Differential regulation of vacuolar H+‐ATPase subunits by transforming growth factor‐β1 in salivary ducts

Abstract Bicarbonate concentration in saliva is controlled by the action of acid–base transporters in salivary duct cells. We show for the first time expression of ATP6V1B1 in submandibular gland and introduce transforming growth factor‐beta (TGF‐β) as a novel regulator of V‐ATPase subunits. Using QRT‐PCR, immunoblotting, biotinylation of surface proteins, immunofluorescence, chromatin immunoprecipitation, and intracellular H( + ) recording with H( + )‐sensitive dye 2′,7′‐bis‐(carboxyethyl)‐5‐(and‐6)‐carboxyfluorescein we show that in the human submandibular gland (HSG) cell line, activation of TGF‐β signaling upregulates ATP6V1E1 and ATP6V1B2, downregulates ATP6V1B1, and has no effect on ATP6V1A. TGF‐β1 effects on ATP6V1B1 are mediated through the canonical, the soluble adenylate cyclase, and ERK signaling. A CREB binding sequence was identified in the ATP6V1B1 promoter and CREB binding decreased after TGF‐β1 treatment. Following acidosis, a bafilomycin‐sensitive and Na + ‐independent cell pH recovery was observed in HSG cells, an effect that was not influenced after disruption of acidic lysosomes. Moreover, neutralization of TGF‐βs, inhibition of TGF‐β receptor, or inhibition of the canonical pathway decreased membrane expression of ATP6V1A and prevented the acidosis‐induced increased V‐ATPase activity. The results suggest multiple modes of action of TGF‐β1 on V‐ATPase subunits in HSG cells: TGF‐β1 may regulate transcription or protein synthesis of certain subunits and trafficking of other subunits in a context‐dependent manner. Moreover, surface V‐ATPase is active in salivary duct cells and involved in intracellular pH regulation following acidosis.