| The accumulation of carotenoids greatly influences the quality of ripe tomato (Lycopersicon esculentum, Mill.) fruit. The regulatory properties and gene expression of phytoene synthase (PSY), a key enzyme involved in carotenoid biosynthesis, were analyzed in tomato fruit. PSY activity was detected in the soluble fraction showing time- and protein-dependent increase in the formation of phytoene from geranylgeranyl pyrophosphate (GGPP). The enzyme was stimulated by Mg2+ and ATP, and was inhibited by Tween 20 and Triton X-100. The enzyme followed typical Michaelis-Menten kinetics with increasing GGPP. During in vivo studies, pericarp discs from breaker stage tomatoes were ripened in darkness (D), or D interrupted by daily pulses of red light (R), or R followed by far red light (FR). After 14d incubation, R-treated discs had accumulated 12 mg carotenoids/100 g fresh weight, nearly a 50% increase over D- and R/FR-treated discs. This R/FR reversibility of carotenoid accumulation was also observed in PSY activity 8 days post breaker, where it showed peak activity. However, this R/FR regulation of PSY activity was not reflected in transcript levels of PSY1, or 1-deoxy-D-xylulose-5-phosphate synthase, another key regulator of carotenogenesis, suggesting that PSY may be subject to post-translational regulation. Supplementation of R to optimize colour development of harvested tomato fruit treated with the ethylene antagonist 1-methylcyclopropene (MCP) did not augment carotenoid accumulation of pre-climacteric (breaker stage) and mid-climacteric (turning stage) fruit while ethylene perception was blocked by MCP. Phytochrome appears to act as a fine regulator of carotenogenesis, while other factors, such as fruit maturity and rates of respiration and ethylene synthesis, act as coarse regulators. As well, end-of-day R supplementation of fruits on the plant did not accelerate carotenoid accumulation during ripening. This may have been because the artificial light only caused a small improvement in the phytochrome equilibrium established under natural light conditions. However, FR-supplementation reduced the overall rates of carotenoid accumulation, thereby demonstrating that phytochrome regulation of the pathway at nighttime can impact the extent of carotenoid accumulation. |
