S. J. Neill

The carotenoid and abscisic acid content of viviparous kernels and seedlings ofZea mays L.

Carotenoid and abscisic acid (ABA) levels were determined in endosperm, embryos and seedlings of wild-type and viviparous (vp) mutants ofZea mays L. Carotenoid concentrations were determined by absorption spectrometry following purification by high-performance liquid chromatography and ABA concentrations by combined gas chromatography-mass spectrometry. Lutein and zeaxanthin were the terminal carotenoids in wild-type tissue. The carotenoid profiles ofvp-1 andvp-8 tissue were similar to that of the wild type; invp-2, vp-5, vp-7 andvp-9 carotenogenesis was blocked at early stages so that xanthophylls were absent. Except forvp-1, where the ABA content was similar to the wild type, the ABA content ofvp embryos was substantially reduced, to 6–16% of the corresponding wild type. Thus, the absence of xanthophylls was associated with reduced ABA content, which was in turn correlated with vivipary. Kernels ofvp-8 had a reduced ABA content although xanthophylls were present. Seedlings of carotenoid-deficient mutants rescued from viviparous kernels contained less ABA than did wild-type seedlings grown in the same way. Furthermore, the ABA concentration of such seedlings did not increase in response to water deficit. Conversely,vp-1 seedlings contained normal levels of carotenoids and ABA. Carotenoid-deficient seedlings did not contain appreciable amounts of chlorophyll so that chloroplast development was not normal. Thus ABA-deficiency could be associated with abnormal plastid development rather than the absence of carotenoids per se.

Xanthoxin levels and metabolism in the wild-type and wilty mutants of tomato

Using 13C-labelled internal standards and gas chromatography-mass spectrometry/multiple-ion monitoring the levels of xanthoxin (Xan) and 2-trans-xanthoxin (t-Xan) have been determined in stressed and non-stressed leaves of wildtype tomato (Lycopersicon esculentum Mill cv. Ailsa Craig), and the wilty mutants, notabilis (not), flacca (flc) and sitiens (sit). Levels of Xan were very low in all tissues. Ratios of t-Xan: Xan ranged from 10:1 to <500:1. In the wild-type and flc, t-Xan levels increased following stress. The results from feeding experiments using [13C]Xan and t-Xan demonstrated that whilst wild-type and not plants readily converted Xan into abscisic acid (ABA), flc and sit plants converted only a small amount of applied Xan into ABA. In all plants t-Xan was not converted into ABA. These results indicate that the flc and sit mutants are impaired in ABA biosynthesis because they are unable to convert Xan into ABA, whereas the not mutant is blocked at a metabolic step prior to Xan. Another possible ABA precursor, ABA-1′,4′-trans-diol (ABA-t-diol) was found to occur in wild-type and mutant tissue. All four tissues could convert [2H]ABA-t-diol to ABA. Incubation of stressed leaves in the presence of 18O2 provided evidence consistent with Xan and ABA originating via oxidative cleavage of a xanthophyll such as violaxanthin.

Seed development and vivipary in Zea mays L.

Seed development was investigated in kernels of developing wild-type and viviparous (vp-1) Zea mays L. Embryos and endosperm of wild-type kernels began to dehydrate at approx. 35 d after pollination (DAP); viviparous embryos did not desiccate but accumulated fresh weight via coleoptile growth in the caryopses. Concentrations of endogenous abscisic acid (ABA) in the embryo were relatively high early in development, being approx. 150 ng·g-1 fresh weight at 20 DAP. The ABA content declined thereafter, falling to approx. 50 ng·g-1 at 30 DAP. Endosperm ABA content was always low, being less than 20 ng·g-1. There were no differences between wild-type and vp-1 tissues. Immature kernels did not germinate when removed from the ear until late in development. The ability to germinate was correlated with decreasing moisture content in the endosperm at the time of removal; premature drying of immature kernels resulted in greatly increased germination following imbibition. Excised embryos germinated precociously when removed from the endosperm as early as 25 DAP. Such germination could be prevented by treatment with 10-5 M ABA or by lowering the solute potential (Ψs) of the medium with 0.3 M mannitol. Treatment of excised embryos with ABA led to internal ABA concentrations comparable to those in embryos in which germination was inhibited in situ. Mannitol treatment did not have this effect, although water-deficit stress of excised embryos resulted in substantial ABA production. Germinated vp-1 embryos were less sensitive to growth inhibition by ABA or mannitol than germinating wild-type embryos. The vp-1 seedlings were not wilty and their transpiration rates were reduced in response to ABA or water shortage.

The biosynthesis of abscisic acid in Cercospora rosicola

Abstract 1′-Deoxyabscisic acid (1′-deoxy-ABA) has been isolated from cultures of Cercospora rosicola which are actively synthesizing abscisic acid (ABA)

Periodicity of response to abscisic acid in lateral buds of willow (Salix viminalis L.).

