Notch enhances Ca entry by activating calcium-sensing receptors and inhibiting voltage-gated K channels.

The increase in cytosolic Ca concentration ([Ca]) and upregulation of calcium-sensing receptor (CaSR) and stromal interaction molecule 2 (STIM2) along with inhibition of voltage-gated K (K) channels in pulmonary arterial smooth muscle cells (PASMC) have been implicated in the development of pulmonary arterial hypertension; however, the precise upstream mechanisms remain elusive. Activation of CaSR, a G protein-coupled receptor (GPCR), results in Ca release from the endoplasmic/sarcoplasmic reticulum (ER/SR) and Ca influx through receptor-operated and store-operated Ca channels (SOC). Upon Ca depletion from the SR, STIM forms clusters to mediate store-operated Ca entry. Activity of K channels, like KCNA5/K1.5 and KCNA2/K1.2, contributes to regulating membrane potential, and inhibition of K channels results in membrane depolarization that increases [Ca] by opening voltage-dependent Ca channels. In this study, we show that activation of Notch by its ligand Jag-1 promotes the clustering of STIM2, and clustered STIM2 subsequently enhances the CaSR-induced Ca influx through SOC channels. Extracellular Ca-mediated activation of CaSR increases [Ca] in -transfected HEK293 cells. Treatment of -transfected cells with Jag-1 further enhances CaSR-mediated increase in [Ca]. Moreover, CaSR-mediated increase in [Ca] was significantly augmented in cells co-transfected with and . CaSR activation results in STIM2 clustering in -cotransfected cells. Notch activation also induces significant clustering of STIM2. Furthermore, activation of Notch attenuates whole cell K currents in - and -transfected cells. Together, these results suggest that Notch activation enhances CaSR-mediated increases in [Ca] by enhancing store-operated Ca entry and inhibits KCNA5/K1.5 and KCNA2/K1.2, ultimately leading to voltage-activated Ca entry.