Functional coupling between TRPV4 channel and TMEM16F modulates human trophoblast fusion

TMEM16F, a Ca²⁺-activated phospholipid scramblase (CaPLSase), is critical for placental trophoblast syncytialization, HIV infection, and SARS-CoV2-mediated syncytialization, however, how TMEM16F is activated during cell fusion is unclear. Here, using trophoblasts as a model for cell fusion, we demonstrate that Ca²⁺ influx through the Ca²⁺ permeable transient receptor potential vanilloid channel TRPV4 is critical for TMEM16F activation and plays a role in subsequent human trophoblast fusion. GSK1016790A, a TRPV4 specific agonist, robustly activates TMEM16F in trophoblasts. We also show that TRPV4 and TMEM16F are functionally coupled within Ca²⁺ microdomains in a human trophoblast cell line using patch-clamp electrophysiology. Pharmacological inhibition or gene silencing of TRPV4 hinders TMEM16F activation and subsequent trophoblast syncytialization. Our study uncovers the functional expression of TRPV4 and one of the physiological activation mechanisms of TMEM16F in human trophoblasts, thus providing us with novel strategies to regulate CaPLSase activity as a critical checkpoint of physiologically and disease-relevant cell fusion events.