ABCMdb

PMID: 19619161

Gilbert L, Alhagdow M, Nunes-Nesi A, Quemener B, Guillon F, Bouchet B, Faurobert M, Gouble B, Page D, Garcia V, Petit J, Stevens R, Causse M, Fernie AR, Lahaye M, Rothan C, Baldet P
GDP-D-mannose 3,5-epimerase (GME) plays a key role at the intersection of ascorbate and non-cellulosic cell-wall biosynthesis in tomato.
Plant J. 2009 Nov;60(3):499-508. Epub 2009 Jul 8., [PubMed]

Sentences

No. Mutations Sentence Comment

30 ABCC8 p.Pro35Ser RESULTS Ascorbate content, regulation of ascorbate biosynthesis and plant phenotype are altered in GME-silenced plants Using a CaMV 35S promoter-controlled RNAi strategy targeting a DNA fragment common to the two GME genes found in tomato (SlGME1 and SlGME2), we generated 15 independent tomato primary transformants (P35S:SlgmeRNAi lines). Login to comment 43 ABCC8 p.Pro35Ser Ascorbate content was reduced by 40-60% in young fully expanded leaves and by 20-40% in fruits at 20 days post-anthesis (DPA) from P35S:SlgmeRNAi lines (Figure 2c). Login to comment 46 ABCC8 p.Pro35Ser The results of transcript expression profiling using TOM1 tomato microarray in the P35S:SlgmeRNAi transgenic and control plants are accessible at http://terry.bordeaux.inra.fr:8080/files/. Login to comment 59 ABCC8 p.Pro35Ser Gene expression, protein and ascorbate content in P35S:SlgmeRNAi transgenic and control plants. Login to comment 60 ABCC8 p.Pro35Ser (a) The relative abundances of SlGME mRNAs were determined in young leaves and fruit at 20 DPA in P35S:SlgmeRNAi plants (lines L-66, L-70 and L-108) compared to control plants. Login to comment 62 ABCC8 p.Pro35Ser (b) Comparison of specific regions of silver-stained two-dimensional gel electrophoresis gels of the proteome of fruit pericarp at 20 DPA from P35S:SlgmeRNAi line L-108 and control plants using 100 lg of proteins (see Table S4). Login to comment 63 ABCC8 p.Pro35Ser (c) Ascorbate content in young leaves and fruit at 20 DPA from P35S:SlgmeRNAi lines L-66, L-70 and L-108 and control (wild-type) plants. Login to comment 70 ABCC8 p.Pro35Ser the P35S:SlgmeRNAi L-66 and L-108 lines (Figure 4a). Login to comment 74 ABCC8 p.Pro35Ser ABCC8 p.Pro35Ser In order to investigate to what extent the growth defect observed in the P35S:SlgmeRNAi plants was related to depletion of the ascorbate pool, we germinated P35S:SlgmeRNAi (L-108) and control seeds on L-ascorbate- or L-galactose-supplemented media. Login to comment 76 ABCC8 p.Pro35Ser However, supplementation failed to enhance plant growth, and seedling height and fresh weight remained significantly lower than that of the control (Figure 4b), suggesting that plant growth defects cannot be solely attributed to the reduced ascorbate biosynthetic capacity of the P35S:SlgmeRNAi plants. Login to comment 77 ABCC8 p.Pro35Ser The GME-silenced lines have increased fragility In addition to the visual phenotypes, P35S:SlgmeRNAi plants appeared to be very fragile in the greenhouse. Login to comment 83 ABCC8 p.Pro35Ser The water potential of transgenic leaves (L-108) was reduced by 50%, but no significant changes were observed for osmotic P35S:SlgmeRNAi line-108P35S:SlgmeRNAi line-66 Line-108 Line-66 Control 16.2 ± 2.3 9.8 ± 2.1 * 6.3 ± 1.8 * Chlorophyll (mg g FW-1 ) Photosynthesis (nmolCO2 µmol photon-1 ) 1.43 ± 0.16 1.09 ± 0.13 * 1.05 ± 0.21 * Paraquat effect (% chlorophyll loss) 46.5 ± 2.8 67.9 ± 4.6 * 76.5 ± 6.5 * (a) (b) Figure 3. Login to comment 85 ABCC8 p.Pro35Ser (a) Eight-week-old P35S:SlgmeRNAi plants that were severely affected, with a strong bleaching phenotype. Login to comment 86 ABCC8 p.Pro35Ser (b) Photosynthesis was assayed in mature leaves from four P35S:SlgmeRNAi plants of each line and the control at several light intensities, and chlorophyll content was measured. Login to comment 93 ABCC8 p.Pro35Ser Growth of plantlets and fruits of P35S:SlgmeRNAi transgenic and control plants. Login to comment 94 ABCC8 p.Pro35Ser (a) Plantlets at 30 days after sowing and sections of 20 DPA fruit pericarp from P35S:SlgmeRNAi lines L-66, L-70 and L-108 and the control. Login to comment 99 ABCC8 p.Pro35Ser (b) Seedlings at 10 days after sowing of P35S:SlgmeRNAi line L-108 and the control on MS medium, or MS supplemented with 1 mM L-galactose (Gal) or 0.25 mM L-ascorbic acid (AsA). Login to comment 104 ABCC8 p.Pro35Ser Together with data from GC-MS metabolome analyses of 20 DPA fruit indicating that the most significant metabolic changes observed in P35S:SlgmeRNAi plants were linked to central and cell-wall-related metabolism (Table S2), these results suggested that the plant fragility and soft fruit traits are possibly linked to modifications of cell-wall polysaccharides. Login to comment 105 ABCC8 p.Pro35Ser Modification of non-cellulosic cell-wall polysaccharides Cell-wall composition analyses of the P35S:SlgmeRNAi lines L-66 and L-108 performed on alcohol-insoluble residue (AIR) indicated that wall preparations of both stem and 20 DPA fruit were enriched in mannose and were depleted in galactose relative to control plants (Figure 6 and Table S3). Login to comment 120 ABCC8 p.Pro35Ser Change in the composition of the polysaccharide constituents of the cell wall in the stem and 20 DPA fruit from control plants and P35S:SlgmeRNAi lines L-66 (squares) and L-108 (diamonds). Login to comment 129 ABCC8 p.Pro35Ser Fragility of the stem, fruit firmness, water (Ww) and osmotic (Ws) potentials, and turgor pressure (Wp) of the leaf in P35S:SlgmeRNAi transgenic line L-108 and control plants. Login to comment 144 ABCC8 p.Pro35Ser In the P35S:SlgmeRNAi lines, the significant decrease in GME expression and protein level reduced the capacity of the plant to produce ascorbate (Figure 2) and impaired plant development (Figure 4). Login to comment 145 ABCC8 p.Pro35Ser Under normal growth conditions, the size of the P35S:SlgmeRNAi plants and fruits correlated perfectly with the ascorbate concentration (Figure 2), the photosynthetic capacity of the plant, and its sensitivity to oxidative stress (Figure 3). Login to comment