Abscisic Acid (With Diagram) | Plant Growth Hormones

Let us make an in-depth study of the Abscisic Acid (ABA). After reading this article you will learn about 1. Discovery and Chemical Nature of ABA 2. Detection & Estimation of ABA 3. Physiological Effects of Abscisic Acid 4. Biosynthesis of ABA in Plants 5. Degradation/Inactivation of ABA in Plants 6. Occurrence and Distribution of ABA in Plants and 7. ABA Transport in Plant.

Discovery and Chemical Nature of ABA:

In 1963, a substance strongly antagonistic to growth was isolated by F.T. Addicott from young cotton fruits and named Abscisin II. Later on, this name was changed to Abscisic acid (ABA). The chemical name of abscisic acid whose structure is given in fig. 17.29.

Is [3-methyl 5-1′ (1′- hydroxy, 4′-oxy-2′, 6′ , 6′-trimethyl-2-cyclohexane-l-yl)-cis, trans-2,4-penta-dienoic acid].

Eagles and Wareing (1963, 64), at the same time pointed out the presence of a substance in birch leaves (Betula pubescens, a deciduous plant) which inhibited growth and induced dor­mancy of buds and, therefore, named it ‘dormin’. But, very soon as a result of the work of Cornforth et al (1965), it was found to be identical with abscisic acid.

i. Abscisic acid is a 15-C sesquiterpene compound (molecular formula C₁₅H₂₀O₄) com­posed of three isoprene residues and having a cyclohexane ring with keto and one hydroxyl group and a side chain with a terminal carboxylic group in its structure.

ii. ABA resembles terminal portion of some carotenoids such as violaxanthin and neoxanthin (see Fig. 17.31) and appears to be a breakdown product of such carotenoids.

iii. Any change in its molecular structure results in loss of activity.

iv. ABA occurs in cis and trans-isomeric forms that are decided by orientation of -COOH group around 2nd carbon atom in the molecule (Fig. 17.29).

v. Almost all naturally occurring ABA in plants exist in cis form that is biologically active and the name abscisic and usually refers to this form.

vi. Trans-ABA is inactive form but can be inter convertible with cis-ABA.

vii. Due to the presence of a chiral centre (asymmetric carbon atom) at l’-position, ABA also occurs in two enantiomeric forms called S and R or (+) and (-) forms respectively which can be distinguished from each other by their optical rotatory dispersion curves. In the S or (+) form, which is dextrorotatory, the -OH at 1′ position faces below the plane of the ring and is shown by dashed bond line while in R or (-) form, which is laevorotatory, it faces above the plane of the ring and is shown by a thick wedge shaped bond line (Fig. 17.30).

viii. Although both S and R enantiomers of cis-ABA are biologically active, the R form is not active in fast responses of ABA such as stomatal closure.

ix. Unlike cis and trans forms of ABA, the S and R forms are not inter-convertible in plant.

x. Commercially available synthetic sample of ABA is a racemic mixture of equal amounts of both S and R forms.

Detection & Estimation of ABA:

1. Bioassay Methods:

A number of different bioassays have been developed to detect and estimate ABA in plant extracts which are based on,

(i) Acceleration of the abscission of cotyledonary petioles in explants of 14 day old cot­ton seedlings;

(ii) Inhibition of IAA-induced straight growth of Avena (oat) coleoptile;

(iii) Inhibition of GA-induced synthesis of a-amylase in aleurone layer of germinating bar­ley (Hordeum);

(iv) Inhibition of growth of duckweed Lemna minor.

(v) Inhibition of germination of isolated wheat (Triticum) embryos and,

(vi) Stimulation of stomatal closure.

Of all above bioassays, the last one that is based on stimulation of stomatal closure has fast response of guard cells and is comparatively more sensitive. It can detect as low as 10^-9 M ABA in plant extracts. Preliminary purification measures are however, necessary in all bioassays.

