Immunoaffinity chromatography of abscisic acid combined with electrospray liquid chromatography–mass spectrometry

Hradecká, Veronika; Novák, Ondřej; Havlı́cěk, Libor; Strnad, Miroslav

doi:10.1016/j.jchromb.2006.09.034

Cited by 45 publications

(31 citation statements)

References 55 publications

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“…However, the spatial localization of ABA within the growing root tip is unknown. We developed an optimized immunofluorescence technique using a commercial rabbit anti-abscisic acid antibody (anti-ABA) that recognizes ABA and ABA conjugates (Hradecká et al, 2007;Turecková et al, 2009). By using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC), which activates carboxyl groups to conjugate to nearby amino groups, we immobilized ABA within the tissue (Sotta et al, 1985;Sossountzov et al, 1986;Peng et al, 2006).…”

Section: Resultsmentioning

confidence: 99%

“…Whole 7-d-old Arabidopsis seedlings were vacuum-infiltrated for 2 h and then fixed overnight at 4°C in 50 mM sodium PBS, pH 7.2, containing 4% (v/v) paraformaldehyde (EMS), 0.2% (v/v) glutaraldehyde (Sigma-Aldrich), and 2% (v/v) EDC (Thermo Scientific), an essential step allowing the fixation of ABA in situ, via crosslinking by its carboxyl group (Sotta et al, 1985;Sossountzov et al, 1986;Peng et al, 2006). Fixed roots were washed three times with PBS and then were simultaneously blocked and permeabilized with 3% (w/v) nonfat dried milk solution in 0.01 M PBS (PBS-milk), pH 7.2, containing 0.1% cellulose, 0.1% pectinase, and 0.1% Triton for 1 h. Permeabilized root tips were washed three times with PBS and incubated with 0.25 µg anti-ABA polyclonal antibodies (rabbit anti-ABA; Hradecká et al, 2007;Turecková et al, 2009) (C1, AS09446; Agrisera) overnight at 4°C. The stained roots were washed and transferred to a 400-fold dilution of secondary antibody goat anti-rabbit IgG-conjugated to Alexa Fluor 488 (excitation, 488 nm; emission, 505 to 530 nm) (Invitrogen, Molecular Probes) or Alexa Fluor 555 (543-nm argon laser line and detected via a 560-to 615-nm band-pass filter [red]) for 2 h at room temperature.…”

Section: Immunofluorescence and Confocal Microscopymentioning

confidence: 99%

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Environmental Nitrate Stimulates Abscisic Acid Accumulation in Arabidopsis Root Tips by Releasing It from Inactive Stores

2016

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Abscisic acid (ABA) signaling plays a major role in root system development, regulating growth and root architecture. However, the precise localization of ABA remains undetermined. Here, we present a mechanism in which nitrate signaling stimulates the release of bioactive ABA from the inactive storage form, ABA-glucose ester (ABA-GE). We found that ABA accumulated in the endodermis and quiescent center of Arabidopsis thaliana root tips, mimicking the pattern of SCARECROW expression, and (to lower levels) in the vascular cylinder. Nitrate treatment increased ABA levels in root tips; this stimulation requires the activity of the endoplasmic reticulum-localized, ABA-GE-deconjugating enzyme b-GLUCOSIDASE1, but not de novo ABA biosynthesis. Immunogold labeling demonstrated that ABA is associated with cytoplasmic structures near, but not within, the endoplasmic reticulum. These findings demonstrate a mechanism for nitrate-regulated root growth via regulation of ABA accumulation in the root tip, providing insight into the environmental regulation of root growth.

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Section: Resultsmentioning

confidence: 99%

Section: Immunofluorescence and Confocal Microscopymentioning

confidence: 99%

Environmental Nitrate Stimulates Abscisic Acid Accumulation in Arabidopsis Root Tips by Releasing It from Inactive Stores

2016

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“…Its role in the growth and development of these seaweeds needs to be further investigated. However, in vascular plants, ABA levels are strongly dependent on the duration of the water stress, for example in Arabidopsis thaliana (Go´mez-Cadenas et al, 2002;Lo´pez-Carbonell & Ja´uregui, 2005) and tobacco (Hradecka´et al, 2007). ABA catabolism has been examined in Brassica napus siliques (Zhou et al, 2003), lettuce seeds (Chiwocha et al, 2003), Western white pine seeds (Feurtado et al, 2004(Feurtado et al, , 2007 and in Douglas fir seeds (von Aderkas et al, 2005).…”

Section: Discussionmentioning

confidence: 99%

Endogenous cytokinins, auxins and abscisic acid inUlva fasciata(Chlorophyta) andDictyota humifusa(Phaeophyta): towards understanding their biosynthesis and homoeostasis

