The following points highlight the three types of hormones which regulates the growth and development of plants. The hormones are: 1. Auxins 2. Cytokinins 3. Gibberellins.
Hormone # 1. Auxins:
Both naturally occurring and synthetic auxins are available and are widely used in cultures for clonal propagation work. Naturally occurring auxin is indole acetic acid (IAA). Synthetic ones are 2, 4-Dichlorophenoxy acetic acid (2, 4-D), Napthalene acetic acid (NAA), Dicamba and Picloram (Fig. 6.1).
Tissue Specific Action:
The site of biosynthesis of IAA in plants is restricted to apices, shoot meristem and active growing regions of plant system. In nature, auxin exhibits polar movement. Polarity of movement has been demonstrated for synthetic auxins like IBA, 2, 4-D and NAA. Both paranchymatous and vascular tissues (xylem and phloem) are capable of transporting auxins. Its movement in root is in acropetal fashion. Electrical field generated within plant is directly involved in downward movement of auxin stream.
Two major concepts are involved in site of auxin action, which results in growth of plant tissue. One of the main action centers for auxin is at cell wall loosening. Other concepts are focussed on nucleic acid metabolism. Auxin’s involvement in loosening of cell wall is required for cell division and consequently triggers growth and development.
Cell wall loosening is initiated by inducing secretion of hydrogen ion into the cell wall and increases acidification of the wall and dissolves micro fibrils. Subsequently, cell increases its extensibility by variation in water potential. Potassium ions are inducted into the cell to counteract electrogenic export of protons (h+). As a result, water potential is decreased which facilitates entry of water and finally cell expands.
Auxins display significant effect on RNA metabolites possibly by modulating the transcription of RNA molecules. There have been several reports on auxin enhanced polypeptide synthesis in plants. Plant tissue treated with auxin induces accumulation of several sets of new mRNA just after 20 min in culture.
It has also been documented that auxin enhances rRNA synthesis because of considerable increase in RNA polymerase I as well as its activity. Synthetic auxin, 2, 4-D treated tissues have been found to enhance the level of translatable mRNA. IAA induced rapid mRNA accumulation was observed within 10-20 min.
In addition, activation of transcription, post translations processing is accountable for auxin actions. Some of the hypothesis correlated close relationship between early auxin induced mRNA accumulation, hydrogen ion secretion and initiation of cell elongation.
mRNA induction → H+ secretion → cell elongation. Auxin can interact with DNA and facilitate expression of wide array of genes by removing repressor proteins or activation of transaction proteins. For more details, see Section I.
In vitro Functions of Auxin:
Auxins, besides their role in cell elongation, are commonly recommended for in vitro induction of callus. Most frequently used auxin for callus induction is 2, 4-D. In vitro usage of 2, 4-D at higher concentration has serious implications by generating genetic variation. Synthetic auxins like NAA are preferred over naturally occurring IAA.
Both NAA and IAA are preferred in plants for rhizogenesis. IAA in certain extent is unstable due to oxidation and irreversible conjugation. The 2, 4-D is generally used between 1 to 3 mg/L except for grasses and cereals. Higher level of 2, 4-D is necessary for callus induction among cereals and grasses.
The beneficial role of 2, 4-D is somatic embryogenesis is widely accepted. Its role is specifically displayed in the induction of embryogenic potential cells. In the combined action of plant hormones, auxins in combination with cytokinins in careful chosen concentration can play crucial role in inducing caulogenesis or rhizogenesis.
Hormone # 2. Cytokinins:
Cytokinins are an important class of plant growth substances which are essential for the regulation of growth and morphogenesis in tissue culture. Both naturally occurring and synthetic analogous are available. At least 25 different natural cytokinin growth substances have been identified (Fig. 6.2). Plant tissue culture work widely employs two of the naturally occurring cytokinins—zeatin (4-deoxy-3-methyl-trans-2-butenyl amino purine) and 2-iP (N6-(202 – isopentanyl).
Adenine and zeatin exhibit high biological activity due to trans-isomers. Synthetic cytokinins such as BAP or BA (6-benzyl amino purine) and kinetin (6-furfuryl amino purine) possess high degree of cytokinin activity.
Naturally occurring cytokinin may be degraded by cytokinin oxidases. Loss of activity is also due to irreversible conjugation of sugar or amino acids. Therefore, synthetic cytokinins (BAP or kinetin) are generally employed in most of the micro-propagation work.
Tissue Specific Action:
Cytokinins are generally synthesized in root meristem or cells retaining growth potential. Actively dividing cells in stem, leaves and cambium tissue are also involved in biosynthesis of cytokinins. They are translocated acropetally in shoots. The molecules occur not only as free bases but also as constituents of molecules like tRNA.
Naturally occurring cytokinin, IPA (Iso pentanyl adenosine) occurs immediately next to anticodon on tRNA. Recent report claims the occurrence of cytokinin active riboside also in ribosomal RNA (rRNA). Therefore, it appears that cytokinin moiety in particular species of tRNA are functionally significant. Cytokinins are also involved in gene activation for RNA synthesis and participate in enhancing enzyme activity.
In Vitro Functions:
Cytokinins appear to be implicated in sugar metabolism. It stimulates cell division and seems to be involved in regulation of tubulin protein. Callus formation from most of the dicotyledonous plants required cytokinin along with high ratio of auxins. Efficiency of cytokinin in promoting direct or indirect shoot formation is documented.
Adenine has been shown to promote shoot formation from the cells. In micropropagation work, cytokinins are generally used in multiple shoot formation directly or indirectly. In in vitro layering process, cytokinins sprout axillary buds and trigger axillary shoot formation. High concentrations of cytokinin generally inhibit root formation.
Experimental process in root culture of carrot plant showed that regeneration of shoot was possible from the callus under the influence of cytokinin. Extended work reveals that proliferation of cells from the carrot pith tissue becomes autonomous for cytokinins. Several reports have shown cytokinins are required for somatic embryogenesis.
Hormone # 3. Gibberellins:
Gibberellins are characterized by their gibbane skeleton. Although several gibberellins have been isolated from plant tissue, only gibberellic acid in view of its biological activity has been frequently used in in vitro techniques. Gibberellin is being considered by horticulturists as an important hormone due to its role in reversal of dwarfism and inters nodal elongation.
GA synthesis takes place in young leaves, root tips and predominantly in developing seeds. It can stimulate cell division and cell enlargement. In germinating seeds, gibberellic acid controls the production of hydrolytic enzymes that digest carbohydrates, fats and protein reserves of the storage endosperm tissue.
Gibberellin when applied on the whole plant can influence the growth and development; promote flowering and inducing fruit set. In micro-propagation, gibberellin is added into the media along with auxin and cytokinin particularly in I and II stage. In shoot tip culture it is added to enhance tissue growth.