ACID INCUBATION INCREASES NHE-3 MESSENGER-RNA ABUNDANCE IN OKP CELLS

With the use of degenerate primers based on conserved amino acid seque nces in human, rat, and rabbit Na/H exchanger-3 (NHE-3), a polymerase chain reaction product was obtained from reverse-transcribed OKP (a cl onal opossum kidney cell line) mRNA and used to screen an OKP cDNA lib rary. The clone obtained predicted an amino acid sequence that was 86% identical to rat NHE-3, 33% to NHE-1, 35% to NHE-2, and 30% to NHE-4. Expression of the corresponding cRNA in Xenopus oocytes induced Na-22 uptake with ethylisopropylamiloride (EIPA) resistance similar to that of the OKP Na/H antiporter. On RNA blot, the cDNA labeled a 9.5-kb tr anscript whose abundance was increased 2.2-fold by 24-h incubation of OKP cells at pH 7.0 and 2.5-fold by 24-h incubation at pH 6.8. The aci d-induced increase in NHE-3 mRNA was detectable at 12 h and increased further at 24 h. Incubation in acid media caused an increase in EIPA-r esistant Na/H antiporter activity that preceded the increase in NHE-3 mRNA. In summary, OKP cells express an NHE-3 transcript that encodes a n EIPA-resistant Na/H antiporter and is chronically regulated by acid.