Abstract
Alkaloid uptake into vacuoles isolated from a Fumaria capreolata L. cell suspension culture was investigated. The uptake is carrier-mediated as shown by its substrate saturation, its sensitivity to metabolic inhibitors and especially by its exclusive preference for the (S)-forms of reticuline and scoulerine while the (R)-enantiomers which do not occur in this plant species were strictly discriminated. The carrier has a high affinity for (S)-reticuline with a K m=0.3 μM. The rate of alkaloid uptake was 6 pmol·h-1·μl-1 vacuole, and 0.03 mg alkaloid·mg-1 vacuolar protein were taken up. Transport was stimulated five-to seven-fold by ATP and was inhibited by the ATPase inhibitors N,N′-dicyclohexylcarbodiimide and 4-4′-diisothiocyanatostilbene-2,2′ disulfonic acid, as well as by the protonophore carbonyl cyanide m-chlorophenylhydrazone. A number of alkaloids did not compete with labelled (S)-reticuline for uptake into vacuoles. The uptake system is absolutely specific for alkaloids indigenous to the plant from which the vacuoles were isolated. Slight modifications of the topography of an alkaloid molecule even with full retention of its electrical charge results in its exclusion. Alkaloid efflux was also shown to be mediated by a highly specific energy-dependent carrier. These results contradict the previously proposed ion-trap mechanism for alkaloid accumulation in vacuoles. A highly specific carrier-mediated and energy-dependent proton antiport system for alkaloid uptake and release is postulated.
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Abbreviations
- CCCP:
-
carbonylcyanide m-chlorophenylhydrazone
- DCCD:
-
N,N′-dicyclohexylcarbodiimide
- DIDS:
-
4-4′-diisothiocyanatostilbene-2,2′ disulfonic acid
References
Amann, M., Nagakura, N., Zenk, M.H. (1984) (S)-Tetrahydro-protoberberine oxidase the final enzyme in protoberberine biosynthesis. Tetrahedron Lett. 25, 953–954
Amann, M., Wanner, G., Zenk, M.H. (1986) Intracellular compartmentation of two key enzymes of berberine biosynthesis in plant cell cultures, Planta 167, (in press)
Arens, H., Stöckigt, J., Weiler, E.W., Zenk, M.H. (1978) Radioimmunoassay for the determination of the indole alkaloids ajmalicine and serpentine in plants. Planta Med. 34, 37–46
Brooks, S.C., Brooks, M.M. (1941). The permeability of living cells, cited in: Levitt, J. (1956) Osmosis and permeability. In: Handbuch der Pflanzenphysiologie Bd. 3: Pflanze und Wasser, pp. 22–29, Ruhland, W., ed. Springer, Berlin Göttingen Heidelberg
Cordell, G.A. (1981) Introduction to alkaloids: A biogenetic approach, John Wiley & Sons, New York Chichester Brisbane Toronto
Deus, B., Zenk, M.H. (1982) Exploitation of plant cells for the production of natural compounds. Biotechnol. Bioeng. 24, 1965–1974
Deus-Neumann, B., Zenk, M.H. (1984) A highly selective alkaloid uptake system in vacuoles of higher plants. Planta 162, 250–260
Doll, S., Rodier, F., Willenbrink, J. (1979) Accumulation of sucrose in vacuoles isolated from red beet tissue. Planta 144, 407–411
Grob, K., Matile, Ph. (1979) Vacuolar location of glucosinolates in horseradish root cells. Plant Sci. Lett. 14, 327–335
Guern, J., Kurdjian, A., Barbier, H., Manigault, P., Quiquampoix, H., Renaudin, J.P. (1982) Difference in electrochemical potential for protons on both sides of the tonoplast and vacuolar accumulation systems. Actual. Bot. 3, 95–105
Heldt, H.W., Sauer, F. (1971) The inner membrane of the chloroplast envelope as the site of specific metabolite transport. Biochim. Biophys. Acta 234, 83–91
Höfler, H. (1947) Was lehrt die Fluoreszenzmikroskopie von der Plasmapermeabilität und Stoffspeicherung. Mikroskopie (Wien) 2, 13–29
Homeyer, B.C., Roberts, M.F. (1984) Alkaloid sequestration by Papaver somniferum latex. Z. Naturforsch. 39, Teil C, 876–881
Kaiser, G., Heber, U. (1984) Sucrose transport into vacuoles isolated from barley mesophyll protoplasts. Planta 161, 562–568
