Biosynthesis of auxin-induced ethylene in mung bean hypocotyls

The pathway of ethylene biosynthesis in auxin-treated mung beanhypocotyls was investigated by comparing the specific radioactivitiesof ethylene produced and S-adenosylmethionine (SAM) in the tissuefollowing the administration of 3,4-¹⁴C-methionine, and by analyzingthe methionine metabolites. When the rate of auxin-induced ethyleneproduction was low due to a low concentration of auxin, thespecific radioactivity of ethylene released was always higherthan that of SAM in the tissue. When the tissue was treatedwith auxin, the tissue produced and accumulated a methioninemetabolite which was converted into ethylene more efficientlythan methionine. The metabolite was identified as 1-aminocyclopropane-l-carboxylicacid (ACC) by means of paper and thin-layer chromatography,high voltage paper electrophoresis and co-crystallization. ACCformation was neither inhibited by low oxygen nor by the inhibitoryprotein of ethylene synthesis, but inhibited by aminoethoxyvinylglycine(AVG). ACC application to the tissue greatly reduced incorporationof 3,4-¹⁴C-methionine into ethylene. The control tissue thatwas not treated with auxin also converted ACC into ethyleneindicating that the enzyme which converts ACC into ethyleneis already present in the tissue and that auxin induced productionof the enzymatic system responsible for the conversion of methionineinto ACC. Ethylene synthesis from ACC was not inhibited by AVG,abscisic acid, cycloheximide or actinomycin D, but inhibitedby low oxygen and the inhibitory protein. (Received November 21, 1979; )     

Heat shock proteins and chilling sensitivity of mung bean hypocotyls

Excised mung bean (Vigna radiata L.) hypocotyl sections wereexposed to 40 C for up to 4 h in the presence or absence of50 µM cycloheximide (CHX) before being held at a non-chilling(20 C) or chilling (2.5 C) temperature. Mung bean hypocotyltissue is chilling sensitive, and the rate of solute leakageis highly correlated with the extent of chilling injury. A 3h heat shock at 40 C reduced chilling-induced solute leakageby up to 40%, but leakage was similar to non-heat-shocked hypocotylswhen CHX was present. Specific proteins were labelled when hypocotylswere exposed to [³⁵S] methionine during the last hour of heatshock. The nine most intense bands on the autoradiographs ofSDS-PAGE gels of extracted protein corresponded to molecularweights of 114, 79, 73, 70, 60, 56, 51, 46, and 18 kDa. The18 kDa band reached a maximum after 1 h at 40 C and then rapidlydecreased in intensity as the heat shock continued, becomingundetectable at 4 h. The four most intense bands after 3 h at40 C corresponded to molecular weights of 79, 70, 51, and 46kDa. The synthesis of these four hsps was markedly reduced whenthe hypocotyl sections were exposed to CHX during heat shock.During chilling for 6 d, the levels of hsps 79 and 70 remainedsignificantly higher in tissue that was heat shocked prior tochilling than in tissue that was not heat shocked. In contrast,the levels of hsps 51 and 46 were similar in bothheat-shockedand control tissues. Heat-shock-induced chilling tolerance waslost between 6 and 9 d ofstorage at 2.5 C; this loss coincidedwith the decay of hsps 79 and 70 to control levels. These resultssuggest that heat shock induces an increase in both chillingtolerance and the de novo synthesis of specific heat shock proteins;namely hsps 79 and 70. This is the first report showing a relationshipbetween heat-shock-induced chilling tolerance and specific heat-shock-inducedproteins. Key words: Ion leakage, protein synthesis, Vigna radiata     

Inhibition of auxin-induced ethylene production by salicylic acid in mung bean hypocotyls

