J Biol Chem 1997 Jun 27;27226:16140-6.

Water and glycerol permeabilities of aquaporins 1-5 and MIP determined quantitatively by expression of epitope-tagged constructs in Xenopus oocytes.

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The goal of this study was to compare single channel water and glycerol permeabilities of mammalian aquaporins (AQP) 1-5 and the major intrinsic protein of lens fiber (MIP). Each of the six cloned cDNAs from rat was left untagged or was epitope-tagged with c-Myc or FLAG at either the N or C terminus so that results would not depend on epitope identity or location. The constructs were expressed in Xenopus oocytes for measurement of osmotic water permeability (Pf), [3H]glycerol uptake, and protein expression. Each of the 30 epitope-tagged constructs was expressed strongly at the oocyte plasma membrane. The 10-min uptake of [3H]glycerol was increased significantly (range of 4.5-8-fold over control) in oocytes expressing untagged AQP3 (GLIP) and each of the four tagged AQP3 constructs; [3H]glycerol uptake was not increased in oocytes expressing AQP1, AQP2, AQP4, AQP5, or MIP. In oocytes microinjected with 5 ng of cRNA, average Pf values (in cm/s x 10(-3)) were 0.67 +/- 0.06 (control), 19 +/- 2 (AQP1), 10 +/- 1 (AQP2), 8 +/- 2 (AQP3), 29 +/- 1 (AQP4), 10 +/- 1 (AQP5), and 1.3 +/- 0.2 (MIP), and they were relatively insensitive to the presence, identity, or location of the epitope tag. Pf values were not affected by protein kinase A or C activation. After normalization for plasma membrane expression by immunoprecipitation of microdissected plasma membranes, single channel water permeabilities (pf, referenced to the AQP1 pf of 6 x 10(-14) cm3/s) were (in cm3/s x 10(-14)) 3.3 +/- 0.2 (AQP2), 2.1 +/- 0.3 (AQP3), 24 +/- 0.6 (AQP4), 5.0 +/- 0.4 (AQP5), and 0.25 +/- 0.05 (MIP); pf values were insensitive to epitope identity and location. These results indicate very different intrinsic water permeabilities for the mammalian aquaporin homologs, with the pf value for AQP4 remarkably higher than those for the others. The pf values establish limits on aquaporin tissue densities required for physiological function and suggest significant structural and functional differences among the aquaporins.

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Species referenced: Xenopus
Genes referenced: aqp1 aqp2 aqp3 aqp4 aqp5 mip myc tnpo1