THE PLASMA-MEMBRANE OF ARABIDOPSIS-THALIANA CONTAINS A MERCURY-INSENSITIVE AQUAPORIN THAT IS A HOMOLOG OF THE TONOPLAST WATER CHANNEL PROTEIN TIP

Plant cells contain proteins that are members of the major intrinsic p rotein (MIP) family, an ancient family of membrane channel proteins ch aracterized by six membrane-spanning domains and two asparagine-prolin e-alanine (NPA) amino acid motifs in the two halves of the protein. We recently demonstrated that gamma-TIP, one of the MIP homologs found i n the vacuolar membrane of plant cells, is an aquaporin or water chann el protein (C. Maurel, J. Reizer, J.I. Schroeder, M.J. Chrispeels [199 3] EMBO J 12: 2241-2247). RD28, another MIP homolog in Arabidopsis tha liana, was first identified as being encoded by a turgor-responsive tr anscript. To find out if RD28 is a water channel protein, rd28 cRNA wa s injected into Xenopus laevis oocytes. Expression of RD28 caused a 10 - to 15-fold increase in the osmotic water permeability of the oocytes , indicating that the protein creates water channels in the plasma mem brane of the oocytes and is an aquaporin just like its homolog gamma-T IP. Although RD28 has several cysteine residues, its activity is not i nhibited by mercury, and in this respect it differs from gamma-TIP and all but one of the mammalian water channels that have been described. Introduction of a cysteine residue next to the second conserved NPA m otif creates a mercury-sensitive water channel, suggesting that this c onserved loop is critical to the activity of the protein. Antibodies d irected at the C terminus of RD28 were used in combination with a two- phase partitioning method to demonstrate that RD28 is located in the p lasma membrane. The protein is present in leaves and roots of well-wat ered plants, suggesting that its presence in plants does not require a specific desiccation regime. These results demonstrate that plant cel ls contain constitutively expressed aquaporins in their plasma membran es (RD28), as well as in their tonoplasts (gamma-TIP).