Salicylic acid (SA), a common plant phenolic compound, influences diverse physiological and biochemical processes in plants. To gain insight into the mode of interaction between auxin, ethylene, and SA, the effect of SA on auxininduced ethylene production in mung bean hypocotyls was investigated. Auxin markedly induced ethylene production, while SA inhibited the auxin-induced ethylene synthesis in a dose-dependent manner. At 1 mM of SA, auxininduced ethylene production decreased more than 60% in hypocotyls. Results showed that the accumulation of ACC was not affected by SA during the entire period of auxin treatment, indicating that the inhibition of auxin-induced ethylene production by SA was not due to the decrease in ACC synthase activity, the rate-limiting step for ethylene biosynthesis. By contrast, SA effectively reduced not only the basal level of ACC oxidase activity but also the wound-and ethylene-induced ACC oxidase activity, the last step of ethylene production, in a dose-dependent manner. Northern and immuno blot analyses indicate that SA does not exert any inhibitory effect on the ACC oxidase gene expression, whereas it effectively inhibits both the in vivo and in vitro ACC oxidase enzyme activity, thereby abolishing auxin-induced ethylene production in mung bean hypocotyl tissue. It appears that SA inhibits ACC oxidase enzyme activity through the reversible interaction with Fe²⁺, an essential cofactor of this enzyme. These results are consistent with the notion that ethylene production is controlled by an intimate regulatory interaction between auxin and SA in mung bean hypocotyl tissue.     

Influence of ethylene on adventitious root formation in mung bean hypocotyl cuttings

The relationship between ethylene and adventitious root formation in mung bean hypocotyl cuttings was studied.Ethephon, an ethylene-releasing compound, at 5 x 10 -5 M increased root number and root dry weight on hypo-cotyl cuttings. When ethephon was applied to hypocotyl at different times after excision, there were two effectivetimes for root production i.e. between 06 h and 18-24 h. These two time periods correspond to the induction phase and the late initiation phase of root development, respectively. After excision, three peaks of ethylene productionwere observed. The first peak commencing at 6 h started the sequence of reactions leading root formation, the second peak appearing at 12 h coincided with the beginning of the increase of the IAA level during primordia initiation, and the third peak showing at 48 h played a role in root differentiation and growth. Ethylene stimulated rooting by enhancing the increase in auxins. Thus it appears that the IAA-induced ethylene production may be a factor involved in the stimulation of adventitious root formation.     

A brassinosteroid-cytokinin interaction on ethylene production by etiolated mung bean segments

Three cytokinins and their ribosides were tested in conjunction with brassinosteroid (BR) for their effects on ethylene production in etiolated mung bean ( Vigna radiata L. Rwilcz cv. Berken) hypocotyl segments. When varying concentrations of BR were tried in combination with a fixed amount of kinetin (10 μ M ), there was an increase in ethylene production with increasing concentrations of BR up to 3 μ M , and a decline beyond. A stimulation in ethylene production was observed with increasing concentrations of each of the cytokinins used. However, when applied in conjunction with 3 μ M BR, all cytokinin concentrations produced similar stimulatory patterns. When 3 μ M BR or 10 μ M 6-benzylaminopurine (BAP) was applied in conjunction with increasing concentrations of IAA, there was a stimulation in ethylene production with increasing concentrations of IAA. However, 3 μ M BR and 10 μ M BAP together with varying concentrations of IAA failed to alter the level of ethylene production.     

Relationship of steroidal structure to ethylene production by etiolated mung bean segments

Several brassinosteroid (BR) analogues, cholesterol and aldosterone were evaluated for their effectiveness alone and in combination with indole-3-acetic acid (IAA) in stimulating ethylene production by etiolated mung bean ( Vigna radiata L. Rwilcz cv. Berken) hypocotyl segments. Changing the conformation of the two hydroxyl groups on C-22 and C-23 positions from α to β did not greatly reduce the efficiency of these compounds to stimulate ethylene production alone or in combination with IAA. There was little difference in activity observed when the conformation of the methyl group in the C-24 position was changed from α to β. However, when hydroxyls were deleted from the side chain in the C-22 and C-23 positions, the compound was rendered inactive alone or in combination with IAA. The compound was also inactivated by removing the 7-oxa function on the B-ring and by substituting an ethyl group for the methyl group in the C-24 position. Both aldosterone and cholesterol were ineffective in promoting ethylene production. This study shows that very stringent structural features are required for a steroid to have BR-like activity and to act synergistically with auxin in the promotion of ethylene synthesis.     

