Phosphate position is key in mediating transmembrane ion channel TMEM16A-phosphatidylinositol 4,5-bisphosphate interaction

TransMEMbrane 16A (TMEM16A) is a Ca2+-activated Cl- channel that plays critical roles in regulating diverse physiologic processes, including vascular tone, sensory signal transduction, and mucosal secretion. In addition to Ca2+, TMEM16A activation requires the membrane lipid phosphatidylinositol 4,5-bisphosphate (PI(4,5)P-2). However, the structural determinants mediating this interaction are not clear. Here, we interrogated the parts of the PI(4,5)P-2 head group that mediate its interaction with TMEM16A by using patch- and two-electrode voltage-clamp recordings on oocytes from the African clawed frog Xenopus laevis, which endogenously express TMEM16A channels. During continuous application of Ca2+ to excised inside-out patches, we found that TMEM16A-conducted currents decayed shortly after patch excision. Following this rundown, we show that the application of a synthetic PI(4,5)P-2 analog produced current recovery. Furthermore, inducible dephosphorylation of PI(4,5)P-2 reduces TMEM16A-conducted currents. Application of PIP2 analogs with different phosphate orientations yielded distinct amounts of current recovery, and only lipids that include a phosphate at the 40 position effectively recovered TMEM16A currents. Taken together, these findings improve our understanding of how PI(4,5)P-2 binds to and potentiates TMEM16A channels.

Automated summary

This study investigates the interaction between TMEM16A channel and the membrane lipid phosphatidylinositol 4,5-bisphosphate (PI(4,5)P-2). The findings show that only lipids containing a phosphate at the 40 position effectively recover TMEM16A currents. These findings improve our understanding of how PI(4,5)P-2 binds to and potentiates TMEM16A channels.