The Role of Ethylene and Cold Temperature in the Regulation of the Apple POLYGALACTURONASE1 Gene and Fruit Softening

Emma Tacken,Daisy Wang,Jo Putterill,Judith Bowen,Sakuntala Karunairetnam,Jason W Johnston,Hilary Ireland,Keith Schultz,Ross G Atkinson,Roger P Hellens,Kularajathevan Gunaseelan,Robert J Schaffer

doi:10.1104/pp.109.151092

Abstract

Fruit softening in apple (Malus x domestica) is associated with an increase in the ripening hormone ethylene. Here, we show that in cv Royal Gala apples that have the ethylene biosynthetic gene ACC OXIDASE1 suppressed, a cold treatment preconditions the apples to soften independently of added ethylene. When a cold treatment is followed by an ethylene treatment, a more rapid softening occurs than in apples that have not had a cold treatment. Apple fruit softening has been associated with the increase in the expression of cell wall hydrolase genes. One such gene, POLYGALACTURONASE1 (PG1), increases in expression both with ethylene and following a cold treatment. Transcriptional regulation of PG1 through the ethylene pathway is likely to be through an ETHYLENE-INSENSITIVE3-like transcription factor, which increases in expression during apple fruit development and transactivates the PG1 promoter in transient assays in the presence of ethylene. A cold-related gene that resembles a COLD BINDING FACTOR (CBF) class of gene also transactivates the PG1 promoter. The transactivation by the CBF-like gene is greatly enhanced by the addition of exogenous ethylene. These observations give a possible molecular mechanism for the cold- and ethylene-regulated control of fruit softening and suggest that either these two pathways act independently and synergistically with each other or cold enhances the ethylene response such that background levels of ethylene in the ethylene-suppressed apples is sufficient to induce fruit softening in apples.

Highlights

Fruit softening in apple (Malus 3 domestica) is associated with an increase in the ripening hormone ethylene
Transgenic apple plants overexpressing PG1 have reduced cell-to-cell adhesion in the leaves (Atkinson et al, 2002), and suppression of PG1 results in firmer fruit (Atkinson et al, 2008). While these PG-suppressed apples were firmer than the controls, they were significantly softer than the ACC OXIDASE1 (ACO1)-suppressed apples, suggesting that in apples a suite of enzymes is required for fruit softening
Previous attempts at quantifying the importance of cold in apple fruit softening have been complicated by the presence of increasing endogenous ethylene production during ripening, making it difficult to determine if ripening responses were due to ethylene, cold, or a combination of these factors

Summary

Introduction

Fruit softening in apple (Malus 3 domestica) is associated with an increase in the ripening hormone ethylene. Apple fruit softening has been associated with the increase in the expression of cell wall hydrolase genes. One such gene, POLYGALACTURONASE1 (PG1), increases in expression both with ethylene and following a cold treatment. The transactivation by the CBF-like gene is greatly enhanced by the addition of exogenous ethylene These observations give a possible molecular mechanism for the cold- and ethylene-regulated control of fruit softening and suggest that either these two pathways act independently and synergistically with each other or cold enhances the ethylene response such that background levels of ethylene in the ethylene-suppressed apples is sufficient to induce fruit softening in apples. Sampling times are labeled a to i, with eight apples sampled at each time, as follows: a, sampled immediately; b, d, and e, stored in an ethylene-free environment at 20°C and sampled after

Methods

Results

Conclusion