Panel A shows bar graphs of water and solute permeability in oocytes expressing various HA-tagged aquaporin channels along with immunoblots. The y-axes represent different units of permeability, and the x-axes list the constructs. Panel B displays similar bar graphs for isolated Yolk Platelets (YPs) along with immunoblots. Panels C, D, F, and G present bar graphs of mercury inhibition and reversibility of water permeability in YPs expressing wild-type and mutant aquaporin channels. The y-axes show water uptake rates, and the x-axes list the constructs. Panels E and H show immunoblots of YPM protein extracts, with molecular mass markers on the left. The immunoblots are normalized to PDI. The graphs and immunoblots collectively illustrate the permeability properties and mercury sensitivity of various aquaporin channels.
Mammalian and piscine AQP12 channels are mercury-sensitive polytransporters. (A) Water and solute (glycerol, urea, H2O2, and ammonia) permeability of X. laevis–uninjected oocytes and oocytes expressing HA-tagged WT HsAQP1, HsAQP12, DrAqp1aa or DrAqp12, or chimeric channels in which the entire CT of the AQP1 and AQP12 paralogs from the same species were swapped. Oocytes expressing the WT or chimeric DrAqp7 (DrAqp7-HA and DrAqp7-12CT-HA, respectively) were used as positive controls for solute transport. (B) Water and solute permeability of YPs isolated from oocytes injected with the different constructs as in A. In A and B, the bottom panels show representative immunoblots of TM, PM, and YPM protein extracts from the oocytes using the α-HA antibody and the α-PDI antibody as a loading control. In A and B, data (mean ± SEM, n = 5–8 oocytes in A, and n = 4–12 biological replicates combined from two independent experiments in B) were statistically analyzed by unpaired Student’s t test (*P < 0.05; **P < 0.01; ***P < 0.001; with respect to the controls). (C, D, F, and G) Mercurial inhibition (100 µM HgCl2) of water permeability, and reversibility by βME, of YPs expressing WT HsAQP12-HA (C) or DrAqp12-HA (F), or mutant channels at potential mercury-binding Cys residues. YP-targeted HsAQP1-12CT-HA (D) or DrAqp1aa-12CT-HA (G) were also tested as positive controls for mercury inhibition. For each construct, data (mean ± SEM, n = 4 independent experiments) were statistically analyzed by one-way ANOVA followed by Tukey’s multiple comparisons test. Bars with different superscripts are significantly different (P < 0.05). (E and H) Immunoblots of the different constructs in the YPM, normalized to PDI. Molecular mass markers (kDa) are on the left. CT, C termini; WT, wild type; βME, β-mercaptoethanol. Source data are available for this figure: SourceData F3.