Mode of action of abscisic acid in triggering ethylene biosynthesis and softening during ripening in mango fruit

Abstract

The role of abscisic acid (ABA) in triggering ethylene biosynthesis and ripening of mango fruit was investigated by applying ABA [S-(+)-cis,trans-abscisic acid] and an inhibitor of its biosynthesis [nordihydroguaiaretic acid (NDGA)]. Application of 1 mM ABA accelerated ethylene biosynthesis through promoting the activities of ethylene biosynthesis enzymes (1-aminocyclopropane-1-carboxylic acid synthase, ACS; 1-aminocyclopropane-1-carboxylic acid oxidase, ACO) and accumulation of 1-aminocyclopropane-1-carboxylic acid (ACC), enhanced fruit softening and activity of endo-polygalacturonase and reduced pectin esterase activity in the pulp. The activities of ethylene biosynthesis and softening enzymes were significantly delayed and/or suppressed in the pulp of NDGA-treated fruit. The ABA-treated fruit had higher total sugars and sucrose as well as degradation of total organic acids, and citric and fumaric acids compared with NDGA treatment. These results suggest that ABA is involved in regulating mango fruit ripening and its effects are, at least in part, mediated by changes in ethylene production.

Introduction

Abscisic acid (ABA) has been reported to play a crucial role in fruit maturation and senescence (Giovannoni, 2001, Giovannoni, 2004, Rodrigo et al., 2003, Zhang et al., 2009a, Zhang et al., 2009b). Lower levels of endogenous ABA in unripe fruit and its substantial accumulation during fruit maturation suggest that ABA plays a key role in modulating ripening and senescence in climacteric fruit such as peach (Rudnicki et al., 1968, Zhang et al., 2009a), avocado (Adato et al., 1976), tomato (Buta and Spaulding, 1994, Sheng et al., 2008, Zhang et al., 2009b), banana (Lohani et al., 2004), apple (Buesa et al., 1994), and non-climacteric fruit including grape (Inaba et al., 1976, Zhang et al., 2009a) and orange (Kojima, 1996). Additionally, the de-greening stage of ABA-deficient orange mutants commenced later than in the wild type consistent with a crucial role for ABA in maturation and ripening of orange fruit (Rodrigo et al., 2003). Some earlier reports indicated that endogenous levels of ABA increased towards harvest in the fruit skin and pulp of ‘Nam Dok Mai’, ‘Nang Klangwan’ (Kondo et al., 2004) and ‘Alphonso’ (Murti and Upreti, 1995) mangoes. Our previous study showed that the accumulation of endogenous ABA during the climacteric rise stage might initiate climacteric ethylene production during ripening of ‘Kensington Pride’ mango fruit (Zaharah et al., 2012).

Further, ABA (10⁻⁶ M) application has been reported to hasten the ripening process and induce some structural changes in ‘Alphonso’ and ‘Langra’ mangoes (Palejwala et al., 1988, Parikh et al., 1990). Application of ABA or high endogenous levels have also been reported to stimulate ethylene production, and promote ripening in other climacteric fruit such as tomato (Sheng et al., 2008, Zhang et al., 2009b, Zhu et al., 2003), peach (Zhang et al., 2009a) and non-climacteric grape berries (Deytieux et al., 2005, Zhang et al., 2009a). Zhang et al. (2009b) reported that the application of 100 μM ABA up-regulated the expression of the Le-ACO1 and Le-ACS2 genes, which encode 1-aminocyclopropane-1-carboxylic acid oxidase (ACO, EC 1.14.17.4) and 1-aminocyclopropane-1-carboxylic acid synthase (ACS, EC 4.4.1.14), and consequently increased ethylene production and accelerated tomato fruit ripening. Further, the application of inhibitors of ABA biosynthesis [fluridone or nordihydroguaiaretic acid (NDGA)], inhibited the expression of both Le-ACO1 and Le-ACS2 genes, and delayed tomato fruit ripening (Zhang et al., 2009b). The accumulation of ABA (3000 ng g⁻¹ fresh weight) during mango fruit ripening also appears to induce climacteric ethylene production which may be modulating its ripening process (Zaharah et al., 2012). However, the mechanism by which ABA regulates ethylene biosynthesis and mango fruit ripening has not been examined in detail.

Mango fruit softening is associated with increased activities of polygalacturonase (PG), exo-polygalacturonase (exo-PG; EC 3.2.1.67), endo-polygalacturonase (endo-PG; EC 3.2.1.15), pectin esterase (PE; EC 3.1.1.11), pectin lyase (PL; EC 4.2.2.2) and endo-1,4-β-d-glucanase (EGase or cellulase; EC 3.1.1.4), which are initiated by ethylene (Chourasia et al., 2006, Chourasia et al., 2008, Singh and Singh, 2011). An endo-β-1,4-glucanase homologue, MiCel1 from mango shows fruit-specific and ripening-related expression which was positively correlated with an increase in EGase activity particularly during the later stages of ripening (Chourasia et al., 2008). Previously, the exogenous application of ABA has also been reported to increase the activity of PG, but has inconsistent effects on the activity of pectin methyl esterase (PME) during ‘Zihua’ mango fruit ripening (Zhou et al., 1996). Overall the research on the effects of applied ABA and its inhibitors in regulating the activities of fruit softening enzymes during mango ripening is sporadic and inconclusive. Therefore, the aim of the present study was to investigate how applied ABA and its biosynthesis inhibitor regulate ethylene biosynthesis and fruit softening, including the activities of ethylene biosynthetic enzymes such as ACS and ACO, ACC content, as well as fruit softening enzymes including PE, exo-PG, endo-PG, EGase and the levels of sugars and organic acids in the pulp of the fruit during ripening.