Plant Physiology (Biology 327)  - Dr. Stephen G. Saupe;  College of St. Benedict/ St. John's University;  Biology Department; Collegeville, MN  56321; (320) 363 - 2782; (320) 363 - 3202, fax;    ssaupe@csbsju.edu

PLANT HORMONES - GIBBERELLINS

I. General

• "foolish seedling" disease (bakanae) - rice plants tall, weak and spindly with little grain - was observed early this century by Japanese farmers.
• associated with the fungus, Gibberella fujikuroi.
• liquid from the culture medium applied to plants caused symptoms (Kurosawa, 1926).
• three gibberellins were isolated and identified from the medium & fungus
• independent confirmation in labs in Europe and US.
• gibberellins known from angiosperms, gymnosperms, ferns, mosses, algae, fungi and even a few bacteria.

II. Chemistry

• diterpenes (C20, though some have lost a carbon and are C19) � based on isoprene skeleton
• characterized by having a complex system of 4-5 rings (ent-kaurene ring system) and a carboxyl (acidic) side chain
• more than 110 different gibberellins are known - abbreviated GA₁...GA_n.
• No more than 12 or so different gibberellins occur in any one species
• only a few (about 15) of the GA's have biological activity; the rest are likely breakdown products of, or precursors to, the active ones
• GA₁ is the most active GA
• GA₃ was the first one discovered and is readily available commercially (produced by G. fujikuroi cultures).
• C19 GA's are more active than C20
• GA's can occur in a free or conjugated form (i.e., glucosides)

III. Biosynthesis

A.  Site - young leaves, roots, and developing seeds (developing endosperm) and fruits.

B.  Pathway

• terpene pathway
• basic terpene building block is isoprene (isopentenylpryophosphate, IPP), a five carbon unit
• five major steps: (1) synthesis of IPP; (2) condensation of 4 isoprene units to form geranylgeranylpyrophosphate (GGPP); (3) GGPP cyclizes to form the kaurene ring system; (4) a methyl group of kaurene is oxidized to a carboxyl group to form GA₁₂-aldehyde; (5) GA₁₂-aldehyde is the precursor to the other GA's
• two routes to the production of IPP (isoprene): (1) Mevalonate-dependent pathway - occurs in cytosol; mevalonate is especially important for sterol biosynthesis; mevalonate is derived from acetyl CoA; and (2) Mevalonate independent pathway � especially in plastids, important for carotenoid biosynthesis

C.  GA synthesis inhibitors

• Kaurene synthesis inhibitors - Phosphon D, CCC (cycocel), and Amo1618.
• Kaurene oxidation inhibitors - Ancymidol (A-rest) and paclobutural
• Another inhibitor is B-Nine (alar).

IV. Bioassays/Analysis

A.  Three common bioassays used for gibberellin are:

• Lettuce hypocotyl elongation
• Dwarf rice (var. Tanginbozu rice) leaf sheath elongation
• alpha-amylase production in barley

B.  Instrumental methods � now the method of choice, especially GC-MS

V. Transport

• made in the tissue in which it is used
• transport occurs through xylem, phloem, or cell-to-cell.
• phloem seems to be most important transport route
• transport is not polar, as it is for auxin.
• not purely diffusion.

• Formation of conjugates
• Slow hydrolysis

A. Promotes stem elongation
    When applied to intact plants, GA usually causes an increase, unlike auxin. It overcomes dwarfism in mutants that have a mutation in the GA synthesis pathway. dwarf = short; wild type = tall; dwarf + GA = tall. Thus, GA application: (1) stimulates elongation; and (2) acts on intact plants.

GA stimulates stem elongation by:

1. stimulating cell division. Specifically, GA increases the transition from G1 � S phase of the cell cycle;
2. increasing amylase (and other hydrolytic enzymes) production � increases hydrolysis of starch � provides glucose and other sugars that: (a) lower water potential which provides driving force for water uptake; (b) provide energy through cell respiration; and (c) provide materials for building cell walls; and
3. increase cell wall plasticity (by a mechanism other than how auxin works).

B. Overcomes dormancy in seeds and buds
    Treating dormant seeds with GA stimulates germination (see below)

C. Involved in parthenocarpic fruit development - remember lab

D. Flowering
    Recall the hormone exercise we did? To summarize: LD = tall (bolts); SD = short; SD + GA = tall. Thus, GA stimulates bolting in Long Day plants and can substitute for long days or cold treatments that are necessary for flowering.

E. Mobilization of food reserves in grass seed germination
    GA is produced by the scutellum (cotyledon) of the embryo → stimulates the production of amylase by the aleurone layer � amylase hydrolyzes starch to simple sugars → absorbed by scutellum and translocated to embryo for growth.

    The production of amylase occurs de novo. That is, gibberellin stimulates transcription. In short: GA → binds to membrane receptor → interacts with a protein complex (heterotrimeric G protein) that → activates a GA signaling intermediate → turns off a repressor → transcription of GA-MYB mRNA → translated in cytosol to make GA-MYB protein → returns to nucleus to bind to alpha-amylase gene promoter region → activates transcription of alpha-amylase mRNA → translated in ribosomes on RER → transported to golgi → secretory vesicles release alpha-amylase. This last step is apparently regulated by a calcium dependent mechanism that was also activated by the heterotrimeric G protein complex.

    Brewers take advantage of GA's ability to stimulate germination and enzymes which are important in the brewing process.

F. Juvenility
    Plants exist in a juvenile and adult form. As in humans, the main difference is whether the plants are able to flower (reproduce). In some plants there is little morphological difference between juvenile and adult forms, whereas in others, the two forms are very distinct. For example, in beans, the first (juvenile) leaves are entire (heart-shaped) while the adult leaves are trifoliate. Lancewood is a New Zealand plant that has very distinctive juvenile and adult forms. In fact, they are so different that botanists originally mistook the two forms for different species. The juvenile form is unbranched with long (ca. 12 in) linear, drooping leaves that look a little like the ribs of a folded umbrella. When the plant reaches about 15 foot it switches to the adult form which is branched and has smaller, ovate leaves. It has been suggested that this is a modification to prevent predation by moa, a large, formerly common but now extinct bird.

    Gibberellin stimulates the transition between the juvenile and adult forms. In ivy, the adult form (unlobed leaves, shorter internodes) is converted to the juvenile form (lobed leaves, longer internodes) by GA treatment.

G. Sex expression
    In plants with separate male and female flowers, GA application can determine sex. For example, in cucumber, hemp and spinach, GA treatment increases the proportion of male flowers. In maize, GA treatment causes female flower development.

IX. Commercial Applications

• increase size of grapes (spray at time of blooming and fruit set stage)
• increase distance between grapes in a cluster to minimize fungi/disease
• Breweries - increase starch digestion for malting process
• Delay senescence - spray on fruit like naval oranges
• celery stalk elongation (but must use carefully because of increased storage problems)
• sugar cane � increased growth and yields
• Minimize lodging