Inhibition of cell differentiation by G alpha q in the renal epithelial cell line LLC-PK1.

Abstract

LLC-PK1, an epithelial cell line derived from the kidney proximal tubule, was used to study the ability of the G protein alpha-subunit, G alpha q, to regulate cell differentiation. A constitutively active mutant protein, alpha qQ209L, was expressed using the LacSwitch-inducible mammalian expression system. Induction of alpha qQ209L expression with isopropyl-beta-D-thiogalactopyranoside (IPTG) enhanced phospholipase C activity maximally by 6- to 7.5-fold. Increasing concentrations of IPTG progressively inhibited the activity of two differentiation markers, Na(+)-dependent hexose transport and alkaline phosphatase activity. Induction of alpha qQ209L expression also caused a change from an epithelial to a spindle-shaped morphology. The effects of alpha qQ209L expression on cell differentiation were similar to those observed with 12-O-tetradecanoylphorbol 13-acetate (TPA) treatment. However, protein kinase C (PKC) levels were downregulated in TPA-treated cells but not in alpha qQ209L-expressing cells, suggesting that the regulation of PKC by G alpha q may be different from regulation by TPA. Interestingly, the PKC inhibitor GF-109203X did not inhibit the effect of IPTG on the development of Na(+)-dependent hexose transport in alpha qQ209L-expressing cells. These data implicate PKC delta and PKC epsilon in the pathway used by G alpha q to block the development of Na(+)-dependent hexose transport in IPTG-treated cells.