2. Physicochemical Methods:

The old bioassay methods have now been replaced with more accurate and reliable mod­ern physicochemical methods of separation and quantification of the hormone. These include high performance liquid chromatography (HPLC) and gas chromatography (GC) which are then followed by mass spectrometry (MS) to provide proof of structure. Such methods can detect as low as 10^-13g ABA in plant extracts. Preliminary purification steps including thin layer chro­matography (TLC) are also required in such methods.

3. Immunoassay Methods:

Like other plant hormones (auxins, gibberellins and cytokinins), highly sensitive and spe­cific immunoassays are also applied for quantification of ABA which can detect as low as 10^-13g ABA in partly purified or crude extracts. In immunoassay, specific anti hormonal antibodies are obtained by injecting the hormone (in this case ABA) in mice or rabbit. These antibodies are then used to react with the hormone in a cuvette assay.

Physiological Effects of Abscisic Acid (ABA):

(1) Stomatal Closing:

The role of ABA in causing stomatal closure in plants undergoing water-stress is now widely recognised. It has been suggested by various workers that in response to the water-stress, the permeability of the chloroplast membranes of mesophyll cells to ABA is greatly increased.

As a re­sult, the ABA synthesized and stored in mesophyll chloroplasts diffuses out into the cyto­plasm. It then moves from one mesophyll cell to another through plasmodesmata and finally reaches to the guard cells where it causes closing of stomata. Fresh biosynthesis of ABA continues in mesophyll chloroplasts during period of water-stress.

When water potential of the plant is restored (i.e., increased), the movement of ABA into the guard cells stops. ABA disappears from the guard cells a little later. The application of exogenous ABA to leaves of normal plants causes closing of stomata within a few minutes. It has been suggested that ABA causes closing of stomata by inhibiting the ATP-mediated H⁺/K⁺ ions exchange pumps in guard cells.

(2) Other Effects:

ABA has also been shown to play some role in:

(i) Inducing bud dormancy in some temper­ate zone trees such as birch (Betula pubescens), Acer, Fraxinus etc.;

(ii) Inducing dormancy of seeds which require stratification (i.e., exposure to low temp, for germination);

(iii) Process of tuberization;

(iv) Senescence of leaves;

(v) Fruit ripening;

(vi) Abscission of leaves, flowers and fruits

(vii) Increasing the resistance of temperate zone plants to frost injury,

(viii) Inhibition of GA-induced synthesis of a-amylase in aleurone layers of germinating barley;

(ix) Inhibition of precocious germination and vivipary and

(x) Increase in root : shoot ratio at low water potentials .(The role of ABA in causing abscission of leaves, flowers and fruits is controversial. ABA may stimulate abscission of organs only in a few species that too probably through increase in production of ethylene. The primary hormone causing abscission is not ABA but ethylene).

Biosynthesis of ABA in Plants:

Extensive studies done by researchers with ABA deficient mutants of tomato, Arabidopsis and other plants have clearly shown that ABA is synthesized in higher plants not from simple terpenoid precursors directly through 15-C farnesyl diphosphate (FPP), but indirectly through carotenoid pathway as breakdown product of 40-C xanthophyll such as violaxanthin or neoxanthin (Fig. 17.31).

i. The initial steps of ABA biosynthesis take place in chloroplasts or other plastids while final steps occur in cytosol.

ii. Violaxanthin is synthesized from zeaxanthin (also a 40-C xanthophyll) in a reaction that is catalysed by the enzyme zeaxanthin epoxidase (ZEP). This enzyme is encoded by ABA1 locus of Arabidopsis.

iii. Violaxanthin is converted into 9′-cis-neoxanthin. The latter is then cleaved into a 15-C compound xanthoxal (previously called xanthoxin) and a 25-C epoxy aldehyde in the presence of the enzyme 9′-cis-epoxycarotenoid dioxygenase (NCED). (This enzyme can also catalyse cleavage of violaxanthin into xanthoxal and a 25-C allenic apoaldehyde).

iv. Xanthoxal (xanthoxin) is finally converted into ABA in cytosol via two oxidation steps catalysed by the enzymes aldehyde oxidases involving abscisyl aldehyde (and/or possibly xanthoxic acid) as intermediates. The enzymes aldehyde oxidases require Mo as cofactor.

