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One-year-old grapevines (Vitis labrusca L. `Concord') were supplied twice weekly for 5 weeks with 0, 5, 10, 15, or 20 mm nitrogen (N) in a modified Hoagland's solution to generate a wide range of leaf N status. Both light-saturated CO₂ assimilation at ambient CO₂ and at saturating CO₂ increased curvilinearly as leaf N increased. Although stomatal conductance showed a similar response to leaf N as CO₂ assimilation, calculated intercellular CO₂ concentrations decreased. On a leaf area basis, activities of key enzymes in the Calvin cycle, ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), NADP-glyceraldehyde-3-phosphate dehydrogenase (GAPDH), phosphoribulokinase (PRK), and key enzymes in sucrose and starch synthesis, fructose-1,6-bisphosphatase (FBPase), sucrose phosphate synthase (SPS), and ADP-glucose pyrophosphorylase (AGPase), increased linearly with increasing leaf N content. When expressed on a leaf N basis, activities of the Calvin cycle enzymes increased with increasing leaf N, whereas activities of FBPase, SPS, and AGPase did not show significant change. As leaf N increased, concentrations of glucose-6-phosphate (G6P), fructose-6-phosphate (F6P), and 3-phosphoglycerate (PGA) increased curvilinearly. The ratio of G6P/F6P remained unchanged over the leaf N range except for a significant drop at the lowest leaf N. Concentrations of glucose, fructose, and sucrose at dusk increased linearly with increasing leaf N, and there was no difference between predawn and dusk measurements. As leaf N increased, starch concentration increased linearly at dusk, but decreased linearly at predawn. The calculated carbon export from starch degradation during the night increased with increasing leaf N. These results showed that 1) grapes leaves accumulated less soluble carbohydrates under N-limitation; 2) the elevated starch level in low N leaves at predawn was the result of the reduced carbon export from starch degradation during the night; and 3) the reduced capacity of CO₂ assimilation in low N leaves was caused by the coordinated decreases in the activities of key enzymes involved in CO₂ assimilation as a result of direct N limitation, not by the indirect feedback repression of CO₂ assimilation via sugar accumulation.

To determine the cause of zonal chlorosis of `Honeycrisp' apple leaves, we compared CO₂ assimilation, carbohydrate metabolism, xanthophyll cycle and the antioxidant system between chlorotic leaves and normal leaves. Chlorotic leaves accumulated higher levels of non-structural carbohydrates, particularly starch, sorbitol, sucrose, and fructose at both dusk and predawn, and no difference was found in total non-structural carbohydrates between predawn and dusk. CO₂ assimilation and the key enzymes in the Calvin cycle, ribulose 1,5-bisphosphate carboxylase/oxygenase, NADP-glyceraldehyde-3-phosphate dehydrogenase, phosphoribulokinase, stromal fructose-1,6-bisphosphatase, and enzymes in starch and sorbitol synthesis, ADP-glucose pyrophosphorylase, cytosolic fructose-1,6-bisphosphatase, and aldose 6-phosphate reductase were significantly lower in chlorotic leaves than in normal leaves. However, sucrose phosphate synthase activity was higher in chlorotic leaves. Thermal dissipation of excitation energy was enhanced in chlorotic leaves under full sun, lowering the efficiency of excitation energy transfer to PSII reaction centers. This was accompanied by a corresponding increase in both xanthophyll cycle pool size (on a chlorophyll basis) and conversion of violaxanthin to antheraxanthin and zeaxanthin. The antioxidant system was up-regulated in chlorotic leaves in response to the increased generation of reactive oxygen species. These findings support the hypothesis that phloem loading and/or transport is partially or completely blocked in chlorotic leaves, and that excessive accumulation of non-structural carbohydrates may cause feedback suppression of CO₂ assimilation via direct interference with chloroplast function and/or indirect repression of photosynthetic enzymes.

