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Expression profiling of ascorbic acid-related genes during tomato fruit development and ripening and in response to stress conditions.

Ioannidi E, Kalamaki MS, Engineer C, Pateraki I, Alexandrou D, Mellidou I, Giovannonni J, Kanellis AK - J. Exp. Bot. (2009)

Bottom Line: L-ascorbate (the reduced form of vitamin C) participates in diverse biological processes including pathogen defence mechanisms, and the modulation of plant growth and morphology, and also acts as an enzyme cofactor and redox status indicator.Important aspects of the hypoxic and post-anoxic response in tomato fruit are discussed.The data suggest that L-galactose-1-phosphate phosphatase could play an important role in regulating ascorbic acid accumulation during tomato fruit development and ripening.

View Article: PubMed Central - PubMed

Affiliation: Group of Biotechnology of Pharmaceutical Plants, Division of Pharmacognosy-Pharmacology, Department of Pharmaceutical Sciences, Aristotle University of Thessaloniki, 541 24 Thessaloniki, Greece.

ABSTRACT
L-ascorbate (the reduced form of vitamin C) participates in diverse biological processes including pathogen defence mechanisms, and the modulation of plant growth and morphology, and also acts as an enzyme cofactor and redox status indicator. One of its chief biological functions is as an antioxidant. L-ascorbate intake has been implicated in the prevention/alleviation of varied human ailments and diseases including cancer. To study the regulation of accumulation of this important nutraceutical in fruit, the expression of 24 tomato (Solanum lycopersicon) genes involved in the biosynthesis, oxidation, and recycling of L-ascorbate during the development and ripening of fruit have been characterized. Taken together with L-ascorbate abundance data, the results show distinct changes in the expression profiles for these genes, implicating them in nodal regulatory roles during the process of L-ascorbate accumulation in tomato fruit. The expression of these genes was further studied in the context of abiotic and post-harvest stress, including the effects of heat, cold, wounding, oxygen supply, and ethylene. Important aspects of the hypoxic and post-anoxic response in tomato fruit are discussed. The data suggest that L-galactose-1-phosphate phosphatase could play an important role in regulating ascorbic acid accumulation during tomato fruit development and ripening.

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Expression of AA biosynthetic genes during tomato fruit ripening. (A) RNA blot analysis of AA biosynthetic genes. (B) The main and alternative AA biosynthetic routes. (C) Ascorbate levels during development and ripening. Total RNA (20 μg) isolated from different stages ranging from anthesis, the first, second, and third weeks from anthesis (FL) (W1, W2, and W3), IG (immature green), MG (mature green), BR (breaker), TR (turning), PK (pink), LR (light red), R (red), to RR (ripe red) was fractionated in formaldehyde denaturing agarose gels, transferred to nylon membranes, stained with 0.04% methylene blue (to observe the equality of loading), and hybridized with radiolabelled probes for AA biosynthetic genes. All experiments were performed in triplicate.
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fig1: Expression of AA biosynthetic genes during tomato fruit ripening. (A) RNA blot analysis of AA biosynthetic genes. (B) The main and alternative AA biosynthetic routes. (C) Ascorbate levels during development and ripening. Total RNA (20 μg) isolated from different stages ranging from anthesis, the first, second, and third weeks from anthesis (FL) (W1, W2, and W3), IG (immature green), MG (mature green), BR (breaker), TR (turning), PK (pink), LR (light red), R (red), to RR (ripe red) was fractionated in formaldehyde denaturing agarose gels, transferred to nylon membranes, stained with 0.04% methylene blue (to observe the equality of loading), and hybridized with radiolabelled probes for AA biosynthetic genes. All experiments were performed in triplicate.

