Laboratory Investigation

Kidney International (1994) 46, 627–638; doi:10.1038/ki.1994.315

Segmental localization of mRNAs encoding Na+-K+-ATPase alpha- and bold beta-subunit isoforms in rat kidney using RT-PCR

William L Clapp1, Paula Bowman1, Geraldine S Shaw1, Pinkal Patel1 and Bruce C Kone1

1DCI Laboratory of Molecular Biology in Nephrology, Division of Nephrology, Hypertension and Transplantation, and The Hypertension Center, University of Florida College of Medicine, Gainesville, Florida, USA

Correspondence: Bruce C Kone MD, Division of Nephrology, Hypertension and Transplantation, University of Florida College of Medicine, Box 100224, JHMHC, Gainesville, Florida 32610-0224, USA.

Received 9 July 1993; Revised 22 March 1994; Accepted 24 March 1994.

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Abstract

Segmental localization of mRNAs encoding Na+-K+-ATPase alpha- and beta-subunit isoforms in rat kidney using RT-PCR. To characterize the expression of genes encoding the alpha- and beta-subunit isoforms of the Na+-K+-ATPase in rat kidney, we used reverse transcription (RT)-PCR of microdissected renal structures combined with quantitation of subunit isoform mRNAs in the major renal parenchymal zones. Transcripts for alpha1, alpha2, alpha3, beta1, and beta2 subunit isoforms were detected by RT-PCR in microdissected glomeruli, proximal convoluted tubules, medullary thick ascending limbs of Henle, cortical and inner medullary collecting ducts. The truncated alpha1 (alpha1-T) isoform was also amplified from cortex, outer and inner medulla and isolated glomeruli, but it was not detected in these nephron segments. The DNA sequence of the renal alpha1-T PCR product was identical to that of the cDNA previously cloned from aortic smooth muscle cells. RNA dot-blot analysis indicated that the alpha1, alpha2, and alpha3 isoforms contributed approx70%, approx20%, and approx10%, respectively, of the total alpha isoform mRNA in each parenchymal zone. RNase protection assays determined that the beta1 and beta2 isoforms accounted for approx95% and approx5%, respectively, of the beta isoform mRNA in each zone. These data provide definitive evidence for the differential expression of mRNAs encoding all the alpha and beta isoforms in the renal parenchyma, and for the coexpression of these isoforms in the nephron segments examined. The results suggest the potential expression of up to eight different Na+-K+-ATPase isoenzymes in the kidney, and for multiple molecular levels of regulation of renal Na+-K+-ATPase expression.

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