Functional Characterization of the Amanita muscaria Monosaccharide Transporter, AmMst1

Abstract

Fungal carbohydrate nutrition is an important aspect of ectomycorrhizal symbiosis. At the plant/fungus interface, fungal and root cortical cells compete for monosaccharides, generated from plant-derived sucrose. Therefore, the kinetic properties of the monosaccharide uptake systems are decisive for the monosaccharide yield of each partner.

For the functional characterization of a hexose transporter (AmMst1) of the ectomycorrhizal fungus Amanita muscaria, the entire cDNA was expressed in a Saccharomyces cerevisiae strain unable to take up hexoses. Uptake experiments with ¹⁴C-labelled monosaccharides resulted in K_M values of 0.46 mM for glucose and 4.20 mM for fructose, revealing a strong preference of AmMst1 for glucose as substrate. Glucose uptake by AmMst1 was strongly favoured even in the presence of a large excess of fructose. Comparable affinities of AmMst1 for glucose, 3-O-methyl glucose and mannose were obtained. In contrast, AmMst1 imported galactose with a much lower efficiency, revealing that this transporter distinguishes pyranoses by steric hindrance at the C-4 position.

While yeast contains numerous hexose transporter genes, the AmMst1 gene seems to be the main, if not the only, hexose transporter that is expressed in A. muscaria, as concluded from the comparison of hexose import properties of A. muscaria protoplasts and AmMst1 expressed in yeast.

Ectomycorrhiza - monosaccharide transporter

References

• 01 Boles,  E., and Hollenberg,  C. P.. (1997);  The molecular genetics of hexose transport in yeasts.  FEMS Microbiological Reviews. 21 85-111
• 02 Brunelli,  J. P., and Pall,  M. L.. (1993);  A series of yeast shuttle vectors for expression of cDNAs and other DNA sequences.  Yeast. 9 1309-1318
• 03 Buckhout,  T. J., and Tubbe,  A.. (1996) Structure, mechanism of catalysis and regulation of sugar transporter in plants. Photoassimilate Distribution in Plants and Crops. Source-Sink Relationship. Zamski, E. and Schaffer, A. A., eds. New York; Marcel Dekker pp. 229-260
• 04 Burgstaller,  W.. (1997);  Transport of small ions and molecules through the plasma membrane of filamenrous fungi.  Critical Reviews in Microbiol.. 23 1-46
• 05 Chen,  X. Y., and Hampp,  R.. (1993);  Sugar uptake by protoplasts of the ectomycorrhizal fungus Amanita muscaria (L. ex fr.) Hooker.  New Phytologist. 125 601-608
• 06 Dohmen,  R. J.,, Strasser,  A. W. M.,, Höner,  C. B.,, and Hollenberg,  C. P.. (1991);  An efficient transformation procedure enabling long-term storage of competent cells of various yeast genera.  Yeast. 7 691-692
• 07 Gogarten,  J. P., and Bentrup,  F.-W.. (1989);  Substrate specifity of the hexose carrier in the plasmalemma of Chenopodium suspension cells probed by transmembrane exchange diffusion.  Planta. 178 52-60
• 08 Hampp,  R., and Schaeffer,  C.. (1995) Mycorrhiza - carbohydrate and energy metabolism. Mycorrhiza. Varma, A. and Hock, B., eds. Berlin; Springer-Verlag pp. 267-296
• 09 Harley,  J. L., and Smith,  S. E.. (1993) Mycorrhizal Symbiosis. London; Academic Press
• 10 Jennings,  D. J.. (1995) The physiology of fungal nutrition. Cambridge; University Press
• 11 Komor,  E.,, Schobert,  C.,, and Cho,  B. H.. (1985);  Sugar specificity and sugar-proton interaction for the hexose-proton-symport system of Chlorella. .  European Journal of Biochemistry. 146 649-656
• 12 Lewis,  D. H., and Harley,  J. L.. (1965 a);  Carbohydrate physiology of mycorrhizal roots of beech. III. Movement of sugars between host and fungus.  New Phytologist. 64 265-275
• 13 Lewis,  D. H., and Harley,  J. L.. (1965 b);  Carbohydrate physiology of mycorrhizal roots of beech I. Identity of endogenous sugars and utilization of exogenous sugars.  New Phytologist. 64 224-237
• 14 Lewis,  D. H., and Harley,  J. L.. (1965 c);  Carbohydrate physiology of mycorrhizal roots of beech II. Utilizaton of exogenous sugars by uninfected and mycorrhizal roots.  New Phytologist. 64 238-255
• 15 Melin,  E., and Nilsson,  H.. (1957);  Transport of ¹⁴C-labelled photosynthate to the fungal associate of pine mycorrhiza.  Svensk Botanisk Tidskrift. 51 166-186
• 16 Nehls,  U.,, Wiese,  J.,, Guttenberger,  M.,, and Hampp,  R.. (1998);  Carbon allocation in ectomycorrhizas: Identification and expression analysis of an Amanita muscaria monosaccharide transporter.  Molecular Plant Microbe Interaction. 11 167-176
• 17 Reifenberger,  E.,, Freidel,  K.,, and Ciriacy,  M.. (1995);  Identification of novel HXT genes in Saccharomyces cerevisiae reveals the impact of individual hexose transporters on glycolytic flux.  Molecular Microbiology. 16 157-167
• 18 Salzer,  P., and Hager,  A.. (1991);  Sucrose utilization of the ectomycorrhizal fungi Amanita muscaria and Hebeloma crustuliniforme depends on the cell wall-bound invertase activity of their host Picea abies. .  Botanica Acta. 104 439-445
• 19 Salzer,  P., and Hager,  A.. (1993);  Characterization of wall-bound invertase isoforms of Picea abies cells and regulation by ectomycorrhizal fungi.  Physiologia Plantarum. 88 52-59
• 20 Schaeffer,  C.,, Wallenda,  T.,, Guttenberger,  M.,, and Hampp,  R.. (1995);  Acid invertase in mycorrhizal and non-mycorrhizal roots of Norway spruce (Picea abies [L.] Karst.) seedlings.  New Phytologist. 129 417-424
• 21 Smith,  S. E., and Read,  D. J.. (1997) Mycorrhizal Symbiosis. 2nd ed. London; Academic Press pp. 1171-1184
• 22 Tubbe,  A., and Buckhout,  T. J.. (1992);  In vitro analysis of the H⁺-hexose symporter of the plasma membrane of sugarbeets (Beta vulgaris L.).  Plant Physiology. 99 945-951
• 23 Wallenda,  T.. (1996) Untersuchungen zur Physiologie der Pilzpartner von Ektomykorrhizen der Fichte. Tübingen, Germany; Ph.D. thesis. Universität Tübingen

Section Editor: U. Lüttge