Mentions: In plants, AA is highly abundant and accumulates in intracellular concentrations of 2–25 mM (Davey et al., 2000). It is primarily known for its antioxidant properties, but it also acts as a cofactor for various enzymes and further contributes to the regulation of cell division and expansion (Smirnoff and Wheeler, 2000). It is essential for plant growth (Alhagdow et al., 2007; Dowdle et al., 2007), participates in stress resistance, and seems to control flowering time and the commencement of senescence (Davey et al., 2000). In addition, AA and its oxidized form dehydroascorbic acid (DHA) can act as signalling agents (Pastori et al., 2003; Fotopoulos et al., 2008) participating in the interaction with the environment, for instance to ozone (Sanmartin et al., 2003), pathogens and oxidizing agents (Fotopoulos et al., 2006), and water loss (Fotopoulos et al., 2008). The benefits that arise from the increase of AA production in plants are profound. However, until recently, many aspects of AA biosynthesis, metabolism, and function remained unsettled (Davey et al., 2000; Smirnoff and Wheeler, 2000; Smirnoff et al., 2001; Conklin, 2001; Hancock and Viola, 2005; Ishikawa et al., 2006). The recent deciphering of the pathway of plant L-AA biosynthesis (see Fig. 1) (Wheeler, et al., 1998; Ishikawa et al., 2006), as well as the cloning of various enzymes of the pathway, has refocused attention on this important micronutrient, particularly as a potential route to improve nutritional quality and plant abiotic stress resistance. Ascorbate is synthesized from mannose-6-phosphate via GDP-mannose and GDP-L-galactose (GDP-L-Gal; Wheeler et al., 1998). The first part of the pathway also provides precursors for the synthesis of certain cell wall polysaccharides containing mannose, L-Gal, and L-fucose. The second part of the pathway is committed to ascorbate synthesis (Hancock and Viola, 2005). Free L-Gal is released from GDP-L-Gal via the action of GDP-L-Gal phosphorylase and L-Gal-1-phosphate phosphatase (GPP), and then is oxidized by L-Gal dehydrogenase to form L-galactono-1,4-lactone. L-Galactono-1,4-lactone is oxidized to ascorbate by L-galactono-1,4-lactone dehydrogenase which is located on the inner mitochondrial membrane. Apart from the last step, it is likely that all the other enzymes are cytosolic.


Expression profiling of ascorbic acid-related genes during tomato fruit development and ripening and in response to stress conditions.

Ioannidi E, Kalamaki MS, Engineer C, Pateraki I, Alexandrou D, Mellidou I, Giovannonni J, Kanellis AK - J. Exp. Bot. (2009)

Expression of AA biosynthetic genes during tomato fruit ripening. (A) RNA blot analysis of AA biosynthetic genes. (B) The main and alternative AA biosynthetic routes. (C) Ascorbate levels during development and ripening. Total RNA (20 μg) isolated from different stages ranging from anthesis, the first, second, and third weeks from anthesis (FL) (W1, W2, and W3), IG (immature green), MG (mature green), BR (breaker), TR (turning), PK (pink), LR (light red), R (red), to RR (ripe red) was fractionated in formaldehyde denaturing agarose gels, transferred to nylon membranes, stained with 0.04% methylene blue (to observe the equality of loading), and hybridized with radiolabelled probes for AA biosynthetic genes. All experiments were performed in triplicate.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2651456&req=5

