Protein Fragmentation As a Regulatory Mechanism: Insights from Two Different Ca2+ Channels, RyR1 and IP3R

The ryanodine receptor 1 (RyR1) is the major skeletal muscle Ca 2+ release channel and as such a key player in excitation-contraction coupling. Intriguingly, a recent paper reported RyR1 fragmentation 24 h after high-intensity interval exercise (six 30-s all-out cycling bouts with 4 min recoveries in between) in vastus lateralis muscle biopsies taken from recreationally active men (Place et al., 2015). In contrast to what could have been hypothesized based on the role of RyR1 in excitation-contraction coupling, this RyR1 fragmentation did not appear to result in excitation-contraction coupling failure, as both maximal voluntary contraction force and forces evoked by supramaximal electrical stimulations had fully recovered 24 h after exercise. Overall, these results suggest that fragmented RyR1 retain their capacity to release Ca 2+ in response to an action potential. If at first this result might appear surprising, as fragmentation is generally thought to result in dysfunctional channels, a recent review published by Yule and colleagues highlighted that fragmentation might actually serve as a regulating mechanism, at least for the inositol 1,4,5-triphosphate receptor (IP 3 R), the major Ca 2+ release channel in non-excitable cells (Wang et al., 2016). Briefly, in addition of being physiologically activated by inositol 1,4,5-triphosphate (IP 3), IP 3 R can be modulated by intracellular Ca 2+ , ATP, cAMP, as well as by post-translational changes such as phosphorylation and redox modifications, similarly to RyR1 (Lanner et al., 2010). By proteolytic cleavage, also IP 3 R may become fragmented (Hirota et al. then argued that the model used in the earlier studies presented important limitations precluding such conclusions. For example in the study of Assefa et al. (2004), a construct encoding only the IP 3 R caspase-cleaved C-terminal domain was expressed in DT40-3KO cells (chicken B-lymphocytes with all IP 3 R isoforms knocked-out) and resulted in an enhanced Ca 2+ leak. According to Wang et al. (2016) these results are not informative of the functionality of fragmented IP 3 Rs as (i) the C-terminal portion of the IP 3 R might have been overexpressed, and (ii) it was expressed in a background without the IP 3 R N-terminal cytoplasmic domain. Using DT40-3KO cells expressing IP 3 R isoform 1 (IP 3 R1), they then showed, by separation on a native non-denaturating gel, that both the N-and C-terminal fragments of the channel remained associated following IP 3 R fragmentation induced by staurosporine (Alzayady et al., 2013). To …