Enzymic Assay of 10<sup>−7</sup> to 10<sup>−14</sup> Moles of Sucrose in Plant Tissues

Jones, Michael G. K.; Outlaw, William H.; Lowry, Oliver H.

doi:10.1104/pp.60.3.379

Cited by 493 publications

(286 citation statements)

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“…Samples were then kept at Ϫ20°C until analysis. Glc, Fru, and Suc were measured through the reduction of NADP by Glc-6-P dehydrogenase after the sequential addition of hexokinase, phosphoglucose isomerase, and invertase (Jones et al, 1977). The method was adapted for use on an ELISA plate, using reduced volumes and a microplate reader to measure absorbance changes.…”

Section: Measurements Of Carbohydratesmentioning

confidence: 99%

Decrease in Phosphoribulokinase Activity by Antisense RNA in Transgenic Tobacco. Relationship between Photosynthesis, Growth, and Allocation at Different Nitrogen Levels1

et al. 1999

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3 ). The activation state of PRK increased as the amount of enzyme was decreased genetically at both levels of N. At high N a 94% decrease in PRK activity had only a small effect (20%) on photosynthesis and growth. At low N a 94% decrease in PRK activity had a greater effect on leaf photosynthesis (decreased by up to 50%) and whole-plant photosynthesis (decreased by up to 35%) than at high N. These plants were up to 35% smaller than plants with higher PRK activities because they had less structural dry matter and less starch, which was decreased by 3-to 4-fold, but still accumulated to 24% to 31% of dry weight; young leaves contained more starch than older leaves in older plants. Leaves had a higher ion and water content, and specific leaf area was higher, but allocation between shoot and root was unaltered. In conclusion, low N in addition to a 94% decrease in PRK by antisense reduces the activity of PRK sufficient to diminish photosynthesis, which limits biomass production under conditions normally considered sink limited.The relationship between photosynthesis and growth is a complex one, with growth rate not being well correlated with the rate of photosynthesis on a leaf-area basis . This is because growth also depends on the investment of biomass in growing sinks and investment in leaf area (Chapin et al., 1990;). There have been many thorough investigations of the relationship between photosynthesis, growth, environment, and genotype, but they have not provided definitive information on the relationship between photosynthesis and growth. This is because of the complex interaction between factors, so it has not been easy to decide whether changes in sink growth result from changes in photosynthesis or vice versa. The elucidation of the relationship between photosynthesis and growth requires specific changes in photosynthesis or sink growth independent of other changes. Genetic manipulation of enzymes involved in photosynthesis and/or sink processes provides a means to achieve this. The use of transgenic plants with altered amounts of Rubisco revolutionized the analysis of photosynthesis and its interaction with the whole plant (Quick et al., 1991b;Stitt and Schulze, 1994). However, because Rubisco constitutes such a large proportion of the protein in a leaf (up to 40%;Woodrow and Berry, 1988), a decrease in amounts of Rubisco substantially disrupts the N balance of the plant, making it difficult to establish direct links between photosynthesis and growth and allocation than if there were a more specific alteration in the rate of photosynthesis. What is required is genetic modification of a photosynthetic enzyme that accounts for a fraction of the N content in the leaf.PRK is one such enzyme, accounting for less than 1% of the protein in a leaf (Evans, 1989). Tobacco (Nicotiana tabacum L.) plants were transformed with an inverted cDNA encoding tobacco PRK (Knight and Gray, 1994). First, it was determined that the effect of this modification on photosynthesis under standard growth-chamber conditions was minim...

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Section: Measurements Of Carbohydratesmentioning

confidence: 99%

Decrease in Phosphoribulokinase Activity by Antisense RNA in Transgenic Tobacco. Relationship between Photosynthesis, Growth, and Allocation at Different Nitrogen Levels1

et al. 1999

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“…The sucrose formed in the cytoplasm has two fates: it may be exported or it may be 'stored' within the cell. Most of the sucrose present in mesophyll cells of species which accumulate sucrose is thought to be contained within the vacuole (5,17,28). Direct studies of transport of sucrose (18) and hexose sugars (E Martinoia, G Kaiser, personal communication) into barley vacuoles suggests that the transport is passive and occurs by facilitated diffusion.…”

Section: Changes In Acid Invertase During Leaf Developmentmentioning

confidence: 99%

Biochemical Mechanism for Regulation of Sucrose Accumulation in Leaves during Photosynthesis

Huber

1989

Plant Physiol.

