amino acids, thus promotingcoleoptile growth. (Received September 24, 1987; Accepted February 3, 1988)     

Compartments and Fluxes of K, NA, and CL in Avena Coleoptile Cells

By the compartmental analysis method of MacRobbie and Dainty, and Pitman, estimates of K⁺, Na⁺, and Cl⁻ concentrations and fluxes were obtained for the cytoplasm and vacuole of coleoptile cells of oat, Avena sativa L. cv. Victory. Double labeling was used in experiments with ⁴²K plus ²²Na and with ⁴²K plus ³⁶Cl in a complete nutrient solution. At the plasmalemma, according to the Ussing-Teorell flux ratio equation, Na⁺ is pumped out and Cl⁻ is actively transported inward. The results with K⁺ are less conclusive, but it is probably pumped in. At the tonoplast there is an active inward transport of Na⁺ and probably of K⁺, but the status of Cl⁻ is uncertain, depending upon whether there is an electrical potential difference between the cytoplasm and vacuole. The results suggest that ion selectivity resides mostly in the plasmalemma. Possible errors in the estimates and interpretations are discussed.     

beta-d-Glucan of Avena Coleoptile Cell Walls

A specific glucanase was used to liberate a noncellulosic beta-d-glucan from isolated cell walls of Avena sativa coleoptile tissue. Cell walls of this tissue contain as much as 7 to 9 mg of glucan/100 mg of dry wall. Because of the specific action pattern of the enzyme, a linkage sequence of.. 1 --> 4 Glc 1 --> 3 Glc 1 --> 4 Glc.. is indicated and the predominance of trisaccharide and tetrasaccharide as hydrolytic products suggests a rather regular repeating pattern in the polysaccharide. The trisaccharide and the tetrasaccharide are tentatively identified as 3-O-beta-cellobiosyl-d-glucose and 3-O-beta-cellotriosyl-d-glucose, respectively. Recovery of these oligosaccharides following glucanase treatment of native wall material was feasible only after wall-bound glucosidases were inactivated. In the absence of enzyme inactivation the released fragments were recovered as glucose. The beta-d-glucan was not extracted from walls by either hot water or protease treatment.Cell walls prepared from auxin-treated Avena coleoptile segments yielded less glucan than did segments incubated in buffer suggesting an auxin effect on the quantity of this wall component. No IAA-induced change in the ratio of the trisaccharide and tetrasaccharide could be detected, suggesting no shift in the 1,3 to 1,4 linkage ratio. While the enzyme acts directly on the beta-d-glucan, no elongation response was apparent when Avena sections were treated with the purified glucanase. The presence of the glucan was not associated with any wound response which could be attributed to the preparation of coleoptile segments. The relationship of glucan metabolism to auxin growth responses is discussed.     

Auxin Has No Effect on Modification of External pH by Soybean Hypocotyl Cells

The cellular adjustment of the pH of the external environment of soybean (Glycine max) hypocotyl elongating cells, frequently assumed to be hydrogen ion secretion when the pH is lowered, is unaffected by auxin. These elongating cells actively adjust the external hydrogen ion concentration (from any pH in the range of 4-8) to pH 5.4 + 0.2. This pH adjustment occurs in a medium which does not contain potassium. Growth-optimum auxin concentrations have no effect on cellular pH adjustment of the external medium, whether added at the beginning of the experiment or after the equilibrium pH is attained. The pH adjustment by the cells occurs rapidly and in spite of the presence of a cuticle.