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Tissue-specific expression of mRNA encoding rat kidney water channel CHIP28k by in situ hybridization
H Hasegawa1, R Zhang, A Dohrman
1Department of Medicine, University of California, San Francisco 94143-0532.
Abstract:
The tissue distribution of mRNA encoding rat kidney water channel CHIP28k was determined by in situ hybridization. cDNA encoding rat kidney CHIP28k was isolated by homology to human erythrocyte CHIP28 (G. M. Preston and P. Agre. Proc. Natl. Acad. Sci. USA 88: 11110-11114, 1991) and used to construct 155-base 35S-labeled cRNA sense and antisense probes corresponding to base pair 7-162. Fixed and frozen tissues were cut in 6- to 12- microns sections, hybridized with probes at 55 degrees C for 16 h, and exposed for 5-9 days. In renal cortex, CHIP28k mRNA was detected intensely on proximal tubule epithelial cells but not in glomeruli or collecting duct. Hybridization to proximal tubule was strongest in deep renal cortex. In no study was there significant hybridization of sense cRNA probe. In renal papilla, CHIP28k mRNA was detected in only a fraction of tubules corresponding to thin limbs of Henle. Hybridization in spleen was observed in red splenic pulp containing erythroid precursors but not in white pulp. In colon, there was selective hybridization in crypt epithelial cells but not in villus epithelial cells or nonepithelial structures. In lung, hybridization was observed in alveolar epithelial cells. In eye, there was selective hybridization in corneal endothelium and ciliary body. No hybridization was observed in any cell types in liver. Northern analysis revealed a 2.8-kilobase mRNA encoding CHIP28k in kidney cortex and papilla but not in brain, skeletal muscle, and liver. These results indicate a wide and highly selective tissue distribution of mRNA encoding the CHIP28k water channel.(ABSTRACT TRUNCATED AT 250 WORDS)
Insights
This study maps the tissue distribution of rat kidney water channel CHIP28k mRNA. CHIP28k mRNA is highly and selectively expressed in specific kidney tubule cells, spleen, colon, lung, and eye tissues.
Area of Science:
- Molecular Biology
- Physiology
- Renal Physiology
Background:
- The aquaporin family of water channels facilitates water transport across cell membranes.
- CHIP28k is a water channel protein identified in rat kidney and homologous to human erythrocyte CHIP28.
- Understanding the tissue-specific expression of water channels is crucial for comprehending physiological functions.
Purpose of the Study:
- To determine the tissue distribution of messenger RNA (mRNA) encoding the rat kidney water channel CHIP28k.
- To investigate the cellular localization of CHIP28k mRNA in various rat tissues using in situ hybridization.
Main Methods:
- Isolation of complementary DNA (cDNA) encoding rat kidney CHIP28k.
- Construction of 35S-labeled cRNA sense and antisense probes.
- In situ hybridization on fixed and frozen tissue sections.
- Northern blot analysis to confirm mRNA size and distribution.
Main Results:
- CHIP28k mRNA was intensely detected in proximal tubule epithelial cells of the renal cortex and in tubules of the renal papilla (thin limbs of Henle).
- Selective hybridization was observed in spleen (red pulp), colon (crypt epithelium), lung (alveolar epithelium), and eye (corneal endothelium, ciliary body).
- No significant CHIP28k mRNA hybridization was detected in glomeruli, collecting ducts, liver, brain, or skeletal muscle.
Conclusions:
- The mRNA encoding the CHIP28k water channel exhibits a wide and highly selective tissue distribution.
- These findings suggest specific roles for CHIP28k in water transport in various organs, particularly in the kidney's proximal tubules.