Contrasting effects of phosphatidylinositol 4,5-bisphosphate on cloned TMEM16A and TMEM16B channels.

BACKGROUND AND PURPOSE: Ca2+ -activated Cl- channels (CaCCs) are gated open by a rise in intracellular Ca2+ concentration ([Ca2+ ]i ), typically provoked by activation of Gq -protein coupled receptors (Gq PCR). Gq PCR activation initiates depletion of plasmalemmal phosphatidylinositol 4,5-bisphosphate (PIP2 ). Here, we determined whether PIP2 acts as a signalling lipid for CaCCs coded by the TMEM16A and TMEM16B genes. EXPERIMENTAL APPROACH: Patch-clamp electrophysiology, in conjunction with genetically encoded systems to control cellular PIP2 content, was used to define the mechanism of action of PIP2 on TMEM16A and TMEM16B channels. KEY RESULTS: A water-soluble PIP2 analogue (diC8-PIP2 ) activated TMEM16A channels by up to fivefold and inhibited TMEM16B by ~0.2-fold. The effects of diC8-PIP2 on TMEM16A currents were especially pronounced at low [Ca2+ ]i . In contrast, diC8-PIP2 modulation of TMEM16B channels did not vary over a broad [Ca2+ ]i range but was only detectable at highly depolarized membrane potentials. Modulation of TMEM16A and TMEM16B currents was due to changes in channel gating, while single channel conductance was unaltered. Co-expression of TMEM16A or TMEM16B with a Danio rerio voltage-sensitive phosphatase (DrVSP), which degrades PIP2 , led to reduction and enhancement of TMEM16A and TMEM16B currents respectively. These effects were abolished by an inactivating mutation in DrVSP and antagonized by simultaneous co-expression of a phosphatidylinositol-4-phosphate 5-kinase that catalyses PIP2 formation. CONCLUSIONS AND IMPLICATIONS: PIP2 acts as a modifier of TMEM16A and TMEM16B channel gating. Drugs interacting with PIP2 signalling may affect TMEM16A and TMEM16B channel gating and have potential uses in basic science and implications for therapy.