PIP₂ and septin control STIM1/Orai1 assembly by regulating cytoskeletal remodeling via a CDC42-WASP/WAVE-ARP2/3...

Store-operated calcium entry (SOCE) is triggered by assembly of Orai1 with STIM proteins in ER-PM junctions. Plasma membrane PIP₂ as well as PIP₂-binding protein, SEPT4, significantly impact Orai1-STIM1 interaction. While septins and PIP₂ can organize the actin cytoskeleton, it is unclear whether the status of actin within the junctions contributes to SOCE. We report herein that actin remodeling modulates STIM1 clustering. Our findings show that a PIP₂- and SEPT4-dependent mechanism involving CDC42, WASP/WAVE, and ARP2 regulates actin remodeling into a ring-like structure around STIM1 puncta. CDC42 localization in the ER-plasma membrane region is enhanced following ER-Ca²⁺ store depletion. PIP₂ depletion or knockdown of SEPT4 attenuate the recruitment of CDC42 to the ER-PM region. Importantly, knockdown of SEPT4, or CDC42+ARP2, disrupts the organization of actin as well as STIM1 clustering. Consequently, Orai1 recruitment to STIM1 puncta, SOCE, and NFAT translocation to the nucleus are all attenuated. Ca²⁺ influx induced by STIM1-C terminus is not affected by CDC42 knockdown. In aggregate, our findings reveal that PIP₂ and SEPT4 affect Orai1/STIM1 clustering by coordinating actin remodeling within ER-PM junctions. This dynamic reorganization of actin has an important role in regulation of SOCE and downstream Ca²⁺-dependent effector functions.