Expression, functional characterization and tissue distribution of a Na+/H+ exchanger cloned from Xenopus laevis
S Busch1, D Rosskopf, H J Lang
1Institut für Pharmakologie, Universitätsklinikum Essen, Hufelandstr. 55, D-45122 Essen, Germany. stefan.busch@uni-essen.de
Abstract:
We examined the functional properties of a Na+/H+ exchanger cloned from Xenopus laevis oocytes (XL-NHE) upon stable transfection into PS120 fibroblasts which lack endogenous Na+/H+ exchange. In contrast to untransfected cells, XL-NHE-transfected cells displayed Na+-dependent alkalinization upon acidification with nigericin. XL-NHE activity was inhibited by amiloride, ethylisopropylamiloride, HOE694 [(3-methylsulphonyl-4-piperidinobenzoyl)-guanidine methanesulphonate] and HOE642 [4-isopropyl-3-methylsulphonylbenzoyl)-guanidine methanesulphonate], Ki values being calculated at 5 micromol/l, 25 nmol/l, 300 nmol/l and 180 nmol/l, respectively. The Na+ dependence of pHi recovery was compatible with simple Michaelis-Menten kinetics, the Km for Na+ being 22.0+/-3.2 mmol/l and the Hill coefficient for Na+ being approximately 1. XL-NHE was activated by phorbol ester, whereas forskolin exerted no effect, suggesting the involvement of phospholipase C/protein kinase C signalling pathways rather than protein kinase A signalling pathways in XL-NHE stimulation. Using reverse transcription polymerase chain reaction, XL-NHE message could be detected in various Xenopus tissues including heart, brain, skeletal muscle, reticulocytes, A6-kidney cells and oocytes.
