Natural Growth Regulator Research Articles

Production of monoclonal antibodies to a steroid plant growth regulator

The active component of the brassin complex of higher plants has been shown to be a C28 steroidal lactone called brassinolide. This compound has been shown to enhance both cell division and cell growth. Monoclonal antibodies directed against a synthetic brassinosteroid have been produced in CAF1 mice. The mice were immunized with a saline solution containing the steroid hapten noncovalently bound to chloroform–methanol extracted fetal calf serum. Hybridomas were generated by fusing primed mouse spleen cells with SP2/0 myeloma cells. Sixty-one hybridoma clones were screened; four of these gave positive enzyme-linked immunosorbent assay (ELISA) values with the brassinosteroid hapten. One of these clones produced an immunoglobulin G antibody with a high specificity for the synthetic antigen. The brassinosteroid hapten which differs from the natural brassinolide by the position of a methyl group on the C24 carbon gave a considerably higher ELISA value than the natural growth regulator and several other plant steroids. The distribution of brassinolide in Brassica napus tissues was examined by ELISA with the monoclonal antibody.

Abscisic acid as a natural growth regulator

Abscisic acid (ABA) is accepted as one of the five major classes of natural plant growth regulators. In many tests ABA inhibits growth and metabolism, and enhances degradative changes, as in ripening and senescence. Its sites of biosynthesis appear not to be strictly localized and it is transported within the plant both within the phloem and by cell-to-cell transport. The best authenticated example of its function as a growth regulator is in the geotropic responses of roots in which growth curvature is apparently brought about by the asymmetric distribution of ABA produced in the root cap. The evidence for a rôle of ABA in the dormancy of seeds and buds, and in the ripening and abscission of fruits, is suggestive but at present incomplete. Experiments involving both exogenous and endogenous ABA suggest that ABA may play a significant role in tuberization in potato and other species. ABA inhibits hormone-induced nucleic acid and protein synthesis and there is evidence for its action both at the transcription and at the translation levels. On the other hand, ABA induces very rapid inhibition of growth which appears to be mediated via its effects on cell membrane properties. Its effects on potassium uptake may be mediated through inhibition of proton excretion by the cell. ABA counteracts the effects of other growth promoting hormones in various tests but the nature of this interaction is unknown. It seems likely that ABA plays a general role in the regulation of growth and certain aspects of metabolism, through its interaction with other growth substances.

Growth regulating effects of phenylacetic acid and p-hydroxyphenylacetic acid on Fucus spiralis L. (Phaeophyceae, Fucales) in axenic culture

Abstract Fucus spiralis L. has been brought into axenic culture by treatment of young frond tips in saturated calcium hypochlorite solution. In axenic culture the Fucus plants grow mostly with cylindrical thalli, which branch once or twice and remain about 1 cm high for several months. Additions of p-hydroxyphenylacetic acid (p-OHPAA) in a concentration range of 10−7–10−5 m as well as of phenylacetic acid (PAA) in concentrations around 10−7 m could induce normal morphology. Broad and slightly wavy fronds were formed and a growth was induced quite comparable to that in bacteria-infected cultures. As these two phenylacetic acids have been identified previously in the brown alga Undaria, the current experiments show that p-OHPAA and PAA may be natural growth regulators in brown algae.

Variations in Levels of Four Natural Growth Regulators During Growth and Flowering of ‘Gloria’ Azalea1

