Role of Ca(2+) in the Stability and Function of TMEM16F and 16K

Area of Science:

• Molecular biology
• Cell biology
• Biochemistry

Background:

• The TMEM16 family comprises 10 proteins in humans and mice.
• TMEM16F functions as a Ca(2+)-dependent phospholipid scramblase at the plasma membrane.
• The precise mechanism of Ca(2+) activation of TMEM16F remains unclear.

Purpose of the Study:

• To elucidate the mechanism by which Ca(2+) activates TMEM16F's phospholipid scramblase activity.
• To investigate the role of TMEM16K in TMEM16F function.

Main Methods:

• Blue-native polyacrylamide gel electrophoresis (BN-PAGE) to detect protein complexes.
• Comprehensive mutagenesis of acidic residues in TMEM16K.
• Point mutations in TMEM16F to assess functional impact.

Main Results:

• Ca(2+) binding to TMEM16K forms a stable complex with TMEM16F.
• Ca(2+) stabilizes the structure of TMEM16K and TMEM16F, preventing aggregation.
• Five conserved acidic residues in TMEM16K are critical for Ca(2+) binding.
• Mutations in homologous residues in TMEM16F impair Ca(2+)-dependent phospholipid scrambling.

Conclusions:

• Ca(2+) directly binds to TMEM16F, inducing conformational changes.
• These Ca(2+)-induced changes enhance TMEM16F stability and phospholipid scramblase function.
• TMEM16K may play a role in mediating Ca(2+) binding to TMEM16F.