Fruit Ripening and Ethylene HORT 301: Plant Physiology December 9th, 2009 Dr. Michael Van Oosten

Phytochrome: regulation of Embryo and Seed development Fruit ripening Cells and cell growth Seed development Seed germination Genes and enzymes Fertilization and embryogenesis Flowering Skotomorphogenesis Biotic stress Abiotic stress Phloem translocation Water transport Photomorphogenesis Mineral nutrition Photoreceptors Nitrogen fixation Phytochrome Secondary metabolism Primary metabolism Phytochrome: regulation of light responses Respiration Photosynthesis: light reaction Photorespiration Plant life cycle Photosynthesis: carbon fixation

What Defines a Fruit? A fruit is a ripened ovary A seed is a ripened ovule

Fruit forms in higher angiosperms

The Ripening Process Process of becoming edible Fruit becomes sweeter (accumulation of sugars) Fruit becomes softer (more palatable) Fruit becomes less green (accumulation of pigments/decrease of chlorophyll) Fruit generally becomes more acidic

Fruit ripening Partial digestion of cell walls and middle lamella Degradation of chlorophyll and starch Synthesis of anthocyanins and carotinoids Respiration of organic acids PP22110.jpg 7

Respiration and Ripening Normally when a tissue reaches maturity, respiration rates drop off Climacteric fruits show a rise in respiration during onset of ripening Apples, Bananas, Aavocados Climacteric ripening is triggered by Ethylene Nonclimacteric do have a ethylene/respiration rise Oranges, Grapes, Strawberries

Ethylene during ripening process PP22110.jpg 9

Changes in the Cell Wall Much of the cell wall is degraded Expansins are produced to “loosen” cell wall Middle lamella can be selectively degrade to allow cells to become “unglued” from each other

Ethylene AIR Ethylene effects Fruit ripening/ Senescence Abscission Triple response Root hair formation AIR Elongation inhibiation Increased Radial growth Pathogen defense PP22110.jpg Ethylene effects 11

Ethylene: it’s a gas!! 1864 illuminating gas powered street lights defoliate trees 1901 Russian Dimitry Neljubov identifies ethylene as phytohormone 1917 Doubt identifies ethylene as defoliant 1934 ethylene biosynthesis in plants detected 1935 ethylene is proposed as the “ripening hormone” PP22110.jpg Biologically active at less than 0.1ppm Transported as ACC Synthesized in ripening fruit and senescing tissues Induced by auxin, draught, wounding, cold, stress, fruit ripening, senescence, pathogen attack

Ethylene: it’s a gas!! H H C C H H PP22110.jpg Biologically active at less than 0.1ppm Transported as ACC Synthesized in ripening fruit and senescing tissues Induced by auxin, draught, wounding, cold, stress, fruit ripening, senescence, pathogen attack

Ethylene biosynthesis from methionine SAM Synthetase S-Adenosyl-Methionine (SAM) ACC-Synthase NH3 COO- H2C C + 1-Aminocyclopropan- 1-carbonic acid (ACC) PP22110.jpg O2 ACC-Oxidase H2C CH2 Ethylene 14

Ethylene mutations in tomato

Antisense-Inhibition of ACC-Oxidase stops flower senescence

And Ethylene the defoliant…. Ethylene as a fruit ripening hormone And Ethylene the defoliant…. PP22110.jpg 17

leaf stem Abscission zone at base of leaf at the where it joints the stem

Ethylene induces abscission PP22090.jpg 19

Auxin prevents abscission PP22110.jpg However: unphysiological auxin concentrations have herbicide effects (agent orange)

Phenotype of first gene is masked by phenotype of a second gene Genetic epistasis Phenotype of first gene is masked by phenotype of a second gene etr1-3 ctr1-1 ein4-1 ctr1-1 etr2-1 ctr1-1 ein2-1 ctr1-1 ein3-1 ctr1-1 ein5-1 ctr1-1 constitutive triple response ethylene insensitive ETR1, EIN4, ETR2 PP22110.jpg CTR1 EIN2, EIN3, EIN5

Ethylene Mutants Ethylene insensitive (EIN) Ethylene resistant (ETR) Constitutive Triple Response (CTR) Ethylene Response Sensor (ERS) Ethylene Overproducer (ETO)

Ethylene: triple response PP22110.jpg Triple response: - thickening of hypocotyl, radial growth - reduction of cell elongation in hypocotyl and root - exaggerated curvature of apical hook, reduced geotropism

Understanding the hormone: Searching for ethylene mutants (CTR1) Wild-type air ethylene air ethylene Recessive loss-of-function ctr1 mutations: - Constitutive activation of ethylene response - Ethylene induced genes are always “on” - Constitutive triple response CTR1 leads to inhibition of ethylene response in absense of ethylene PP22090.jpg Constitutive triple response ctr1

Searching for ethylene mutants Wild-type Searching for ethylene mutants Air Ethylene etr1 Ethylene resistant (exogenous ethylene added) PP22090.jpg Constitutive triple Response (no exogenous ethylene) ctr1

Understanding the hormone: Searching for ethylene mutants (ETR1) Wild-type Air Ethylene Air Ethylene PP22090.jpg Etr (ethylene resistant) Ein (ethylene insensitive) etr1 Ethylene resistant Ethylene receptors

Structure of ethylene receptors GMP binding PP22090.jpg

Understanding the hormone: Searching for ethylene mutants (ETO1) Wild-type Understanding the hormone: Searching for ethylene mutants (ETO1) Air Ethylene - Ethylene overproducer show same phenotype as ctr1 - ctr1 not reversible by inhibitors of ethylene biosynthesis - eto phenotype is reversed by ethylene synthesis inhibitors ETO1 has de-regulated ethylene biosynthesis PP22090.jpg Ethylene overproducer eto1 29

Horticultural Question Why does blocking ethylene perception work much better than blocking ethylene production within the plant?

Ethylene and Horticulture 1-Methylcyclopropene  Binds tightly to ethylene receptor and blocks activation Marketed as: EthylBloc (Flowers SmartFresh Apple Kiwi Tomatoes Bananas Plums Avocados Melons INVINSA (drought & heat stress) Corn Soybeans Cotton Sunflower PP22090.jpg