Aseptically cultured lateral buds of Salix viminalis L. collected from field-grown trees exhibited a clear periodicity in their ability to respond to exogenous abscisic acid (ABA). Buds were kept unopened by ABA only when the plants were dormant or entering dormancy. Short days alone did not induce bud dormancy in potted plants but ABA treatment following exposure to an 8-h photoperiod prevented bud opening although ABA treatment of buds from long-day plants did not. Naturally dormant buds taken from shoots of field-grown trees and cultured in the presence of ABA opened following a chilling treatment. In no cases were the induction and breaking of dormancy and response to ABA correlated with endogenous ABA levels in the buds.

The metabolism of α-ionylidene compounds by Cercospora rosicola

Abstract α-Ionylidene ethanol was converted to 4′hydroxy-α -ionylidene acetic acid, 1′-deoxy-ABA and ABA by Cercospora rosicola. Both the 4′-( R ) and 4′-( S -epimers of 4′-hydroxy-α-ionylidene acetic acid were detected but the configuration of the 1′-position was not established. Both epimers were metabolized to 1′-deoxy-ABA and ABA. Both the cis - and trans 1′,4′diols of ABA were also converted to ABA. 1′-Deoxy-ABA was stereospecifically hydroxylated to form ABA. 1′-Hydroxy-α-ionylidene derivatives inhibited ABA production and were only oxidised to ABA in low yield. α-Ionylidene ethanol, α-ionylidene acetic acid and both epimers of 4′-hydroxy-α-ionylidene acetic acid were identified as endogenous compounds.

Measurement of xanthoxin in higher plant tissues using 13C labelled internal standards.

Abstract A new procedure for the reliable measurement of xanthoxin from plant tissues has been developed. This accounts for both its breakdown and its isomerization to 2- trans -xanthoxin, and relies on the use of 12 C-labelled internal standards, HPLC purification and analysis by GC-MS SIM. The antioxidant tert -butylated hydroxyquinoline was found to reduce the amount of isomerization to 2- trans -xanthoxin during extraction. Levels of xanthoxin present in all tissues examined were much lower than previously thought, and 2- trans -xanthoxin: xanthoxin ratios much higher.

Partial isotope fractionation during high-performance liquid chromatography of deuterium-labelled internal standards in plant hormone analysis: A cautionary note.

Deuterium-labelled indole-3-acetic acid, abscisic acid and phthalimido-1-aminocyclopropane-1-carboxylic acid were found to separate from the unlabelled compounds on reverse-phase high-performance liquid chromatography (HPLC). A similar separation was found for the methyl esters of these compounds on normal-phase HPLC. Such separations may lead to substantial errors when these compounds are used as internal standards for quantitation by gas chromatography-mass spectrometry/selective ion detection, unless the complete chromatographic peaks are collected.

Incorporation of α-ionylidene ethanol and α-ionylidene acetic acid into abscisic acid by Cercospora rosicola

Abstract 2 H-Labelled α-ionylidene ethanol and α-ionylidene acetic acid are converted in high yield to 1′-deoxy-abscisic acid (1′-deoxy-ABA) and absc

3‐methyl‐5‐(4′‐oxo‐2′‐6′‐6′‐trimethylcyclohex‐2′‐en‐1′‐yl)‐2,4 pentadienoic acid, a putative precursor of abscisic acid from Cercospora rosicola

Although the plant growth inhibitor abscisic acid (ABA) has been intensively studied by plant physiologists and biochemists, little is known about its pathway of biosynthesis. ABA appears to play an important role in plants subjected to water stress. Under water stress conditions ABA levels in leaves can rise by up to 20-fold and this increase in ABA may be important in regulating stomata1 aperture [ 11. Attempts to gain an insight into the biochemical mechanism whereby plants control their ABA levels have been sorely hampered by a lack of knowledge of its pathway of biosynthesis. Although two pathways, a direct Crs route and an indirect C4,, route via carotenoids have been proposed, no reliable experimental evidence has been produced to substantiate either of these proposals. ‘The stereochemistry of ABA biosynthesis in Avocado fruit is consistent with both suggested routes PI‘ The major problem associated with the elucidation of the pathway of ABA biosynthesis in plants is the extremely low level of this compound found in plant tissues, even under conditions of water stress. Thus no inter~lediates on the ABA pathway have been identified to date, The observation that the plant pathogenic fungus Ccrcospora rosicola produces relatively large levels of ABA 131, has offered the possibility of determining the pathway of ABA in this organism. It may then be possible to check for the existence of a similar pathway in higher plants. This paper describes briefly the isolation of 3-methyl-5-(4’-oxo-2’,6’,6’trimethylcyclohex-2’-en-l’-yl)-2,4 pentadienoic acid (l’-desoxy-ABA) from C. rosicola. We believe that this compound may be the immediate precursor of ABA.