Abstract Three gibberellins; GA3, GA4, GA7; and abscisic acid (ABA) from the shoot tips of greenhouse grown ‘Gloria’ azaleas, Rhododendron sp.L., were tentatively identified using column chromatography, gas-liquid chromatography, and Rumex leaf senesence bioassay. Growth regulators were quantitatively estimated biweekly from 6 weeks after shoot tip removal until anthesis. GA3 levels remained nominal for the normal commercial treatment until after plants were returned to 19°C from the 9° cooler. Endogenous GA3 levels then peaked at 0.6 μg/bud at anthesis. GA4 levels remained fairly constant for all treatments and times at 0.1 μg/bud. GA7 levels remained fairly constant below 2 μg/bud except in the cold-treated plants when endogenous GA7 levels peaked at 0.6 μg/bud at the time that plants were removed from the cooler. ABA levels were similar until 24 wks. from pinch when the levels dropped to undetectable levels in cold-treated plants and increased in treatments not given a cold treatment by peaking at 0.1 μg/bud at 28 wks. from pinch. Of the commercially available gibberellins that were monitored, GA7 seemed to be the best treatment for chemically overcoming flower bud dormancy in azalea.

Ethylene and carbon dioxide as mediators in the response of the bean hypocotyl hook to light and auxins

Ethylene inhibits hook opening in the bean hypocotyl and at high concentrations induces closure of the hook. Indoleacetic acid and 2,4-dichlorophenoxyacetic acid, whose inhibitory effect on hook opening resembles that of ethylene, stimulate ethylene production from the hook tissue, and this ethylene production is physiologically active in inhibiting hook opening. It is concluded that the inhibition of opening by auxin is due at least in a major part to auxin-induced ethylene production by the hook tissue.Carbon dioxide promotes hook opening, apparently by antagonizing the action of endogenous ethylene. The concentration of respiratory CO2 in the internal gas space of the hook tissue is high enough to play a role in the regulation of hook opening.Red light causes a decrease in ethylene production and an increase in CO2 evolution from the hook tissue. These effects are partially reversible by far-red light. It is concluded that both ethylene and CO2 serve as natural growth regulators which mediate the hypocotyl hook-opening response to light in bean seedlings.

Ethylene and carbon dioxide: mediation of hypocotyl hook-opening response.

Ethylene at low concentrations inhibits the light-induced opening of the bean hypocotyl hook; auxin inhibits the opening by inducing production of ethylene. Light causes a decrease in ethylene production and an increase in the production of carbon dioxide. Hook opening appears to be a response in which ethylene serves as a natural growth regulator and in which carbon dioxide may be involved also as a growth regulator through its antagonism of the action of ethylene.

Growth-promoting activity of caffeic acid.

Vendrig and Buffel have recently stated1 that trans-caffeic acid “may be a very important natural growth regulator, not less important than indole-3-acetic acid”. This conclusion was based on the identification of caffeic acid as one of the active growth substances in an ether extract of leaves, and on growth tests with commercial caffeic acid on 2-mm. sections of Avena coleop tiles. We believe that the true explanation of these observations is quite different. In what follows, a good crystalline sample of commercial caffeic acid, m.p. 188°–190°, was used.

