AQUAPORIN-CHIP-RELATED PROTEIN IN FROG URINARY-BLADDER - LOCALIZATIONBY CONFOCAL MICROSCOPY

Aquaporin CHIP, a 28 kDa channel forming protein, has been proposed to function as water channel in both erythrocyte and kidney proximal tub ule. Recently, we have reported that in frog urinary bladder, a model of the kidney collecting tubule, polyclonal antibodies against human e rythrocyte CHIP recognize and immunoprecipitate a 30 kDa protein from the epithelial cell homogenate. In the present work confocal fluoresce nce microscopy was used to determine the cellular and subcellular loca lization of CHIP28-like proteins in the urinary epithelium. A clear la beling of the apical border was found after Triton X-100 permeabilizat ion. The labeling was distributed throughout the apical domain and not restricted to specific domains of the membrane. The staining was also present in the deeper confocal sections where the fluorescence seems to be localized at the cellular contour. No difference in the labeling patterns was observed between resting and ADH-treated bladder. Specif icity of the staining was confirmed by the absence of the labeling pat tern when antiserum was preadsorbed on CHIP28 protein immobilized on I mmobilon P stripes. Our results suggest that CHIP-like proteins are no t proteins inserted in the apical membrane during the antidiuretic res ponse, Moreover, we do not know whether the labeling was due to the pr esence of CHIP28 itself or an as-yet-unidentified protein sharing immu nological analogies with aquaporin CHIP.