Gene expression studies in kiwifruit and gene over-expression in Arabidopsis indicates that GDP-L-galactose guanyltransferase is a major control point of vitamin C biosynthesis

Abstract

Vitamin C (L-ascorbic acid, AsA) is an essential metabolite for plants and animals. Kiwifruit (Actinidia spp.) are a rich dietary source of AsA for humans. To understand AsA biosynthesis in kiwifruit, AsA levels and the relative expression of genes putatively involved in AsA biosynthesis, regeneration, and transport were correlated by quantitative polymerase chain reaction in leaves and during fruit development in four kiwifruit genotypes (three species; A. eriantha, A. chinensis, and A. deliciosa). During fruit development, fruit AsA concentration peaked between 4 and 6 weeks after anthesis with A. eriantha having 3–16-fold higher AsA than other genotypes. The rise in AsA concentration typically occurred close to the peak in expression of the L-galactose pathway biosynthetic genes, particularly the GDP-L-galactose guanyltransferase gene. The high concentration of AsA found in the fruit of A. eriantha is probably due to higher expression of the GDP-mannose-3′,5′-epimerase and GDP-L-galactose guanyltransferase genes. Over-expression of the kiwifruit GDP-L-galactose guanyltransferase gene in Arabidopsis resulted in up to a 4-fold increase in AsA, while up to a 7-fold increase in AsA was observed in transient expression studies where both GDP-L-galactose guanyltransferase and GDP-mannose-3′,5′-epimerase genes were co-expressed. These studies show the importance of GDP-L-galactose guanyltransferase as a rate-limiting step to AsA, and demonstrate how AsA can be significantly increased in plants.

Proteins involved in biotic and abiotic stress responses as the most significant biomarkers in the ripening of Pinot Noir skins

“…In our work, we selected the relative spot (spot 2233) because it was characterized by an extremely positive coefficient for RS37 model and a very negative one for RS38, reflecting that the levels of ManEp peaked in the second ripening stage and dropped in mature skins. This behavior is similar to what reported for kiwifruit: after reaching the top in the amount of the enzyme 2 weeks after anthesis, the levels of ManEp sharply downturned moving towards maturity, overlapping the trend of the levels of ascorbate in the fruit during development (Bulley et al 2009). …”

“…It has been recently indicated that in a comparison among kiwifruit genotypes that differ for the levels of AsA, the concentration of the vitamin is correlated to the high amount of GDP-mannose-3′,5′-epimerase and GDP-Lgalactose guanyltransferase also because, overexpressing the genes of the two enzymes in transgenic kiwifruit plants, a significant increase in AsA levels occurred (Bulley et al 2009). In our work, we selected the relative spot (spot 2233) because it was characterized by an extremely positive coefficient for RS37 model and a very negative one for RS38, reflecting that the levels of ManEp peaked in the second ripening stage and dropped in mature skins.…”

Allelic Variation in Paralogs of GDP-l-Galactose Phosphorylase Is a Major Determinant of Vitamin C Concentrations in Apple Fruit

“…The concentration of AsA will be determined by the net rates of biosynthesis, recycling, degradation, and/or intercellular and intracellular transport, but the relative contribution of these various processes depends on several factors, including genetics, tissue type (Bulley et al, 2009), developmental stage (Hancock et al, 2007;Bulley et al, 2009;Ioannidi et al, 2009), and light intensity (Yabuta et al, 2007;Gautier et al, 2009). The biosynthesis of AsA proceeds via L-Gal (Wheeler et al, 1998), although conclusive evidence for all steps has only relatively recently become available (Conklin et al, 2006;Laing et al, 2007).…”

“…Genes involved in several of these mechanisms have been proposed to be key regulators of fruit AsA concentrations, including GDP-L-Gal phosphorylase (GGP) or vitamin c defective2 (VTC2) in kiwifruit (Actinidia deliciosa; Bulley et al, 2009Bulley et al, , 2012 as well as GDP-Man-3,5-epimerase (GME; Gilbert et al, 2009), L-Gal-1-Pphosphatase (GPP or VTC4; Ioannidi et al, 2009), and MDHAR (Stevens et al, 2007) in tomato. However, apart from GGP (Bulley et al, 2012), overexpression of these structural genes has to date had limited success in altering the fruit AsA pool (Agius et al, 2003;Bulley et al, 2009;Haroldsen et al, 2011).…”

“…However, apart from GGP (Bulley et al, 2012), overexpression of these structural genes has to date had limited success in altering the fruit AsA pool (Agius et al, 2003;Bulley et al, 2009;Haroldsen et al, 2011).…”