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Single-Cell Calcium Imaging for Studying the Activation of Calcium Ion Channels
Published on: December 13, 2024
Orai1 and STIM reconstitute store-operated calcium channel function
Jonathan Soboloff1, Maria A Spassova1, Xiang D Tang1
1Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine, Baltimore, Maryland 21201.
The Journal of Biological Chemistry
|June 13, 2006
Summary
The interaction between STIM1 and Orai1 proteins is crucial for store-operated calcium entry (SOCE). Coexpression reconstitutes functional SOCs, with Orai1 forming the plasma membrane channel component.
Area of Science:
- Cell Biology
- Molecular Biology
- Ion Channel Physiology
Background:
- Store-operated channels (SOCs) regulate calcium influx into cells.
- STIM1 and Orai1 are known essential proteins for SOC activation.
- The precise functional interaction between STIM1 and Orai1 remains unclear.
Purpose of the Study:
- To elucidate the functional interaction between STIM1 and Orai1.
- To determine the roles of STIM1 and Orai1 in store-operated calcium entry (SOCE).
- To investigate the channel properties of Orai1 in the context of STIM1 interaction.
Main Methods:
- Coexpression of STIM1 and Orai1 in human embryonic kidney 293 cells and rat basophilic leukemia cells.
- Measurement of store-operated calcium entry (SOCE) and calcium release-activated calcium current (I(CRAC)).
- Assessment of store-dependent and store-independent calcium entry.
- Pharmacological inhibition using 2-aminoethoxydiphenylborate.
Main Results:
- Coexpression of STIM1 and Orai1 reconstituted functional SOCs, significantly increasing SOCE (up to 103-fold).
- Orai1 alone suppressed SOCE and I(CRAC), but coexpression with STIM1 dramatically enhanced them.
- The enhanced calcium entry was strictly store-dependent and blocked by 2-aminoethoxydiphenylborate.
- STIM2 coexpressed with Orai1 induced constitutive calcium entry, unlike STIM1.
Conclusions:
- Orai1 likely forms the plasma membrane channel pore for calcium entry in SOCs.
- STIM1 acts as the calcium store sensor, mediating the activation of Orai1.
- The findings suggest a critical stoichiometry between STIM1 and Orai1 for proper SOC function.
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