Utilisation of the MVA pathway to produce elevated levels of the sesquiterpene α-copaene in potato tubers

Abstract

Potato flavour is a complex trait resulting from the presence of a combination of volatile and non-volatile compounds. The aim of this work was to investigate the effect of specifically altering the volatile content of tubers and assess its impact on flavour. Tuber-specific over-expression of a potato α-copaene synthase gene resulted in enhanced levels (up to 15-fold higher than controls) of the sesquiterpene α-copaene. A positive correlation (R² = 0.8) between transgene expression level and α-copaene abundance was observed. No significant changes in the levels of volatiles other than α-copaene were detected. Non-volatile flavour compounds (sugars, glycoalkaloids, major umami amino acids and 5′-ribonucleotides) were also determined. Relationships between flavour compounds and sensory evaluation data were investigated. Evaluators could not detect any aroma differences in the transgenic samples compared with controls and no significant differences in taste attributes were found. Thus although successful engineering of potato tubers to accumulate high levels of the flavour volatile α-copaene was achieved, sensory analysis suggests that α-copaene is not a major component of potato flavour.

Introduction

Potato tuber flavour is a major consideration in consumer preference but is a difficult trait to assess quantitatively (McGregor, 2007). Increasing our understanding of the molecular basis of this important quality trait will help us to make improvements in potato germplasm. Potato flavour perception is complex and results from the interaction of both volatile and non-volatile compounds (Maga, 1994, Taylor et al., 2007). This is further complicated because storage conditions and cooking treatments and have a major impact on the levels and type of flavour compounds present (Duckham et al., 2001, Duckham et al., 2002, Oruna-Concha et al., 2001, Oruna-Concha et al., 2002). Previous work has shown that the non-volatile compounds including the major umami amino acids, glutamate and aspartate, and the 5-ribonucleotides, GMP and AMP are key determinants of potato flavour (Morris et al., 2007) whereas chemicals representing the so-called sweet, sour, salty and bitter tastes do not provide a cooked potato taste (Solms, 1971, Solms and Wyler, 1979). Other non-volatile compounds e.g., salts, sugars or glycoalkaloids, do not enhance potato flavour although their presence at high levels may decrease palatability (Sinden et al., 1976, Ross et al., 1978).

Volatile compounds produced by raw and cooked potatoes have been studied extensively (Maga, 1994, Taylor et al., 2007). Recently, boiled tuber volatiles produced by Solanum tuberosum group Phureja cultivars were compared with profiles from S. tuberosum group Tuberosum (Winfield et al., 2005, Shepherd et al., 2007a, Shepherd et al., 2007b). Phureja tubers score consistently higher than Tuberosum when assessed by sensory evaluation (Winfield et al., 2005, Shepherd et al., 2007a, Shepherd et al., 2007b). Interestingly, compared with Tuberosum, Phureja tubers produce significantly higher levels (up to 100-fold) of the sesquiterpene (C₁₅) volatile α-copaene (Ducreux et al., 2008), an important aroma compound in several food plants including sweet potato (Cui et al., 2010), lettuce and carrots (Nielsen and Poll, 2007). Microarray analysis of gene expression facilitated the cloning of a potato sesquiterpene synthase gene that encodes an enzyme that catalyses the conversion of farnesyl diphosphate to α-copaene (Ducreux et al., 2008). With the identification of the potato α-copaene synthase gene, it is now possible to manipulate the tuber volatile profile through transgenic modification, providing a means of testing the importance of this volatile metabolite to sensory evaluation. Indeed, previous reports have shown that it is possible to increase the levels of important flavour volatiles in tomato fruit using a transgenic approach by manipulating terpene biosynthesis (Davidovich-Rikanati et al., 2007).

To date, a number of sesquiterpene synthases have been cloned and characterized in plants (Nagegowda, 2010, Nagegowda and Dudareva, 2007). Several examples of metabolic engineering of sesquiterpenes have been reported in plants including Arabidopsis (Aharoni et al., 2003, Kappers et al., 2005), tobacco (Hohn and Ohlrogge, 1991, Wallaart et al., 2001) and tomato (Davidovich-Rikanati et al., 2008). Although engineering of these compounds is achievable there are inherent requirements and limitations to consider (Aharoni et al., 2006) e.g., the limited availability of the cytosolic sesquiterpene precursor farnesyl diphosphate. The aim of this study was to investigate whether potato tubers could be engineered to produce elevated sesquiterpene levels by tuber-specific over-expression of a native α-copaene synthase gene. If so, how does increasing the level of α-copaene in potato influence potato flavour? To this end, flavour metabolites were measured in cooked tuber samples from control and transgenic lines and correlations with taste panel assessments are reported.