Junctin and triadin each activate skeletal ryanodine receptors but junctin alone mediates functional interactions with calsequestrin

Abstract

Summary Normal Ca2+ signalling in skeletal muscle depends on the membrane associated proteins triadin and junctin and their ability to mediate functional interactions between the Ca2+ binding protein calsequestrin and the type 1 ryanodine receptor in the lumen of the sarcoplasmic reticulum. This important mechanism conserves intracellular Ca2+ stores, but is poorly understood. Triadin and junctin share similar structures and are lumped together in models of interactions between skeletal muscle calsequestrin and ryanodine receptors, however their individual roles have not been examined at a molecular level. We show here that purified skeletal ryanodine receptors are similarly activated by purified triadin or purified junctin added to their luminal side, although a lack of competition indicated that the proteins act at independent sites. Surprisingly, triadin and junctin differed markedly in their ability to transmit information between skeletal calsequestrin and ryanodine receptors. Purified calsequestrin inhibited junctin/triadin-associated, or junctin-associated, ryanodine receptors and the calsequestrin re-associated channel complexes were further inhibited when luminal Ca2+ fell from 1mM to ≤100μM, as seen with native channels (containing endogenous calsequestrin/triadin/junctin). In contrast, skeletal calsequestrin had no effect on the triadin/ryanodine receptor complex and the channel activity of this complex increased when luminal Ca2+ fell, as seen with purified channels prior to triadin/calsequestrin re-association. Therefore in this cell free system, junctin alone mediates signals between luminal Ca2+, skeletal calsequestrin and skeletal ryanodine receptors and may curtail resting Ca2+ leak from the sarcoplasmic reticulum. We suggest that triadin serves a different function which may dominate during excitation- contraction coupling.

Organellar Calcium Buffers

“…Phosphorylated calsequestrin strongly interacts with junctin and hence inhibits the RyR1 channel (Beard et al 2008). Junctin, but not triadin, is required for skeletal calsequestrin to exert regulatory control over RyR1 channels (Wei et al 2009a). At resting SR Ca 2þ levels, skeletal calsequestrin inhibits RyR1; by contrast, cardiac calsequestrin activates both RyR1 and RyR2 (Wei et al 2009b).…”

“…These interactions are thought to be mediated by the transmembrane proteins junctin and triadin, though recent results suggest that only junctin may be crucial for the regulation of RyR proteins by calsequestrin in skeletal muscle (Wei et al 2009a). Interactions between calsequestrin and the RyR depend on the luminal Ca 2þ concentration ).…”

A novel cytoplasmic interaction between junctin and ryanodine receptor calcium release channels

“…4 below and Wei et al, 2009a). The rationale was that if endogenous junctin was bound to native RyRs, then exogenous FLjun added to the luminal solution could not bind to or activate the channels.…”

Mapping domains and mutations on the skeletal muscle ryanodine receptor channel

“…It has been suggested that junctin may be the main protein mediator between CSQ and RyR [64]. Knockdown of junctin in myotubes reduces both depolarization-induced SR Ca 2+ release and the content of Ca 2+ in the SR [61], possibly by disrupting the signaling complex between CSQ and RyR.…”