After reading this article you will learn about 1. Discovery and Chemical Nature of Auxin 2. Distribution of Auxin (IAA) in Plant 3. Biosynthesis of Auxin (IAA) in Plants and 4. Destruction/Inactivation of Auxin in Plant.
Discovery and Chemical Nature of Auxin:
The discovery of auxins dates back to last quarter of the 19th century when Charles Darwin was studying tropisms in plants. He exposed grass coleoptile to unilateral light and observed it to bend towards light. He covered the coleoptile tip with tin foil or cut it off so that it was not acted upon by light and observed that the coleoptile did not bend towards unilateral light. Thus he concluded from his experiments that some ‘stimulus’ is transmitted from upper to the lower part which induced bending of the coleoptile. These observations are recorded in his famous book on the “Power of Movement in Plants” (1880).
(Coleoptile, in monocots e.g., grasses, oats, maize etc., the plumule in the seed remains covered by a protective cap-like structure called as coleoptile. On germination of the seed, the coleoptile grows upward in the form of a tubular covering which surrounds the long narrow primary leaf during early stages of the development of the seedling as shown in Fig. 17.5. After some time, the coleoptile does not keep pace with the rapidly growing primary leaf which comes out of the coleoptile after piercing its tip and becomes exposed).
Many years later, Boysen-Jensen (1910) cut off the coleoptile tip and replaced it with a thin plate of gelatine inserted between the tip and cut stump and observed that the coleoptile could still bend towards unilateral light. Although he did not give any explanation, but it was evident from his experiment that the ‘stimulus’ mentioned by Darwin was in fact a ‘material substance’ which was in control of growth.
The explanation was given by Paal (1919) who cut off the tip of the coleoptile and replaced it asymmetrically on the cut coleoptile stump and discovered that the coleoptile bent away from the side bearing tip even in dark. Thus he concluded that the tip secretes a substance which promotes the growth of part below it.
When the tip is intact and receiving uniform light from all sides the growth is symmetrical. Therefore, the asymmetrical growth of the coleoptile resulting in curvature towards unilateral light must have been due to an asymmetrical distribution of this growth substances (now known as auxin). Larger amounts of this substances on the shaded side cause that side to grow more and the coleoptile to bend towards unilateral light.
F.W. Went (1926, 1928) was successful in isolating this growth substance from Avena coleoptile tips which still retained the growth promoting activity. He cut off the tips of the Avena coleoptile and placed them on small agar-blocks for certain period of time and then placed the agar-blocks asymmetrically on cut coleoptile stumps.
All the coleoptiles showed typical curvature even in dark. He also developed a method for determining the amount of this growth substance (i.e., auxin) which is active in very small amounts in the Avena coleoptile tips. This method or the biossay is famous by the name of Avena Curvature Test.
He concluded from this test that the curvature of the coleoptile is proportional within the limits of statistical error to the number of tips used and the length of time during which they remained on agar-blocks, or in other words, the curvature is proportional to the concentration of the active growth substance (i.e., auxin) in agar-block.
Due to extremely small amount of this growth substance in the Avena coleoptile tips and lack of accurate methods of separating the constituents of the cells, it could not be possible to chemically analyse this growth substance at that time. This probably led to the exploration of other sources, chief of which are human urine, Rhizopus cultures, malt and corn-grain oil.
Kogl and Haagen Smit (1931) isolated an active substance from human urine which was called as Auxin-A (or Auxen triolic acid). Later on in 1934, a similar active substance was isolated from malt and corn-grain oil and was named as Auxin-B (or Auxenolonic acid). Neither of these two auxins has ever been isolated again and there is considerable doubt regarding their existence.
Re-examination of human urine by Kogl e al (1934) and examination of Rhizopus culture by Thimann (1935) led to the isolation of a different substance which was named as heteroauxin (or other auxin). It appeared to be identical with an earlier known chemical compound called Indole-3-Acetic Acid (IAA)
Indole-3-acetic acid (IAA) has now been identified in a great variety of higher plants including the Avena coleoptiles by using chromatographic, chemical and colourimetric methods, and is considered to be the only true natural auxin of the higher plants.
i. Some synthetic compounds which show auxin activity (synthetic auxins), have now been detected in plants and isolated from different plant parts and now considered as natural auxins instead of synthetic auxins. An important example of such compounds is IBA (Indole-3- butyric acid) which has recently been isolated from seeds and leaves of maize and other species. 4-C1-1AA (4-chloro-indole-3-acetic acid) is another example. This chlorinated analogue of IAA has been reported from extracts of legume (pea) seeds. Structures of these two chemical compounds follow:
ii. (A naturally occurring aromatic acid called phenylacetic acid (PAA) has recently been shown to possess auxin like activity)
Distribution of Auxin (IAA) in Plant:
Auxin (IAA) is widely distributed in plant but its relative concentrations differ in different parts of the plant. Since auxin is synthesized in growing tips or meristematic regions of the plant from where it is transported to other plant parts, the highest concentrations of auxin are found in these parts such as growing shoot and root tips, young leaves and developing axillary shoots. The distribution of auxin in monocot and dicot seedlings is shown in Fig. 17.10.
