QUANTITATIVE-ANALYSIS OF AQUAPORIN MESSENGER-RNA EXPRESSION IN RAT-TISSUES BY RNASE PROTECTION ASSAY

The RNase protection assay was applied to quantify mRNA expression of five principal mammalian water channels in 18 different rat tissues, a nd to determine the influence of dehydration on renal water channel ex pression. Probes consisted of labeled cRNAs transcribed from cDNA frag ments of rat CHIP28 (AQP-1, bp 238-575 of coding sequence), AQP-CD (AQ P2, bp 53-606), MIWC (AQP4, bp 235-572), CLIP (AQP3, bp 219-604), and AQP5 (bp 56-612). Results were normalized to expression of rat beta-ac tin by Quantitative densitometry of autoradiograms. CHIP28 mRNA was ex pressed strongly in heart, kidney > placenta, skeletal muscle, and uri nary bladder and detected weakly in eye, lung, trachea, spleen, liver, colon, prostate, and skin. AQP-CD was detected only in kidney. MIWC m RNA expression was highest in brain, followed by eye, trachea, lung, s tomach, kidney, and skeletal muscle. GLIP was found in eye, trachea, k idney, urinary bladder, skin, prostate, placenta, and skeletal muscle. AQP5 was detected in salivary gland, eye, lung, and trachea. An alter natively spliced form of MIWC (sMIWC) was also identified in lung and kidney by RNase protection assay, corresponding to deletion of exon 2 of MIWC. In response to dehydration (3 days, -15% body weight), renal expression of CHIP28 and MIWC were unchanged, whereas expression of AQ P-CD and CLIP were increased significantly by 2.18 +/- 0.04 and 1.36 /- 0.11 fold (SE, n = 5), respectively. These results establish quanti tative values for aquaporin transcript expression in multiple mammalia n tissues. The sensitive RNase protection assay revealed the expressio n of water channels in several tissues not studied previously or in wh ich mRNA levels were too low to detect by Northern blot analysis. The observation of CLIP up-regulation in kidney by dehydration suggests a role in the urinary concentrating mechanism.