Linsmaier, E.M., Skoog, F. (1965) Organic growth factor requirements of tobacco tissue cultures. Physiol. Plant. 18, 100–127
Matile, Ph. (1976) Localization of alkaloids and mechanism of their accumulation in vacuoles of Chelidonium majus laticifers. Nova Acta Leopold., Suppl. 7, 139–156
Matile, Ph. (1978) Biochemistry and function of vacuoles. Annu. Rev. Plant Physiol. 29, 193–213
Matile, Ph. (1984) Das toxische Kompartiment der Pflanzenzelle. Naturwissenschaften 71, 18–24
Matile, Ph., Jans, B., Rickenbacher, R. (1970) Vacuoles of Chelidonium latex: lysosomal properties and accumulation of alkaloids. Biochem. Physiol. Pflanz. 161, 447–458
Müller, E., Neumann, D., Nelles, A., Bräutigam, E. (1976) Alkaloid transport and distribution in Nicotiana and Macleava. Diversity and unity. Nova Acta Leopold., Suppl. 7, 133–138
Neumann, D., Krauss, G., Hieke, M., Gröger, D. (1983) Indole alkaloid formation and storage in cell suspension cultures of Catharanthus roseus Planta Med. 48 20–23
Saunders, J.A. (1979) Investigations of vacuoles isolated from tobacco. 1. Quantitation of nicotine. Plant Physiol. 64, 74–78
Schieder, O. (1975) Regeneration von haploiden und diploiden Datura innoxia Mill. Mesophyll-Protoplasten zu Pflanzen. Z. Pflanzenphysiol. 76, 462–466
Steffens, P., Nagakura, N., Zenk, M.H. (1984) The berberine bridge-forming enzyme in tetrahydroprotoberberine biosynthesis. Tetrahedron Lett. 25, 951–952
Steffens, P., Nagakura, N., Zenk, M.H. (1985) Purification and characterization of the berberine bridge enzyme from Berberis beaniana cell cultures. Phytochemistry, 2577–2583
Sze, H. (1984) H+-translocating ATPases of the plasma membrane and tonoplast of plant cells. Physiol. Plant. 61, 683–691
Sze, H., Churchill, K.A. (1981) Mg/KCl-ATPase of plant plasma membranes is an electrogenic pump. Proc. Natl. Acad. Sci. USA 78, 5578–5582
Tanahashi, T., Zenk, M.H. (1985) Isoquinoline alkaloids from cell suspension cultures of Fumaria capreolata. Plant Cell Rep. 4, 96–99
Thom, M., Komor, E. (1984a) H+-sugar antiport as the mechanism of sugar uptake by sugarcane vacuoles. FEBS Lett. 173, 1–4
Thom, M., Komor, E. (1984b) Effect of magnesium and ATP on ATPase of sugarcane vacuoles. Planta 161, 361–365
Thom, M., Komor, E., Maretzki, A. (1982) Vacuoles of sugarcane suspension cultures. II. Characterization of sugar uptake. Plant Physiol. 69, 1320–1325
Werner, Ch., Matile, Ph. (1985) Accumulation of coumaringlucosides in vacuoles of barley mesophyll protoplasts. J. Plant Physiol. 118, 237–249
Westekemper, P., Wieczorek, U., Gueritte, F., Langlois, N., Langlois, Y., Potier, P., Zenk, M.H. (1980) Radioimmunoassay for the determination of the indole alkaloid vindoline in Catharanthus. Planta Med. 39, 24–37
Willenbrink, J., Doll, S. (1979) Characteristics of the sucrose uptake system of vacuoles isolated from red beet tissue. Kinetics and specificity of the sucrose uptake system. Planta 147, 159–162
Wink, M., Hartmann, Th. (1982) Diurnal fluctuation of quinolizidine alkaloid accumulation in legume plants and photomixotropic cell suspension cultures. Z. Naturforsch. 37 Teil C, 369–375
Wyse, R.E. (1978) Movement of sucrose from the free space into the vacuole of sugarbeet (Beta vulgaris L.) root cells. (Abstr.) Plant Physiol. 61, Suppl., 106
Zenk, M.H. (1985) Enzymology of benzylisoquinoline alkaloid formation. In: The chemistry and biology of isoquinoline alkaloids. Phillipson, J.D., Roberts, M.F., Zenk, M.H., eds. Springer, Berlin Heidelberg New York Tokyo, 240–256
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Dedicated to Professor Harry Beevers, Santa Cruz, on the occasion of his 60th birthday
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Deus-Neumann, B., Zenk, M.H. Accumulation of alkaloids in plant vacuoles does not involve an ion-trap mechanism. Planta 167, 44–53 (1986). https://doi.org/10.1007/BF00446367
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DOI: https://doi.org/10.1007/BF00446367