Essential role of N-terminal SAM regions in STIM1 multimerization and function

Area of Science:

• Cellular Biology
• Molecular Biology
• Biophysics

Background:

• Stromal Interaction Molecule 1 (STIM1) regulates calcium (Ca2+) homeostasis by sensing ER Ca2+ levels.
• STIM1 activates Orai1 calcium channels upon Ca2+ store depletion.
• The structural basis for STIM1 luminal multimerization remained unclear.

Purpose of the Study:

• To elucidate the structural mechanisms underlying STIM1 luminal di/multimerization.
• To identify specific regions within the STIM1 sterile alpha motif (SAM) domain involved in multimerization.
• To investigate the functional consequences of impaired STIM1 multimerization on calcium signaling.

Main Methods:

• Molecular dynamics (MD) simulations to predict multimerization sites.
• Site-directed mutagenesis of identified STIM1 SAM domain regions.
• Biochemical assays and live-cell imaging to assess STIM1 multimerization and Orai1 channel activity.
• Analysis of calcium influx and oscillation frequency.

Main Results:

• MD simulations identified two key multimerization segments (α7 and near α9) in the STIM1 SAM domain.
• Point mutations in these segments disrupted STIM1 higher-order multimerization and puncta formation.
• Impaired STIM1 multimerization significantly reduced store-operated calcium entry and oscillation frequency.
• Functional defects were partially rescued by combining mutations with gain-of-function mutations or Orai1 coexpression.

Conclusions:

• Hydrophobicity-driven interactions within the STIM1 SAM domain are critical for its switch between monomeric and multimeric states.
• STIM1 SAM domain interactions are essential for STIM1 multimerization and subsequent Orai1 channel activation.
• This study reveals the structural basis for STIM1-mediated calcium signaling regulation.