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TRPP2 and STIM1 form a microdomain to regulate store-operated Ca2+ entry and blood vessel tone.

Cell communication and signaling : CCS (2020-09-02)
Jizheng Guo, Ren Zhao, Muyao Zhou, Jie Li, Xiaoqiang Yao, Juan Du, Jiexia Chen, Bing Shen
ABSTRACT

Polycystin-2 (TRPP2) is a Ca2+ permeable nonselective cationic channel essential for maintaining physiological function in live cells. Stromal interaction molecule 1 (STIM1) is an important Ca2+ sensor in store-operated Ca2+ entry (SOCE). Both TRPP2 and STIM1 are expressed in endoplasmic reticular membrane and participate in Ca2+ signaling, suggesting a physical interaction and functional synergism. We performed co-localization, co-immunoprecipitation, and fluorescence resonance energy transfer assay to identify the interactions of TRPP2 and STIM1 in transfected HEK293 cells and native vascular smooth muscle cells (VSMCs). The function of the TRPP2-STIM1 complex in thapsigargin (TG) or adenosine triphosphate (ATP)-induced SOCE was explored using specific small interfering RNA (siRNA). Further, we created TRPP2 conditional knockout (CKO) mouse to investigate the functional role of TRPP2 in agonist-induced vessel contraction. TRPP2 and STIM1 form a complex in transfected HEK293 cells and native VSMCs. Genetic manipulations with TRPP2 siRNA, dominant negative TRPP2 or STIM1 siRNA significantly suppressed ATP and TG-induced intracellular Ca2+ release and SOCE in HEK293 cells. Inositol triphosphate receptor inhibitor 2-aminoethyl diphenylborinate (2APB) abolished ATP-induced Ca2+ release and SOCE in HEK293 cells. In addition, TRPP2 and STIM1 knockdown significantly inhibited ATP- and TG-induced STIM1 puncta formation and SOCE in VSMCs. Importantly, knockdown of TRPP2 and STIM1 or conditional knockout TRPP2 markedly suppressed agonist-induced mouse aorta contraction. Our data indicate that TRPP2 and STIM1 are physically associated and form a functional complex to regulate agonist-induced intracellular Ca2+ mobilization, SOCE and blood vessel tone. Video abstract.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Duolink® In Situ Detection Reagents Red
Sigma-Aldrich
Thapsigargin, ≥98% (HPLC), solid film
Sigma-Aldrich
Duolink® In Situ PLA® Probe Anti-Rabbit MINUS
Sigma-Aldrich
Duolink® In Situ PLA® Probe Anti-Goat PLUS
Sigma-Aldrich
MISSION® esiRNA, targeting human STIM1