In monocot seedling, the highest concentration of auxin is found in the coleoptile tip which decreases progressively toward its base. From the base of the coleoptile, the auxin concentration increases progressively up to the root tip. However, the concentration of auxin at the tip of root is much lower than at the coleoptile tip (Fig. 17.10 A).
In dicot seedling, although the pattern of auxin distribution appears to be complex, but obviously highest auxin concentrations are found in growing regions of shoot, root, young leaves and developing axillary shoots (Fig. 17.10B).
Within the plant, the auxins may be present in two forms-free auxins and bound auxins. Free auxins are those which can be easily extracted by various organic solvents such as diethyl ether or those which are easily diffusable such as that obtained in agar block from cut coleoptile tip.
Bound auxins on the other hand, need more drastic methods for their extraction from plants such as hydrolysis, autolysis, enzymolysis etc., and are not easily diffusable. Bound auxins occur in plant as complexes (conjugated auxins) usually with carbohydrates such as glucose, arabinose or sugar alcohols, or proteins or amino acids such as aspartate, glutamate or with inositol.
i. The free form of auxin is biologically active form of the hormone. In bound or conjugated form (which predominates in plants), the auxin is considered to be biologically inactive.
ii. The metabolism of bound or conjugated auxin might be a major contributing factor in controlling level of free auxin in plants.
Biosynthesis of Auxin (IAA) in Plants:
(A) Tryptophan Dependent Pathways:
In 1935, Thimann demonstrated that a fungus Rhizopus suinus could convert an amino acid tryptophan (trp) into indole-3-acetic acid (IAA). Since then, it is generally held that tryptophan is primary precursor of IAA in plants.
The indole-3-acetic acid (IAA) can be formed from tryptophan by 4 different pathways (Fig 17.11):
(a) TAM (Tryptamine) pathway:
Tryptophan is decarboxylated to form tryptamine (TAM) followed by deamination of the latter resulting in the formation of indole-3-acetaidehyde (lAld). The enzymes involved are tryptophan decarboxylase and tryptamine oxidase respectively. lAld is readily oxidised to indole-3- acetic acid (IAA) by the enzyme lAld dehydrogenase (Fig. 17.11a).
(b) IPA (Indole-3-pyruvic acid) pathway:
Tryptophan is deaminated to form indole-3-pyruvic acid (IPA) followed by decarboxylatin of the latter resulting in the formation of indole-3-acetaldehyde (lAld). The enzymes involved are tryptophan transminase and indole pyruvate decarboxylase (Fig. 17.11b).
One of the above two methods (sometimes both) is most common pathway of formation of IAA in plants.
(c) IAN (Indole-3-acetonitrile) pathway:
It occurs in some plants especially those belonging to families Brassicaceae, Poaceae and Musaceae.
Tryptophan is converted into IAA in the presence of the enzyme nitrilase. Indole-3- acetaldoxime and indoIe-3-acetonitrile (IAN) are the intermediates (Fig. 17.11 C).
(d) Bacterial pathway:
In some pathogenic bacteria such as Agrobacterium tumefaciens and Pseudomonas savastanoi, tryptophan is first converted into indole-3-acetamide (IAM) in the presence of tryptophan monooxygenase. IAM is then hydrolysed to IAA in the presence of the enzyme IAM hydrolase (Fig. 17.11d). The auxin (IAA) produced in this way often causes morphological changes in the host plant cells.
(B) Tryptophan Independent Pathway/s:
In recent years, experimental evidences for the existence of tryptophan independent pathway/s of IAA biosynthesis in higher plants have been obtained from mutants of maize and Arabidopsis (family Brassicaceae). The branch point for biosynthesis of IAA may be either indole or indole-3-glycerol phosphate with IAN and IPA as the possible intermediates. However, neither the immediate precursor of IAA in this pathway has yet been identified, nor relative importance of tryptophan dependent and independent pathways is clearly understood.
Destruction/Inactivation of Auxin in Plant:
Sufficient levels of auxin in plant required for regulation of plant growth are maintained not only by the synthesis of auxin, but also by its destruction or inactivation. Chief method for the destruction (degradation) of auxin in plant is its oxidation by O2 in the presence of the enzyme IAA-oxidase or peroxidase. This oxidation involves removal of CO2 from the carboxylic group of auxin (IAA) and results in the formation of a variety of compounds, but 3-methylene-oxindole is the major end product.
Auxin may be temporarily inactivated in plants by its conversion into its bound form (bound auxin or conjugated auxin) in which auxin is conjugated to a variety of substances such as carbohydrates, amino acids, proteins or inositol etc. Rapid inactivation of auxin may occur by irradiation with X-rays and gamma rays. Ultraviolet light is also known to reduce auxin levels in plants. Inactivation or decomposition of IAA by light has been called as photo-oxidation.