174

136

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It is not known why some species accumulate high concentrations of sucrose in leaves during photosynthesis while others do not. To determine the possible basis, we have studied 10 species, known to differ in the accumulation of sucrose, in terms of activities of sucrose hydrolyzing enzymes. In general, acid invertase activity decreased as leaves expanded; however, activities remaining in mature, fully expanded leaves ranged from low (<10 micromoles per gram fresh weight per hour) to very high (>100 micromoles per gram fresh weight per hour). In contrast, sucrose synthase activities were low and relatively similar among the species (4-10 micromoles per gram fresh weight per hour). Importantly, leaf sucrose concentration, measured at midaftemoon, was negatively correlated with acid invertase activity. We propose that sucrose accumulation in vacuoles of species such as soybean and tobacco is prevented by acid invertase-mediated hydrolysis. Initial attempts were made to characterize the relatively high activity of acid invertase from mature soybean leaves. Two apparent forms of the enzyme were resolved by Mono Q chromatography. The two forms had similar affinity for substrate (apparent Km [sucrose] = 3 millimolar) and did not interconvert upon rechromatography. It appeared that the loss of whole leaf invertase activity during expansion was largely the result of changes in one of the enzyme forms. Overall, the results provide a mechanism to explain why some species do not accumulate sucrose in their leaves. Some futile cycling between sucrose and hexose sugars is postulated to occur in these species, and thus, the energy cost of sucrose production may be higher than is generally thought.In many common higher plants, the principal end products of leaf photosynthesis are the carbohydrates starch and sucrose. However, species differ in the partitioning of photosynthetically fixed carbon between starch and sucrose and the extent to which sucrose normally accumulates in leaves during the photoperiod (9, 13). Accumulation of carbohydrates in leaves is critical, as they are mobilized in darkness to support continued translocation (albeit at a reduced rate) and thereby maintain heterotrophic plant parts.Plant species such as soybean that accumulate starch typically do not accumulate soluble sugars; conversely, species ' Cooperative investigations ofthe U.S. Department ofAgriculture,

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“…Soluble sugars were measured as described by Jones et al (1977). Leaf tissue was weighed and extracted three times with 80% ethanol at 80°C.…”

Section: Sugar Measurementmentioning

confidence: 99%

Monogenic Recessive Mutations Causing Both Late Floral Initiation and Excess Starch Accumulation in Arabidopsis.

et al. 1995

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A recessive Arabidopsis mutation, carbohydrate accumulation mutantl (caml), which maps to position 22.8 on chromosome 3, was identified by screening leaves of ethyl methanesulfonate-mutagenized M2 plants stained with iodine for altered starch content. lncreased starch content in leaves of the c a m l mutant was observed at the onset of flowering. This mutant also had a delayed floral initiation phenotype with more rosette leaves than the parenta1 line. In addition, activities of several enzymes associated with starch metabolism were altered in the c a m l mutant. The late-flowering mutant gigantea (gi) also manifested an elevated starch leve1 in leaves. However, not all late-flowering mutants had increased leaf starch content. Double mutants c a m l a d g l (for AJP-glucose pyrophosphorylase), c a m l pgm (for -phosphoglucomutase), and g i p g m had no observable starch in leaves but showed the late-flowering phenotype, demonstrating tThat the elevated starch content is not the cause of late floral initiation. The pleiotropic effects of c a m l and gi suggest that they may play regulatory roles in starch metabolism and floral initiation. These data suggest that starch accumulation and floral initiation may share a common regulatory pathway.

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Enzymic Assay of 10⁻⁷ to 10⁻¹⁴ Moles of Sucrose in Plant Tissues

Abstract: Procedures are desrbed for measuring sucrose in plant extracts or freeze-dried tissue in the range between 10-7 and 10-14 moles. The method is based on the destruction of pre-exbiting glucose and fructose, followed by the hydrolysis of sucrose and reduction of NADP+

Cited by 493 publications

References 9 publications

Decrease in Phosphoribulokinase Activity by Antisense RNA in Transgenic Tobacco. Relationship between Photosynthesis, Growth, and Allocation at Different Nitrogen Levels1

Decrease in Phosphoribulokinase Activity by Antisense RNA in Transgenic Tobacco. Relationship between Photosynthesis, Growth, and Allocation at Different Nitrogen Levels1

Biochemical Mechanism for Regulation of Sucrose Accumulation in Leaves during Photosynthesis

Monogenic Recessive Mutations Causing Both Late Floral Initiation and Excess Starch Accumulation in Arabidopsis.

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Enzymic Assay of 10−7 to 10−14 Moles of Sucrose in Plant Tissues

Abstract: Procedures are desrbed for measuring sucrose in plant extracts or freeze-dried tissue in the range between 10-7 and 10-14 moles. The method is based on the destruction of pre-exbiting glucose and fructose, followed by the hydrolysis of sucrose and reduction of NADP+

Cited by 493 publications

References 9 publications

Decrease in Phosphoribulokinase Activity by Antisense RNA in Transgenic Tobacco. Relationship between Photosynthesis, Growth, and Allocation at Different Nitrogen Levels1

Decrease in Phosphoribulokinase Activity by Antisense RNA in Transgenic Tobacco. Relationship between Photosynthesis, Growth, and Allocation at Different Nitrogen Levels1

Biochemical Mechanism for Regulation of Sucrose Accumulation in Leaves during Photosynthesis

Monogenic Recessive Mutations Causing Both Late Floral Initiation and Excess Starch Accumulation in Arabidopsis.

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About

Enzymic Assay of 10⁻⁷ to 10⁻¹⁴ Moles of Sucrose in Plant Tissues