Expression of a bacterial mtlD gene in transgenic tobacco leads to production and accumulation of mannitol.

Abstract

A bacterial gene encoding mannitol-lphosphate dehydrogenase, mtlD, was engineered for expression in higher plants. Gene constructions were stably incorporated into tobacco plants. The mtlD gene was expressed and translated into a functional enzyme in tobacco, resulting in the synthesis and accumulation of mannitol, which was identified by NMR and mass spectroscopy. Mannitol concentrations exceeded 6 jumol/g (fresh weight) in the leaves and in the roots of some transformants, whereas this sugar alcohol was not detected in these organs of wild-type tobacco plants or of untransformed tobacco plants that underwent the same regeneration scheme. These experiments demonstrate that branchpoints in plant carbohydrate metabolism can be generated by which novel gene products can utilize endogenous substrates to divert metabolic energy into novel compounds. Additionally, the system described here allows for physiological studies in which the responses of wild-type and transgenic tobacco to various environmental stimuli can be compared directly. Such studies will facilitate our understanding of the roles of sugar alcohols (e.g., in stress tolerance) in higher plants.Metabolic engineering (1) was examined as an approach to gain insight into the roles of sugar alcohols or polyols in higher plants. These simple carbohydrates are widely distributed among organisms including bacteria, algae, fungi, higher plants, insects, and mammals (2-5). In plants, the importance of sugar alcohols is reflected by the estimation that metabolism of these compounds, rather than that of sugars, contributes to about 30% of the annual global primary carbon production (4). Some sugar alcohols, notably mannitol and sorbitol, are major photosynthetic products of, and can accumulate to high levels in, various higher plant species (3,4,6). These sugar alcohols are relatively widespread in higher plants, with mannitol, for example, having been detected in over 50 families (3). Mannitol and sorbitol are also translocated by some higher plant species (e.g., ref. 7), indicating a role as a storage compound. Other sugar alcohols, such as dulcitol and ribitol, are considerably less prevalent in higher plants and are apparently characteristic of species representing only a few families (3, 4).The functions of sugar alcohols in higher plants (and organisms in general) are not clear. A commonly held belief is that these compounds may confer beneficial traits on those species where they are found, rather than being simply intermediates of carbohydrate metabolism. Suggested physiological roles for sugar alcohols include osmoregulation (3,8) and service as compatible solutes (5, 9), storage of reduced carbon and energy (3), regulation of coenzymes (3, 10), and neutralization of hydroxyl radicals (11). Evidence supporting these roles for sugar alcohols has been obtained primarily with fungi and animals; only limited studies exist regarding the functions of these compounds in higher plants (12)(13)(14)(15)(16)(17).It seemed possible, by means of gene engineeri...

Dissecting the roles of osmolyte accumulation during stress

“…1). None the less, in contrast to fructan-accumulating transgenic lines which have higher levels of non-structural carbohydrates (Pilon-Smits et al 1995;Caimi et al 1997;Turk et al 1997), transgenic trehalose- (Romero et al 1997) and mannitol-accumulating plants have lower levels of sucrose (Tarczynski et al 1992;Thomas et al 1995). Whether the slight decreases observed in mtlD transformants might have a significant impact is doubtful (Thomas et al 1995;Karakas et al 1997).…”

“…Unfortunately, very few comprehensive and quantitative data are available concerning the locations of different osmolytes in cytoplasmic compartments. However, recent studies using transgenic plants have indicated that restricting trehalose (Holmström et al 1996) or mannitol (Shen et al 1997) accumulation to chloroplasts may have profoundly different effects to those mediated by the same transgene products expressed in the cytosol (Goddijn et al 1997;Romero et al 1997;Tarczynski et al 1992Tarczynski et al , 1993. Apparently, in both instances, restriction of the accumulated product to the cytosol was associated with negative effects, possibly arising from an interference with organellar metabolism.…”

“…Hexose-mediated repression of malate synthase and isocitrate lyase (Graham et al 1994) as well as α-amylase (Koch 1996) has been reported. Furthermore, sucrose levels are lower in both tobacco (Tarczynski, Jensen & Bohnert 1992) and Arabidopsis (Thomas et al 1995) mtlD-expressing lines. A drastic decline in sucrose levels was observed in vegetative tissues of Arabidopsis mtlD transformants after salt stress (Thomas et al 1995).…”

Distribution, métabolisme et rôle du sorbitol chez les plantes supérieures. Synthèse

“…L'accumulation de sorbitol est plus importante dans la racine que dans la tige (Jefferies et al, 1979). L'implication du sorbitol dans l'ajustement osmotique des feuilles de cerisier (Ranney et (Tarczynski et al, 1992 (Tarczynski et al, 1993 (Thomas et al, 1995).…”

Within-plant distribution of cotton aphid (Hemiptera: Aphididae) in cotton cultivars with colored fibers

“…The influence of the host plant on the success of a phytophagous insect species can be measured from three aspects: stimuli that lead insects to locate and choose the plant; plant conditions that lead to the initiation and maintenance of feeding; and finally the nutritional characteristics of the plant, which enable the development of the insect and its progeny (Ghovlanov 1976). Sucrose is the only sugar found in the cotton phloem sap (Tarczynski et al 1992) and A. gossypii converts most of the sucrose it ingests into oligosaccharides. Carbohydrate concentration and content vary with the age of the plant (Wells 2002).…”