Ca2+-calmodulin increases RyR2 open probability yet reduces ryanoid association with RyR2.

Area of Science:

• Cardiovascular Physiology
• Molecular Cardiology
• Ion Channel Biophysics

Background:

• Calcium-calmodulin (Ca(2+)CaM) activates cardiac ryanodine receptors (RyR2) at physiological concentrations, increasing Ca(2+) sparks and waves.
• Ca(2+)CaM is known to inhibit [(3)H]ryanodine binding to cardiac sarcoplasmic reticulum, a method that doesn't reflect RyR2 open probability (Po).

Purpose of the Study:

• To investigate if Ca(2+)CaM directly influences ryanoid binding to single RyR2 channels, independent of Po.
• To elucidate the mechanism by which Ca(2+)CaM modulates RyR2 function.

Main Methods:

• Utilized the reversible ryanoid, ryanodol, to study Ca(2+)CaM's effect on ryanoid binding kinetics to single RyR2 channels.
• Analyzed voltage-dependent gating of ryanodol-bound RyR2 channels and the influence of Ca(2+)CaM on subconductance states.

Main Results:

• Ca(2+)CaM was found to reduce the association rate of ryanodol to RyR2 without altering the dissociation rate.
• Ca(2+)CaM significantly altered the equilibrium between the M(1) and M(2) subconductance states of ryanodol-bound RyR2 channels.
• These findings suggest Ca(2+)CaM binding induces conformational changes in RyR2, affecting the ryanoid binding site and gating.

Conclusions:

• Ca(2+)CaM binding to RyR2 directly modulates ryanoid binding kinetics and channel gating, explaining discrepancies with traditional binding assays.
• The study provides a molecular mechanism for Ca(2+)CaM's regulation of RyR2 function in cardiac cells.