The effect of brassinosteroid on auxin-induced ethylene production by etiolated mung bean segments

Brassinosteroid, an analogue of brassinolide, (BR) (2α, 3α, 22β, 23β-tetrahydroxy-24β-methyl-B-homo-7-oxa-5α-cholestan-6-one), was tested in conjunction with indole-3-acetic acid (IAA), naphthaleneacetic acid (NAA), 2,4-dichlorophenoxyacetic acid (2,4-D), indole-3-butyric acid (IBA), indole-3-propionic acid (IPA), indole-3-pyruvic acid (IPyA), indole-3-aldehyde (IAld), indole-3-carbinol (ICB) or tryptophan (TRP) for its effects on ethylene production by etiolated mung bean (Vigna radiata (L.) Rwilcz cv. Berken) hypocotyl segements. The enhancement of ethylene production due to BR was greatest in conjunction with 1 μM IBA, 2,4-D, IAA, or NAA (these increases were 2580, 2070, 890, and 300%, respectively). When increasing concentrations of IBA, 2,4-D, IAA, or NAA were used, there was a decrease in the percentage stimulation by BR. Both IPyA and IPA had different optimal concentrations than the other auxins tested. Their BR-enhanced maximum percentage stimulations (1430 and 1580%) were greatest with 5 μM IPya and 10 μM IPA, respectively. There was a marked reduction in the percentage stimulation by BR with either 100 μM IPyA or IPA. The inactive indoles (IAld, ICB, or TRP) did not synergize with BR at any of the concentrations tested. Four hours following treatment those segments in contact with 1 μM BR with or without the addition of 10 μM IAA began to show a stimulation in ethylene production above the control and this stimulation became greater over the following 20 h. It was necessary for BR to be in continual contact with the tissue to have a stimulatory effect on auxin-induced ethylene production. When segments excised from greater distances below the hypocotyl hook, were treated with either IAA alone or in combination with BR, there was a decrease in ethylene production with increasing distance. There was no effect of hypocotyl length on BR stimulation of auxin-induced ethylene production; however, there was a definite decrease in ethylene production when IAA was applied alone.     

Effect of the defoliant thidiazuron on ethylene evolution from mung bean hypocotyl segments

The effect of the defoliant thidiazuron (N-phenyl-N′1,2,3-thiadiazol-5-ylurea) on ethylene evolution from etiolated mung bean hypocotyl segments was examined. Treatment of hypocotyl segments with concentrations of thidiazuron equal to or greater than 30 nanomolar stimulated ethylene evolution. Increased rates of ethylene evolution from thidiazuron-treated tissues could be detected within 90 minutes of treatment and persisted up to 30 hours after treatment. Radioactive methionine was readily taken up by thidiazuron-treated tissues and was converted to ethylene, 1-aminocyclopropane-1-carboxylic acid (ACC) and an acidic conjugate of ACC. Aminoethoxyvinylglycine, aminooxyacetic acid, cobalt chloride, and α-aminoisobutyric acid reduced ethylene evolution from treated tissues. An increase in the endogenous content of free ACC coincided with the increase in ethylene evolution following thidiazuron treatment. Uptake and conversion of exogenous ACC to ethylene were not affected by thidiazuron treatment. No increases in the extractable activities of ACC synthase were detected following thidiazuron treatment.     

Stem phototropism toward blue and ultraviolet light

Positive phototropism is the process through which plants orient their organs toward a directional light source. While the blue light receptors phototropins (phot) play a major role in phototropism toward blue (B) and ultraviolet (UV) radiation, recent research showed that the UVB light receptor UVR8 also triggers phototropism toward UVB. In addition, new details of the molecular mechanisms underlying the activity of these receptors and interaction with other environmental signals have emerged in the past years. In this review, we summarize the current knowledge about hypocotyledoneous and inflorescence stem growth reorientation toward B and UVB, with a focus on the molecular mechanisms.     

Immunochemical identification of mung bean actin like protein and its cellular involvement during germination

Actin like protein, extracted and purified fromVigna radiata (mung bean) seedling, has been found to give positive enzyme-linked immunosorbent assay with mouse monoclonal antiactin antibody. In vivo studies show that cytochalasin B at sublethal dose inhibits the chromosomal movement at metaphase stage during germination. Fromin vitro studies it is found that the actin like protein isolated from mung bean seedling has a cytochalasin B binding property with a Kd value 1.2 × 10⁻⁵ M. From these two specific observations it appears probable that the biological function of mung bean actin like protein is to take part in cell division process directly or indirectly during the time of seedling development.     

