FKBP12 binds to the cardiac ryanodine receptor with negative cooperativity: implications for heart muscle physiology in health and disease

Abstract

Cardiac ryanodine receptors (RyR2) release the Ca 2+ from intracellular stores that is essential for cardiac myocyte contraction. The ion channel opening is tightly regulated by intracellular factors, including the FK506 binding proteins, FKBP12 and FKBP12.6. The impact of these proteins on RyR2 activity and cardiac contraction is debated, with often apparently contradictory experimental results, particularly for FKBP12. The isoform that regulates RyR2 has generally been considered to be FKBP12.6, despite the fact that FKBP12 is the major isoform associated with RyR2 in some species and is bound in similar proportions to FKBP12.6 in others, including sheep and humans. Here, we show time- and concentration-dependent effects of adding FKBP12 to RyR2 channels that were partly depleted of FKBP12/12.6 during isolation. The added FKBP12 displaced most remaining endogenous FKBP12/12.6. The results suggest that FKBP12 activates RyR2 with high affinity and inhibits RyR2 with lower affinity, consistent with a model of negative cooperativity in FKBP12 binding to each of the four subunits in the RyR tetramer. The easy dissociation of some FKBP12/12.6 could dynamically alter RyR2 activity in response to changes in in vivo regulatory factors, indicating a significant role for FKBP12/12.6 in Ca 2+ signalling and cardiac function in healthy and diseased hearts. This article is part of the theme issue ‘The heartbeat: its molecular basis and physiological mechanisms’.

The RyR1 P3528S Substitution Alters Mouse Skeletal Muscle Contractile Properties and RyR1 Ion Channel Gating

“…With this wide range of individual values, the average values reflect the results from the most active channels. Normalizing the data for each channel to its internal control value yields individual values and average data that are independent of intrinsic channel activity [ 18 , 19 , 20 , 21 ]. The results for parameter values expressed relative to values with 300 nM cytoplasmic Ca 2+ are shown in Figure 6 C–E and Figures S2, S4 and S6 and summarized Table 5 .…”

“…Importantly, the Central Domain is the only cytoplasmic domain that is continuous with the transmembrane domain and ion channel gating residues that control channel activity. The greatest mobility in RyRs is in the clamp domain which undergoes a downward rotation upon channel opening and contains binding sites for ligands including the FK506-binding proteins calmodulin and CLIC2 [ 21 , 32 , 33 ]. As a result of the functionally important mobility of this region, its high resolution structure is not well defined, with parts of the backbone structure remaining undefined [ 32 ].…”

“…Open probability and gating parameters are presented as the measured parameter values and are also expressed relative to internal control values with 300 nM cis Ca 2+ calculated for each individual channel. The relative changes in activity gave equal weighting to changes in activity of all RyR channels where control P o values varied from 0.0002 to 0.2 [17,18,21] Specific muscle groups were dissected for histology immediately after CO 2 euthanasia and immersed in a solution containing 10% formalin phosphate-buffered saline (pH = 6.8) and then embedded (using standard Histology procedures in the JCSMR Flow Cytometry Facility). The muscle was sectioned transversely and stained with haematoxylin and eosin (HE).…”

Ion channel trafficking implications in heart failure

“…When one of these elements is perturbed, the RYR2 undergoes a pathological remodeling, which can initiate or amplify the cellular mechanisms contributing to HF. PKA hyperphosphorylation of RYR2 in HF results in the depletion of its stabilizing FK506 binding protein, FKBP12.6 ( 96 ). RYR2 are hyperphosphorylated by PKA and/or CAMKII, causing aberrant Ca 2+ leaks from the SR ( 97 ).…”

Cardiomyocyte electrophysiology and its modulation: current views and future prospects

“…Richardson et al . [ 85 ] report time- and concentration-dependent effects of FKBP12 on previously FKBP12/12.6-depleted RyR2 channels, suggesting negative co-operativity in their FKBP12 binding, potentially significant in regulating RyR-mediated Ca 2+ signalling. Genetic gain of RyR2 or loss of calsequestrin function is associated with the pro-arrhythmic condition catecholaminergic polymorphic ventricular tachycardia (CPVT) experimentally recapitulated in murine hearts carrying genetically altered RyR2 or calsequestrin-2 [ 86 , 87 ].…”