Cookies on this website

We use cookies to ensure that we give you the best experience on our website. If you click 'Accept all cookies' we'll assume that you are happy to receive all cookies and you won't see this message again. If you click 'Reject all non-essential cookies' only necessary cookies providing core functionality such as security, network management, and accessibility will be enabled. Click 'Find out more' for information on how to change your cookie settings.

Phosphorylation of the cardiac ryanodine receptor (RyR2) is thought to be important not only for normal cardiac excitation-contraction coupling but also in exacerbating abnormalities in Ca²+ homeostasis in heart failure. Linking phosphorylation to specific changes in the single-channel function of RyR2 has proved very difficult, yielding much controversy within the field. We therefore investigated the mechanistic changes that take place at the single-channel level after phosphorylating RyR2 and, in particular, the idea that PKA-dependent phosphorylation increases RyR2 sensitivity to cytosolic Ca²+. We show that hyperphosphorylation by exogenous PKA increases open probability (P(o)) but, crucially, RyR2 becomes uncoupled from the influence of cytosolic Ca²+; lowering [Ca²+] to subactivating levels no longer closes the channels. Phosphatase (PP1) treatment reverses these gating changes, returning the channels to a Ca²+-sensitive mode of gating. We additionally found that cytosolic incubation with Mg²+/ATP in the absence of exogenously added kinase could phosphorylate RyR2 in approximately 50% of channels, thereby indicating that an endogenous kinase incorporates into the bilayer together with RyR2. Channels activated by the endogenous kinase exhibited identical changes in gating behavior to those activated by exogenous PKA, including uncoupling from the influence of cytosolic Ca²+. We show that the endogenous kinase is both Ca²+-dependent and sensitive to inhibitors of PKC. Moreover, the Ca²+-dependent, endogenous kinase-induced changes in RyR2 gating do not appear to be related to phosphorylation of serine-2809. Further work is required to investigate the identity and physiological role of this Ca²+-dependent endogenous kinase that can uncouple RyR2 gating from direct cytosolic Ca²+ regulation.

Original publication




Journal article


J Membr Biol

Publication Date





21 - 33


Animals, Calcium, Calcium Channels, Cyclic AMP-Dependent Protein Kinases, Cytosol, Heart Failure, Homeostasis, Humans, Ion Channel Gating, Lipid Bilayers, Myocardium, Myocytes, Cardiac, Phosphoprotein Phosphatases, Phosphorylation, Ryanodine, Ryanodine Receptor Calcium Release Channel, Sarcoplasmic Reticulum, Serine, Sheep