To determine the cause of a characteristic zonal chlorosis of `Honeycrisp' apple (Malus ×domestica Borkh.) leaves, we compared CO₂ assimilation, carbohydrate metabolism, the xanthophyll cycle and the antioxidant system between chlorotic leaves and normal leaves. Chlorotic leaves accumulated higher levels of nonstructural carbohydrates, particularly starch, sorbitol, sucrose, and fructose at both dusk and predawn, and no difference was found in total nonstructural carbohydrates between predawn and dusk. This indicates that carbon export was inhibited in chlorotic leaves. CO₂ assimilation and the key enzymes in the Calvin cycle, ribulose 1,5-bisphosphate carboxylase/oxygenase, NADP-glyceraldehyde-3-phosphate dehydrogenase, phosphoribulokinase, stromal fructose-1,6-bisphosphatase, and the key enzymes in starch and sorbitol synthesis, ADP-glucose pyrophosphorylase, cytosolic fructose-1,6-bisphosphatase, and aldose 6-phosphate reductase were significantly lower in chlorotic leaves than in normal leaves. However, sucrose phosphate synthase activity was higher in chlorotic leaves. In response to a reduced demand for photosynthetic electron transport, thermal dissipation of excitation energy (measured as nonphotochemical quenching of chlorophyll fluorescence) was enhanced in chlorotic leaves under full sun, lowering the efficiency of excitation energy transfer to PSII reaction centers. This was accompanied by a corresponding increase in both xanthophyll cycle pool size (on a chlorophyll basis) and conversion of violaxanthin to antheraxanthin and zeaxanthin. The antioxidant system, including superoxide dismutase and ascorbate peroxidase and the ascorbate pool and glutathione pool, was up-regulated in chlorotic leaves in response to the increased generation of reactive oxygen species via photoreduction of oxygen. These findings support the hypothesis that phloem loading and/or transport is partially or completely blocked in chlorotic leaves, and that excessive accumulation of nonstructural carbohydrates may cause feedback suppression of CO₂ assimilation via direct interference with chloroplast function and/or indirect repression of photosynthetic enzymes.

Own-rooted one-year-old `Concord' grapevines were fertigated twice weekly for 11 weeks with 1, 10, 20, 50, OR 100 μmol iron (Fe) from ferric ethylenediamine di (o-hydroxyphenylacetic) acid in a complete nutrient solution. As Fe supply increased, leaf total Fe content did not change, whereas active Fe (extracted by 2, 2'-dipyridyl) and total chlorophyll content increased curvilinearly. CO₂ assimilation and stomatal conductance increased curvilinearly with increasing active Fe, whereas intercellular CO₂ concentrations decreased linearly. Activities of key Calvin cycle enzymes, Rubisco, NADP-glyceraldehyde-3-phosphate dehydrogenase, phosphoribulokinase, stromal fructose-1,6-bisphosphatase (FBPase), and a key enzyme in sucrose synthesis, cytosolic FBPase, all increased linearly with increasing active Fe. No difference was found in the activities of ADP-glucose pyrophosphorylase and sucrose phosphate synthase of leaves between the lowest and the highest treatments, whereas slightly lower activities were observed in the middle Fe treatments. Content of 3-phosphoglycerate increased curvilinearly with increased active Fe, whereas glucose-6-phosphate and fructose-6-phosphate did not change. Glucose, fructose, sucrose, starch, and total non-structural carbohydrates at both dusk and pre-dawn increased with increasing active Fe. Carbon export from starch breakdown during the night, calculated as the difference between dusk and predawn levels, increased as active Fe increased. In conclusion, Fe limitation reduces the activities of Rubisco and other photosynthetic enzymes, and hence CO₂ assimilation capacity. Fe-deficient grapevines have lower concentrations of non-structural carbohydrates in source leaves, and therefore, are source limited.

Own-rooted 1-year-old `Concord' grapevines (Vitis labruscana Bailey) were fertigated twice weekly for 11 weeks with 1, 10, 20, 50, or 100 μm iron (Fe) from ferric ethylenediamine di (o-hydroxyphenylacetic) acid (Fe-EDDHA) in a complete nutrient solution. As Fe supply increased, leaf total Fe content did not show a significant change, whereas active Fe (extracted by 2,2′-dipyridyl) content increased curvilinearly. Chlorophyll (Chl) content increased as Fe supply increased, with a greater response at the lower Fe rates. Chl a: b ratio remained relatively constant over the range of Fe supply, except for a slight increase at the lowest Fe treatment. Both CO₂ assimilation and stomatal conductance increased curvilinearly with increasing leaf active Fe, whereas intercellular CO₂ concentrations decreased linearly. Activities of key enzymes in the Calvin cycle, ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), NADP-glyceraldehyde-3-phosphate dehydrogenase (GAPDH), phosphoribulokinase (PRK), stromal fructose-1,6-bisphosphatase (FBPase), and a key enzyme in sucrose synthesis, cytosolic FBPase, all increased linearly with increasing leaf active Fe. No significant difference was found in the activities of ADP-glucose pyrophosphorylase (AGPase) and sucrose phosphate synthase (SPS) of leaves between the lowest and the highest Fe treatments, whereas slightly lower activities of AGPase and SPS were observed in the other three Fe treatments. Content of 3-phosphoglycerate (PGA) increased curvilinearly with increasing leaf active Fe, whereas glucose-6-phosphate (G6P), fructose-6-phosphate (F6P), and the ratio of G6P: F6P remained unchanged over the range of Fe supply. Concentrations of glucose, fructose, sucrose, starch, and total nonstructural carbohydrates (TNC) at both dusk and predawn increased with increasing leaf active Fe. Concentrations of starch and TNC at any given leaf active Fe content were higher at dusk than at predawn, but both glucose and fructose showed the opposite trend. No difference in sucrose concentration was found at dusk or predawn. The export of carbon from starch breakdown during the night, calculated as the difference between dusk and predawn measurements, increased as leaf active Fe content increased. The ratio of starch to sucrose at both dusk and predawn also increased with increasing leaf active Fe. In conclusion, Fe limitation reduces the activities of Rubisco and other photosynthetic enzymes, and hence CO₂ assimilation capacity. Fe-deficient grapevines have lower concentrations of nonstructural carbohydrates in source leaves and, therefore, are source limited.