Stirk

Novák

Hradecká

et al. 2009

European Journal of Phycology

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Hormones are present in seaweeds, but little is known about their biosynthesis and regulation. The aim of this study was to investigate variation in concentration and composition of endogenous cytokinins, auxins and abscisic acid in two seaweeds collected over a 1 year period to gain a more complete picture of the types of hormones present during a range of environmental and developmental conditions. Ulva fasciata (Ulvales, Chlorophyta) and Dictyota humifusa (Dictyotales, Phaeophyta) were collected bimonthly from the inter-tidal zone at Rocky Bay, South Africa. Ethanol extracts of the samples containing a mixture of internal standards were purified using a combined DEAE-Sephadex-octadecylsilica column, followed by immuno-affinity chromatography and analysed by high-performance liquid chromatography -mass spectrometry to quantify different cytokinins, auxins and abscisic acid. cis-Zeatin, isopentenyladenine and their ribotide and riboside conjugates were the main cytokinins present in both U. fasciata and D. humifusa with low concentrations of trans-zeatin, dihydrozeatin and aromatic cytokinins. Very low concentrations of O-glucosides and riboside-O-glucosides and no N-glucosides were detected in any of the samples. Based on the cytokinins detected, we propose that as in higher plants, the ribotides are also the first intermediates formed during de novo biosynthesis in seaweeds and are subsequently converted to free-bases and ribosides. The only indole compounds detected in both species were free indole-3-acetic acid and indole-3-acetamide (IAM), suggesting that IAM is the intermediate in auxin biosynthesis. Endogenous abscisic acid was detected in most samples with levels in U. fasciata generally being higher than those measured in D. humifusa.

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“…Extracts were dissolved in 100 mL 99% methanol:1% acetic acid (v/v) topped up to 1 mL with 99% water:1% acetic acid (v/v), purified by solid-phase extraction on an Oasis HLB cartridges (60 mg, 3 mL; Waters), and evaporated to dryness in a Speed-Vac (UniEquip). Subsequently, the evaporated samples were methylated, purified by ABA-specific immunoaffinity extraction (Hradecká et al, 2007), and analyzed by UPLC-ESI (+)-MS/MS (Tureč ková et al, 2009). …”

Section: Analysis Of Endogenous Aba Contentsmentioning

confidence: 99%

Ethylene Interacts with Abscisic Acid to Regulate Endosperm Rupture during Germination: A Comparative Approach UsingLepidium sativumandArabidopsis thaliana

et al. 2009

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The micropylar endosperm cap covering the radicle in the mature seeds of most angiosperms acts as a constraint that regulates seed germination. Here, we report on a comparative seed biology study with the close Brassicaceae relatives Lepidium sativum and Arabidopsis thaliana showing that ethylene biosynthesis and signaling regulate seed germination by a mechanism that requires the coordinated action of the radicle and the endosperm cap. The larger seed size of Lepidium allows direct tissue-specific biomechanical, biochemical, and transcriptome analyses. We show that ethylene promotes endosperm cap weakening of Lepidium and endosperm rupture of both species and that it counteracts the inhibitory action of abscisic acid (ABA) on these two processes. Cross-species microarrays of the Lepidium micropylar endosperm cap and the radicle show that the ethylene-ABA antagonism involves both tissues and has the micropylar endosperm cap as a major target. Ethylene counteracts the ABA-induced inhibition without affecting seed ABA levels. The Arabidopsis loss-of-function mutants ACC oxidase2 (aco2; ethylene biosynthesis) and constitutive triple response1 (ethylene signaling) are impaired in the 1-aminocyclopropane-1-carboxylic acid (ACC)-mediated reversion of the ABA-induced inhibition of seed germination. Ethylene production by the ACC oxidase orthologs Lepidium ACO2 and Arabidopsis ACO2 appears to be a key regulatory step. Endosperm cap weakening and rupture are promoted by ethylene and inhibited by ABA to regulate germination in a process conserved across the Brassicaceae.

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Immunoaffinity chromatography of abscisic acid combined with electrospray liquid chromatography–mass spectrometry

Cited by 45 publications

References 55 publications

Environmental Nitrate Stimulates Abscisic Acid Accumulation in Arabidopsis Root Tips by Releasing It from Inactive Stores

Environmental Nitrate Stimulates Abscisic Acid Accumulation in Arabidopsis Root Tips by Releasing It from Inactive Stores

Endogenous cytokinins, auxins and abscisic acid inUlva fasciata(Chlorophyta) andDictyota humifusa(Phaeophyta): towards understanding their biosynthesis and homoeostasis

Ethylene Interacts with Abscisic Acid to Regulate Endosperm Rupture during Germination: A Comparative Approach UsingLepidium sativumandArabidopsis thaliana

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