Lack of involvement of polyphenol oxidase in ortho-hydroxylation of phenolic compounds in mung bean seedlings

Complete elimination of polyphenol oxidase activity in hypocotyls and leaves of developing mung bean ( Vigna radiata L. Wilczek cv. Berkin) seedlings by tentoxin had no effect on the content of the ortho-hydroxylated flavonoids delphinidin and rutin. Tentoxin completely eliminated polyphenol oxidase-mediated ortho-hydroxylation of p -coumaric acid to caffeic acid. Despite this, tentoxin had no effect on caffeic acid derivative contents in the seedlings. High performance liquid chromatography profiles indicated that elimination of polyphenol oxidase had no effect on either the quality or the quantity of soluble phenolic compounds, These data strongly indicate that polyphenol oxidase is not involved in metabolism of phenolic compounds in developing plant tissues.     

Promotion of rooting in mung bean hypocotyl cuttings with ethrel, an ethylene releasing compound

Ethrel increased the number of roots on mung bean hypocotylcuttings under continuous illumination but decreased their length.IAA, GA₃) kinetin and TIBA inhibited both the number and length.Inhibitory effect of GA3 and kinetin on root number was overcomeby ethrel but on root length was enhanced. Ethrel in combinationwith IAA promoted the number and length of roots synergisticallybut enhanced the inhibitory effect of TIBA on both. (Received June 27, 1970; )     

Uniconazole reduces ethylene and 1-aminocyclopropane-1-carboxylic acid and increases spermine levels in mung bean seedlings

Uniconazole reduced growth of etiolated mung bean seedlings and increased lateral root formation. Ethylene production for whole seedlings was reduced by 80% within 24 h after treatment and 1-aminocyclopropane-1-carboxylic acid concentrations were reduced by approximately 40% in 12 h. Uniconazole treatment increased spermine levels by 100% by day 4, whereas spermidine and putrescine levels were not affected. Uniconazole, by inhibiting ethylene synthesis, may be increasing spermine levels, which in turn stimulate formation of root primordia.     

The effect of ethylene on adventitious root formation in mung bean (Vigna radiata) cuttings

A promotive effect of ethylene on the formation of adventitious roots by mung bean cuttings was demonstrated using a recirculating solution culture system to apply dissolved ethylene. The number of roots increased in proportion to the length of exposure to the gas. Mean root numbers per cutting for a 4-day exposure to ethylene and an air control were 45 and 19, respectively. The tissue was most sensitive to a 24-h ethylene “pulse” 2–3 days after taking cuttings. Rooting was maximal at a concentration of 13 μl 1^?1 ethylene. The ethylene treatment inhibited the growth of roots and terminal buds. Application of Ag⁺, as silver thiosulfate, reversed the effect of ethylene on the two growth responses but had no effect on root numbers. Norbornadiene, another inhibitor of ethylene action, reversed all three ethylene responses.     

Stimulation of ethylene production in the mung bean hypocotyls by cupric ion, calcium ion, and kinetin

The synergistic stimulation of ethylene production by kinetin and Ca²⁺ in hypocotyl segments of mung bean (Phaseolus aureus Roxb.) seedling was further studied. The requirement for Ca²⁺ in this system was specific. Except for Sr²⁺, which mimicked the effect of Ca²⁺, none of the following divalent cations, including Ba²⁺, Mg⁶⁺, Cu²⁺, Hg²⁺, Co²⁺, Ni²⁺, Sn²⁺, and Zn²⁺, showed synergism with kinetin on ethylene production. Fe²⁺, however, showed a slight synergism with kinetin. Some of them (Hg²⁺, Co²⁺, and Ni²⁺) had a strong inhibitory effect, while others (Zn²⁺, Mg²⁺, Sn²⁺, and Ba²⁺) had a slight or no inhibitory effect on ethylene production in the absence or presence of kinetin.     

Genetic divergence and hybrid performance in mung bean

Summary Genetic divergence in 35 populations (10 parents and 25 F₁'s) of mung bean was studied by D² and canonical analyses. The ten parents formed as many as eight separate clusters, suggesting that the genetic divergence between them was quite substantial. The parent BR-2 was highly divergent from all the other entries. It was found that flowering time, maturity, seed density and seed size (100-seed weight) contributed substantially to the divergence. Canonical analysis supported the divergence pattern obtained by D² analysis and the contribution of different characters to genetic divergence. The relationship between genetic divergence (D²) and heterosis was evaluated. In general, there was fair agreement between the extent of heterosis and the genetic divergence between the parents.