The response to reduced light intensity of two contrasting cultivars Puma Sunny (shade intolerant) and Gongzi (shade tolerant) was characterized in terms of plant height, the root/shoot ratio, photosynthetic capacity, and the morphology and ultrastructure of their chloroplasts and phloem companion cells. The initial response to shading of cultivar Puma Sunny plants was to extend their stems, and while the equivalent response of cultivar Gongzi was less marked. Shading depressed the maximum relative electron transport rate (rETR) in both cultivars, and while the efficiency of light capture in cultivar Puma Sunny was compromised by shading, this was not the case for cultivar Gongzi. Low levels of incident light inhibited the formation of starch grains in the chloroplast and increased the volume of interspace between the grana lamellae. In cultivar Puma Sunny, but less so in cultivar Gongzi, the chloroplasts became more slender and the stroma lamellae more swollen. Adjusting chloroplast morphology by developing extra layers of grana lamellae and maintaining the integrity of the phloem companion cells are both adaptations which help make ‘Gongzi’ a more shade-tolerant cultivar.

Loquat (Eriobotrya japonica) is a model fruit for investigating flesh lignification during storage and response to chilling injury. However, the investigations of enzymes and coding genes and loquat fruit lignification under low-temperature storage are still limited. Here, the activity and transcript levels of up-stream enzymes of the phenylpropanoid pathway, including l-phenylalanine ammonia-lyase (PAL), cinnamate 4-hydroxylase (C4H), and 4-coumarate:coenzyme A ligase (4CL), were investigated. The results indicated that activity of these enzymes was positively correlated with loquat fruit lignification and suppression of these increases by heat treatment (HT) and low-temperature conditioning (LTC) significantly alleviated loquat fruit lignification. Coding genes for these enzymes were subsequently isolated based on information from an RNA-seq database and expression of Ej4CL1 was found to be the most responsive to low temperature and inhibition by HT and LTC treatment, whereas the other genes were less responsive to these treatments. Furthermore, function of Ej4CL1 was analyzed by transient overexpression in tobacco leaves, where it stimulated lignin accumulation. Ej4CL1 may be a key candidate that involved in CI-related loquat fruit lignification.

Volatile compounds have a tremendous impact on fruit quality. We evaluated the volatile compound profiles of ripening wild apple fruit (10 Malus baccata accessions and three Malus prunifolia accessions) in the National Field Genebank for Hardy Fruits at Gongzhuling, China. Alcohols, esters, aldehydes, terpenes, hydrocarbons, ethers, heterocycles, carboxylic acids, and ketones were detected in the M. baccata and M. prunifolia fruit, with the first four being the main volatile compounds present. Of the 92 volatiles detected, esters were the most diverse (49 compounds). This wide range of abundant volatile compounds suggests that M. prunifolia is a good resource for breeding apple cultivars with novel and interesting flavors. The M. baccata accession ‘Zhaai Shandingzi’ and the M. prunifolia accession ‘Bai Haitang’ had the widest range of volatile compounds and the highest volatile compound contents of the accessions examined, and will therefore be good breeding materials for developing commercial lines with enhanced flavor and for widening the genetic diversity. The number of different ester compounds present was significantly positively correlated (r = 0.877) with the cube root of the weight of an individual ripe fruit. Principal component analysis (PCA) showed that the contents of ester compounds could be used to distinguish between M. baccata and M. prunifolia species. Therefore, ester compounds could be used as a reference of parental choice in apple breeding.