Simone Pelizzari: Understanding the functional role of RyR1 cytosolic domains required for skeletal muscle excitation–contraction coupling

Speaker: Simone Pelizzari / Medical University of Innsbruck

Abstract: Ryanodine receptor 1 (RyR1) is a homotetrameric Ca²⁺ release channel located in the skeletal muscle sarcoplasmic reticulum (SR) membrane. Unlike cardiac RyR2, RyR1 is not activated by Ca²⁺ influx through voltage-gated L-type Ca²⁺ channels. Instead, skeletal muscle excitation–contraction coupling (ECC) relies on a direct mechanical interaction between RyR1 and the skeletal muscle L-type channel CaV1.1, which functions as the voltage sensor. Notably, this mode of activation is specific for RyR1, not conserved in RyR2 and RyR3. The RyR1 domains that mediate this communication, however, remain poorly defined. For many years, the cytosolic Divergent Region 2 (DR2) was the only RyR1 domain directly implicated in this process. However, multiple lines of evidence indicate that DR2 is neither sufficient nor isoform-specific for skeletal muscle ECC. In fact, the translocation of RyR1 DR2 into the ECC-incompetent RyR3 fails to restore depolarization-induced Ca²⁺ release, demonstrating that DR2 alone is insufficient for coupling to CaV1.1. Conversely, replacing RyR1 DR2 with the corresponding RyR2 sequence preserves ECC, indicating that DR2 is not isoform-specific for skeletal muscle ECC. Consistent with the involvement of additional RyR1 regions, recent in situ cryo-electron tomography has implicated at least one additional cytosolic RyR1 domain in ECC. Here, we investigate the functional contribution and isoform specificity of this additional RyR1 domain to skeletal muscle ECC using a series of chimeric RyR1 constructs in which the homologous domain is replaced with that from RyR2 or RyR3, combined with electrophysiological recordings and fluorescent Ca²⁺ imaging.