Recent advances in understanding the ryanodine receptor calcium release channels and their role in calcium signalling

Abstract

The ryanodine receptor calcium release channel is central to cytoplasmic Ca ²⁺ signalling in skeletal muscle, the heart, and many other tissues, including the central nervous system, lymphocytes, stomach, kidney, adrenal glands, ovaries, testes, thymus, and lungs. The ion channel protein is massive (more than 2.2 MDa) and has a structure that has defied detailed determination until recent developments in cryo-electron microscopy revealed much of its structure at near-atomic resolution. The availability of this high-resolution structure has provided the most significant advances in understanding the function of the ion channel in the past 30 years. We can now visualise the molecular environment of individual amino acid residues that form binding sites for essential modulators of ion channel function and determine its role in Ca ²⁺ signalling. Importantly, the structure has revealed the structural environment of the many deletions and point mutations that disrupt Ca ²⁺ signalling in skeletal and cardiac myopathies and neuropathies. The implications are of vital importance to our understanding of the molecular basis of the ion channel's function and for the design of therapies to counteract the effects of ryanodine receptor-associated disorders.

Keywords: High Resolution Cryo-EM; Ryanodine Receptor; Skeletal and Cardiac Muscle.

Figure 1.. The near-atomic resolution structure of RyR1 .

( A) The linear sequence of RyR1 domains, starting at the N-terminal domain (NTD). ( B) “Side” view of RyR1 highlighting two of four protomers. Domains in ( B) are colour-coded in the same way as those in ( A). Modified from Figure 1 of Yan et al. . The 10 major domains in each subunit include the N-terminal domain, which harbours many RyR1 and RyR2 disease-causing mutations. They also include the SPRY1 domain that forms part of the binding site for FKPB12, the helical and central domains, which contain protein kinase A and Ca ²⁺/calmodulin protein kinase II (CaMKII) phosphorylation sites, the transmembrane domain containing the ion pore, and the C-terminal domain that forms part of the Ca ²⁺, ATP, and caffeine activation sites ( Figure 2). FKBP12 binds in a cleft formed by the Handle, NTD, and SPRY1/3 domains. Suggested FKBP12 binding residues are located in the Handle domain (P1780, C1781, and S1687) and in a hydrophobic cluster around D720 in the SPRY1 domain . The cytoplasmic surface of the transverse tubule (T-tubule) membrane is shown overlying the RyR to illustrate the way that the bulk of the RyR is sandwiched between the membranes on either side of the T-tubule/sarcoplasmic reticulum (SR) junction. RyR, ryanodine receptor.

Figure 2.. “Side view” of the open RyR1 channel structure .

The structure (PDB code 5TAL) reveals the major domains of the protein and the location of Ca ²⁺, ATP, and caffeine binding sites identified by . Adapted from ; the structural domain nomenclature is as given in . The transmembrane domain (TMD) containing the permeation pathway is shown embedded in the sarcoplasmic reticulum (SR) membrane, which is depicted as a solid pale blue rectangle. NTD, N-terminal domain; RyR, ryanodine receptor.

Figure 3.. Proposed region for ionic interactions between skeletal triadin (Trisk 95) and RyR1.

The near-atomic resolution structure of the pore-forming elements of RyR1 showing two diagonal protomers (Extended Data Figure 8 in ). The grey transmembrane S5 and S6 helices and red pore helix, S5–pore helix linker, and SF linker between the pore helix and S6 helix are shown. Mutagenesis studies suggest that residues D4878, E4907, and E4908 in the outer regions of the pore helix are associated with K218, K220, and K224 in Trisk 95 ^, . The approximate positions of E4907 and E4908 are indicated by the black arrowhead, and the arrow indicates the predicted binding site for Trisk 95. RyR, ryanodine receptor; SR, sarcoplasmic reticulum. Reprinted by permission from Springer Nature: Pflügers Archiv European Journal of Physiology, Three residues in the luminal domain of triadin impact on Trisk 95 activation of skeletal muscle ryanodine receptors, E. Wium, A. F. Dulhunty, N. A. Beard, © 2016.