Isolation and characterization of a Na-H antiporter-deficient mutant of LLC-PK1 cells

Abstract

LLC-PK1 is an established cell line derived from pig kidney epithelium; it differentiates in vitro to form a polarized epithelial sheet, capable of the vectorial transport of solutes and water. We have used a modification of the "proton-suicide" method of Pouyssegur et al. (Proc. Natl. Acad. Sci. USA. 81:4833-4837, 1984) to isolate a mutant of LLC-PK1 cells that is deficient in Na-H antiporter activity. The mutant grows normally at pH 7.0 and above in the presence and absence of bicarbonate; at pH 6.5, however, it requires bicarbonate for growth. Alkalinization of the cytoplasm by the Na-H antiporter thus appears to be essential for growth at acidic pH in the absence of bicarbonate. The antiporter is also essential for the formation of domes (a consequence of vectorial water transport) in the absence of bicarbonate.