Presence in frog urinary bladder of proteins immunologically related to the aquaporin-CHIP

Aquaporin-CHIP, a 28 kDa channel forming protein already referred to as CHIP28, has been identified as the water channel in red blood cells as well as in mammalian renal tubule cells. Another member of the aquaporin family, WCH-CD, has been found in the apical membrane of collecting duct principal cells and may represent the ADH-sensitive water channel. The present study investigates the possible presence of CHIP28-like proteins in amphibian urinary bladder, where the presence of water channels has been postulated. For this purpose, we raised polyclonal antibodies against human erythrocyte CHIP28. Immune serum precipitated a protein of about 30 kDa from the whole homogenate of urinary epithelial cells. By Western blotting, in addition to the reaction with the 30 kDa component, the immune serum reacted with higher molecular weight components from the bladder homogenate. The 30 kDa band was detected by Western blot only in bladders having a high water permeability. Moreover, a 30 kDa protein was also recognized in frog red blood cell membranes by the anti-CHIP28 antibodies. In line with the immunoblotting studies, in immunohistofluorescence anti-CHIP28 antibodies stained frog red blood cells and urinary bladder epithelial cells. However, in whole tissue water permeability studies apical treatment with the anti-CHIP28 antibodies had no effect on either the hydrosmotic response to ADH or on the basal net water flow of the bladder. All together, these results indicate the presence in the frog red blood cells and urinary epithelium of proteins sharing immunological analogies with aquaporin-CHIP.