Abstract
Xenopus oocytes are a versatile expression system particularly suited for membrane transporters and channels. Oocytes have little background activity and therefore offer a very high signal-to-noise ratio for transporter and channel characterization. This chapter provides an overview of the basic methods used for the analysis of membrane transporters in this system, including preparation of oocytes, assays of transport activity, protocols for immunostaining and fluorescence microscopy, and other assays to study surface expression.
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References
Stegen, C., Matskevich, I., Wagner, C.A., Paulmichl, M., Lang, F., and Broer, S. (2000) Swelling-induced taurine release without chloride channel activity in Xenopus laevis oocytes expressing anion channels and transporters. Biochim. Biophys. Acta. 1467, 91–100.
Zeuthen, T., Meinild, A.K., Loo, D.D., Wright, E.M., and Klaerke, D.A. (2001) Isotonic transport by the Na+-glucose cotransporter SGLT1 from humans and rabbit. J. Physiol. 531, 631–644.
Weber, W. (1999) Ion currents of Xenopus laevis oocytes: state of the art. Biochim. Biophys. Acta. 1421, 213–233.
Broer, S., Schneider, H.P., Broer, A., Rahman, B., Hamprecht, B., and Deitmer, J.W. (1998) Characterization of the monocarboxylate transporter 1 expressed in Xenopus laevis oocytes by changes in cytosolic pH. Biochem. J. 333, 167–174.
Broer, A., Hamprecht, B., and Broer, S. (1998) Discrimination of two amino acid transport activities in 4F2 heavy chain-expressing Xenopus laevis oocytes. Biochem. J. 333, 549–554.
Gurdon, J.B., Lane, C.D., Woodland, H.R., and Marbaix, G. (1971) Use of frog eggs and oocytes for the study of messenger RNA and its translation in living cells. Nature 233, 177–182.
Mishina, M., Kurosaki, T., Tobimatsu, T., Morimoto, Y., Noda, M., Yamamoto, T., Terao, M., Lindstrom, J., Takahashi, T., Kuno, M., et al. (1984) Expression of functional acetylcholine receptor from cloned cDNAs. Nature 307, 604–608.
Barnard, E.A., Miledi, R., and Sumikawa, K. (1982) Translation of exogenous messenger RNA coding for nicotinic acetylcholine receptors produces functional receptors in Xenopus oocytes. Proc. R Soc. Lond. B Biol. Sci. 215, 241–246.
Zampighi, G.A., Kreman, M., Boorer, K. J., Loo, D. D., Bezanilla, F., Chandy, G., Hall, J. E., and Wright, E. M. (1995) A method for determining the unitary functional capacity of cloned channels and transporters expressed in Xenopus laevis oocytes. J. Membr. Biol. 148, 65–78.
Miledi, R., Eusebi, F., Martinez-Torres, A., Palma, E., and Trettel, F. (2002) Expression of functional neurotransmitter receptors in Xenopus oocytes after injection of human brain membranes. Proc. Natl. Acad. Sci. USA 99, 13238–13242.
Miledi, R., Palma, E., and Eusebi, F. (2006) Microtransplantation of neurotransmitter receptors from cells to Xenopus oocyte membranes: new procedure for ion channel studies. Methods Mol. Biol. 322, 347–355.
Maller, J.L., Butcher, F.R., and Krebs, E.G. (1979) Early effect of progesterone on levels of cyclic adenosine 3':5'-monophosphate in Xenopus oocytes. J. Biol. Chem. 254, 579–582.
Uezono, Y., Bradley, J., Min, C., McCarty, N.A., Quick, M., Riordan, J.R., Chavkin, C., Zinn, K., Lester, H.A., and Davidson, N. (1993) Receptors that couple to 2 classes of G proteins increase cAMP and activate CFTR expressed in Xenopus oocytes. Receptors Channels 1, 233–241.
Landau, E.M. and Blitzer, R.D. (1994) Chloride current assay for phospholipase C in Xenopus oocytes. Methods Enzymol. 238, 140–154.
Guo, Z., Liliom, K., Fischer, D.J., Bathurst, I.C., Tomei, L.D., Kiefer, M.C., and Tigyi, G. (1996) Molecular cloning of a high-affinity receptor for the growth factor-like lipid mediator lysophosphatidic acid from Xenopus oocytes. Proc. Natl. Acad. Sci. USA 93, 14367–14372.
Ferrell, J.E., Jr. (1999) Xenopus oocyte maturation: new lessons from a good egg. Bioessays 21, 833–842.
Wagner, C.A., Ott, M., Klingel, K., Beck, S., Melzig, J., Friedrich, B., Wild, K. N., Broer, S., Moschen, I., Albers, A., Waldegger, S., Tummler, B., Egan, M.E., Geibel, J.P., Kandolf, R., and Lang, F. (2001) Effects of the serine/threonine kinase SGK1 on the epithelial Na(+) channel (ENaC) and CFTR: implications for cystic fibrosis. Cell Physiol. Biochem. 11, 209–218.
Trotti, D., Peng, J.B., Dunlop, J., and Hediger, M.A. (2001) Inhibition of the glutamate transporter EAAC1 expressed in Xenopus oocytes by phorbol esters. Brain Res. 914, 196–203.
Levis, R.A. and Rae, J.L. (1992) Constructing a patch clamp setup. Methods Enzymol. 207, 14–66.
Corey, J.L., Davidson, N., Lester, H.A., Brecha, N., and Quick, M.W. (1994) Protein kinase C modulates the activity of a cloned gamma-aminobutyric acid transporter expressed in Xenopus oocytes via regulated subcellular redistribution of the transporter. J. Biol. Chem. 269, 14759–14767.
Chubb, S., Kingsland, A.L., Broer, A., and Broer, S. (2006) Mutation of the 4F2 heavy-chain carboxy terminus causes y+ LAT2 light-chain dysfunction. Mol. Membr. Biol. 23, 255–267.
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Bröer, S. (2010). Xenopus laevis Oocytes. In: Yan, Q. (eds) Membrane Transporters in Drug Discovery and Development. Methods in Molecular Biology, vol 637. Humana Press. https://doi.org/10.1007/978-1-60761-700-6_16
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DOI: https://doi.org/10.1007/978-1-60761-700-6_16
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