Degradation/Inactivation of ABA in Plants:

There are two ways in which ABA is degraded or inactivated in plants (Fig. 17.32).

1. Oxidation to Phaseic Acid:

This is the main route of degradation of ABA in plants. ABA is oxidised to phaseic acid (PA) with a subsequent reduction of the keto group on the cyclohexane ring to form dihydro phaseic acid (DPA).

In some cases, DPA may be further metabolized to form 4′-glucoside of DPA.

Both dihydrophaseic acid and its glucoside are metabolically inactive. Phaseic acid is also inactive or exhibits greatly reduced activity in most bioassays but not in all. Like ABA, phaseic acid (PA) can also induce stomatal closure in some species and is as effective in inhibiting GA induced synthesis of a-amylase in aleurone layers of germinating barley.

2. Conjugation as Glucosides:

Free ABA can also be inactivated in plants by covalent conjugation with some simple sugar molecule such as glucose to form ABA-p-D-glucosyl ester (ABA-GE). The latter accu­mulates in vacuole.

Occurrence and Distribution of ABA in Plants:

Abscisic acid (ABA) is a ubiquitous plant hormone in vascular plants. In bryophytes, it has been found in mosses but not in liverworts. Some fungi synthesize ABA as secondary metabolite. ABA has not been detected in any algae. However, a 15-C organic compound called lunularic acid has been found in algae and liverworts that appears to be possible functional equivalent of ABA in these plants.

Within the plant, ABA has been detected in all major organs or living tissues from root caps to apical buds such as roots, stems, buds, leaves, fruits and seeds and also in phloem and xylem sap and in nectar.

ABA is synthesized in all types of cells that contain chloroplasts or other plastids. It occurs predominantly in mature green leaves.

Most plant tissues contain ABA in conc. of 20-100 ng per g fresh weight, but higher conc. of 10 µg and 20 µg per g fresh weight have been reported in avocado fruit pulp and dormant buds of cocklebur (Xanthium spp.) respectively.

The concentration of ABA in specific plant tissues varies greatly at different developmen­tal stages or in response to environmental conditions especially water stress. For instance, in developing seeds ABA conc. may increase 100 fold within a few days and decline as the seed matures. Similarly, under water stressed conditions, ABA level may increase 50 fold in the leaves within a few hours and declines to normal when plant water potential is restored.

The concentration of ABA in plant tissue is regulated by:

(i) Its synthesis,

(ii) Degrada­tion,

(iii) Compartmentation and

(iv) Transport.

In plant, ABA predominantly occurs in its free form but it may also occur in conjugated form as glycoside with some simple sugar molecule such as glucose forming ABA-β-D-glucosyl esters. ABA is biologically inactive in its conjugated or bound form.

ABA Transport in Plant:

The transport of endogenous ABA at various stages of growth in higher plants is not very clear. However, experiments made with ¹⁴C-labelled ABA have shown that,

(i) Externally applied ABA is able to get into tissues rapidly and is distributed freely in all directions within the plant;

(ii) Cell to cell transport of ABA is slow and non-polar;

(iii) ABA is present in phloem and xylem sap and is most probably trans located through­out the plant through vascular tissue;

(iv) ABA synthesized in root cap moves basipetally into the central vascular tissue;

(v) There is some evidence of lateral movement of ABA in root tips in response to grav­ity stimulus where it causes asymmetric inhibition of growth resulting in geotropic curvature.

i. Redistribution of ABA among plant cell compartments is controlled by pH gradient across the membranes. At low pH (6.3 or less) ABA exists in protonated or un-dissociated form (ABAH) which can readily cross most cell membranes. At higher pH (7.2 or more), ABA exists in dissociated form (ABA) that is impermeant and cannot cross the membranes easily. ABA in protonated form tends to diffuse from a compartment with low or acidic pH into a compart­ment with high or alkaline pH. At higher pH, ABAH dissociates into ABA^– and is trapped.

ii. ABA is known to be transported in plant mostly in its free form. However, ABA can also be transported to some extent in its conjugated form as ABA-β-D-glucosyl ester.