fig1: Expression of AA biosynthetic genes during tomato fruit ripening. (A) RNA blot analysis of AA biosynthetic genes. (B) The main and alternative AA biosynthetic routes. (C) Ascorbate levels during development and ripening. Total RNA (20 μg) isolated from different stages ranging from anthesis, the first, second, and third weeks from anthesis (FL) (W1, W2, and W3), IG (immature green), MG (mature green), BR (breaker), TR (turning), PK (pink), LR (light red), R (red), to RR (ripe red) was fractionated in formaldehyde denaturing agarose gels, transferred to nylon membranes, stained with 0.04% methylene blue (to observe the equality of loading), and hybridized with radiolabelled probes for AA biosynthetic genes. All experiments were performed in triplicate.
Mentions: In plants, AA is highly abundant and accumulates in intracellular concentrations of 2–25 mM (Davey et al., 2000). It is primarily known for its antioxidant properties, but it also acts as a cofactor for various enzymes and further contributes to the regulation of cell division and expansion (Smirnoff and Wheeler, 2000). It is essential for plant growth (Alhagdow et al., 2007; Dowdle et al., 2007), participates in stress resistance, and seems to control flowering time and the commencement of senescence (Davey et al., 2000). In addition, AA and its oxidized form dehydroascorbic acid (DHA) can act as signalling agents (Pastori et al., 2003; Fotopoulos et al., 2008) participating in the interaction with the environment, for instance to ozone (Sanmartin et al., 2003), pathogens and oxidizing agents (Fotopoulos et al., 2006), and water loss (Fotopoulos et al., 2008). The benefits that arise from the increase of AA production in plants are profound. However, until recently, many aspects of AA biosynthesis, metabolism, and function remained unsettled (Davey et al., 2000; Smirnoff and Wheeler, 2000; Smirnoff et al., 2001; Conklin, 2001; Hancock and Viola, 2005; Ishikawa et al., 2006). The recent deciphering of the pathway of plant L-AA biosynthesis (see Fig. 1) (Wheeler, et al., 1998; Ishikawa et al., 2006), as well as the cloning of various enzymes of the pathway, has refocused attention on this important micronutrient, particularly as a potential route to improve nutritional quality and plant abiotic stress resistance. Ascorbate is synthesized from mannose-6-phosphate via GDP-mannose and GDP-L-galactose (GDP-L-Gal; Wheeler et al., 1998). The first part of the pathway also provides precursors for the synthesis of certain cell wall polysaccharides containing mannose, L-Gal, and L-fucose. The second part of the pathway is committed to ascorbate synthesis (Hancock and Viola, 2005). Free L-Gal is released from GDP-L-Gal via the action of GDP-L-Gal phosphorylase and L-Gal-1-phosphate phosphatase (GPP), and then is oxidized by L-Gal dehydrogenase to form L-galactono-1,4-lactone. L-Galactono-1,4-lactone is oxidized to ascorbate by L-galactono-1,4-lactone dehydrogenase which is located on the inner mitochondrial membrane. Apart from the last step, it is likely that all the other enzymes are cytosolic.

Bottom Line: L-ascorbate (the reduced form of vitamin C) participates in diverse biological processes including pathogen defence mechanisms, and the modulation of plant growth and morphology, and also acts as an enzyme cofactor and redox status indicator.Important aspects of the hypoxic and post-anoxic response in tomato fruit are discussed.The data suggest that L-galactose-1-phosphate phosphatase could play an important role in regulating ascorbic acid accumulation during tomato fruit development and ripening.

View Article: PubMed Central - PubMed

Affiliation: Group of Biotechnology of Pharmaceutical Plants, Division of Pharmacognosy-Pharmacology, Department of Pharmaceutical Sciences, Aristotle University of Thessaloniki, 541 24 Thessaloniki, Greece.

ABSTRACT
L-ascorbate (the reduced form of vitamin C) participates in diverse biological processes including pathogen defence mechanisms, and the modulation of plant growth and morphology, and also acts as an enzyme cofactor and redox status indicator. One of its chief biological functions is as an antioxidant. L-ascorbate intake has been implicated in the prevention/alleviation of varied human ailments and diseases including cancer. To study the regulation of accumulation of this important nutraceutical in fruit, the expression of 24 tomato (Solanum lycopersicon) genes involved in the biosynthesis, oxidation, and recycling of L-ascorbate during the development and ripening of fruit have been characterized. Taken together with L-ascorbate abundance data, the results show distinct changes in the expression profiles for these genes, implicating them in nodal regulatory roles during the process of L-ascorbate accumulation in tomato fruit. The expression of these genes was further studied in the context of abiotic and post-harvest stress, including the effects of heat, cold, wounding, oxygen supply, and ethylene. Important aspects of the hypoxic and post-anoxic response in tomato fruit are discussed. The data suggest that L-galactose-1-phosphate phosphatase could play an important role in regulating ascorbic acid accumulation during tomato fruit development and ripening.

Show MeSH
Related in: MedlinePlus