INFLUENCE OF SOME GROWTH‐REGULATING SUBSTANCES ON SUNFLOWER AND TOBACCO TISSUE IN VITRO

THE WELL-KNOWN responses of plant materials to various growth regulators have suggested a study of their effects on plant tissues as they were grown in vitro. This is a part of a larger work on fundamental aspects of pathological growth, in which mu%h attention has been given to the crown-gall disease caused by Phytomonas tumefaciens (Smith and Town.) Bergey et al. The early literature was reviewed by Riker and Berge (1935) and by Riker, Spoerl and Gutsche (1946). These growth regulators may be among the critical factors in the balances resulting, respectively, in or pathological growth (Riker, 1942). Tissue cultures have advantages for certain work over studies with whole plants. For example: (1) Simple, callus-like tissue may be grown vegetatively in vitro for unlimited periods free from variations arising from pollination. (2) Since the nutrients consist only of known chemicals, the basal metabolism can be studied with relatively simple and controlled procedures. (3) Changes in both kind and character of cells may be induced under controlled conditions. (4) The tissues may be exposed a long time to a wide range in concentrations of materials, including cell-stimulating and inhibiting agents, while other factors are constant. (5) It is possible to determine how far the tissue develops by itself or is dependent on other parts of the plant. Growth regulators show their presence in whole plants, for example, by epinasty, formation of adventitious roots, inhibition of bud development, stimulation of cambial activity, and delayed abscission of old leaves. A number of natural and synthetic growth regulators may produce various ones of these effects and either stimulate or inhibit abnormal cell proliferation (reviewed by Grieve, 1943; van Overbeek, 1944; Thomson, 1945). The exact role they play in crown gall is not established. Large numbers of growth regulators have been tested on many kinds of whole or decapitated plants (Thompson, Swanson and Norman, 1946,; Zimmerman, 1942, 1943). The influence of certain growth regulators has already been tested on growth in vitro of normal plant tissue (reviewed by Nobecourt, 1943; Gautheret, 1945). Indole-3-acetic acid particularly was important for growth of plant tissue cultures from many species and was indispensable in some cases. Gautheret (1945) noted that indole-butyric acid and naphthalene-acetic acid 1Received for publication April 14, 1947. This work was supported in part by the Donner Foundation and by the Research Committee of the Graduate School from funds supplied by the Wisconsin Alumni Research Foundation. Published with the approval of the Director of the Wisconsin Agricultural Experiment Station. The authors are indebted to Eugene Herrling for preparing fig. 1-10. could be substituted for indole-3-acetic acid when indole-3-acetic acid was needed for tissue cultures. With some species these materials modified the shape and kind of cells in the cultures (Gautheret, 1945). However, the effects of these materials on pathological plant tissue growing in vitro have received only limited attention. Indole-3-acetic acid and alpha-naphthalene-acetic acid -prevented bud, leaf, and stem formation on tobacco callus in a liquid medium (Skoog, 1944). The present paper describes the influence of some representative natural and synthetic growth regulators on excised tissues in vitro. It aims to clarify the stimulating or inhibiting effects on growth of various substances, their critical concentrations, and whether they act alike on two different tissues. An abstract of some of this material has already appeared (Hildebrandt and Riker, 1946b). MATERIALS AND METHODS.-Tissue cultures from two species were used. The sunflower tissue was isolated from a secondary crown-gall on Helianthus annuus L. var. Giant Russian and was free of of the crown-gall bacteria. The tobacco tissue (P. R. White strain) was from the hybrid Nicotiana glauca Grah. 9 X N. langsdorffii Weinm. S. Both tissues have already been described and illustrated (Hildebrandt, Riker and Duggar, 1945). The cultural procedure was essentially as given by Hildebrandt, Riker and Duggar (1945). Four tissue pieces were grown in each 125 ml. Erlenmeyer flask on 50 ml. of a basal, mineral-salt, agar medium. The basal medium for the sunflower tissue has been described (Hildebrandt, Riker and Duggar, 1946a). A similar medium (Hildebrandt, 1944) for the tobacco tissue had the following ingredients in milligrams per liter of distilled water: 400 Na2SO4, 800 Ca(NO3)2 4H20, 180 MgSO4 7H20, 160 KNO3, 65 KCI, 33 NaH2PO4 H2O, 9 MnSO4 4H2O, 3 ZnSO4 7H2O, 3 H3BO3, 1 KI, 20 ferric tartarate, 3 glycine, and 0.1 thiamine hydrochloride. All these chemicals were of reagent grade except boric acid and potassium iodide. In addition each liter contained 20 g. of sucrose and 5 g. of Difco agar. The growth-regulating compounds used and their sources were as follows: cysteinehydrochloride, indole-3-acetic acid, indole-butyric acid, parachlorophenoxy-acetic acid, alpha-naphthalene-acetic acid, alpha-naphthalene-acetamide, and beta-naphthoxy-acetic acid (Eastman Kodak Company); 2,4dichlorophenoxy-acetic acid, and 2,4-dichlorophenoxy-butyric acid (Dow Chemical Company); and sodium 2,4-dichlorophenoxy-acetate (E. D. Whitman, Ohio State University). The sodium 2,4-dichlorophenoxy-acetate, indole3-acetic acid, and cysteine hydrochloride, respec-