The interactive role of STAC3 in the skeletal excitation-contraction coupling machinery
Excitation-contraction (EC) coupling in skeletal muscle is an essential cellular process that requires a physical coupling between the dihydropyridine receptor (DHPR) L-type Ca2+ channel or CaV1.1 in the surface membrane, and the ryanodine receptor (RyR1) Ca2+ release channel in the sarcoplasmic reticulum (SR) membrane. However, the exact nature of the molecular interaction between the dihydropyridine receptor and the ryanodine receptor in skeletal muscle remains unclear. Recently, the adaptor...[Show more]
|dc.description.abstract||Excitation-contraction (EC) coupling in skeletal muscle is an essential cellular process that requires a physical coupling between the dihydropyridine receptor (DHPR) L-type Ca2+ channel or CaV1.1 in the surface membrane, and the ryanodine receptor (RyR1) Ca2+ release channel in the sarcoplasmic reticulum (SR) membrane. However, the exact nature of the molecular interaction between the dihydropyridine receptor and the ryanodine receptor in skeletal muscle remains unclear. Recently, the adaptor protein STAC3 has been identified as another essential component of the skeletal EC coupling machinery. STAC3 coprecipitates with both the CaV1.1 and RyR1, thereby indicating that it is a critical part of the CaV1.1/RyR1 triadic complex that is central to skeletal EC coupling. More importantly, deletion of STAC3 in mice results in complete paralysis and perinatal lethality with severe musculoskeletal defects that are characteristic of models lacking EC coupling. Additionally, in humans, a point mutation (W284S) in STAC3 is implicated in Native American Myopathy (NAM), a congenital disease characterized by muscle weakness and a predisposition to malignant hyperthermia. Recent works have focused on the functional interactions between STAC3 and CaV, with two distinct interactions having already been identified. Nonetheless, there is still little evidence of a functional interaction between STAC3 and RyR1. More recently, however, STAC3 was found to colocalize to the triads in the absence of CaV1.1, indicating that STAC3 could also directly interact with RyR1 and thus, mediate a functional interaction between CaV1.1 and RyR1. This study investigates the biophysical characteristics of STAC3 and its functional interaction with RyR1. Using single-channel bilayers, the tandem SH3 domains of STAC3 were found to interact with and activate RyR1. The introduction of the NAM mutation altered the secondary structure of STAC3SH3s and significantly reduced protein stability, with STAC3SH3s NAM failing to activate RyR1 channel activity. More importantly, the first SH3 domain of STAC3 was found to be critical for RyR1 activation. In addition, cross-linking experiments identified a second interaction between STAC3 and RyR1. A PxxP motif containing sequence was identified on RyR1, which bound to the second SH3 domain of STAC3 with micromolar affinity. Collectively, these findings provide the first direct evidence of an interaction between STAC3 and RyR1 and are in line with STAC3's recently proposed role as a functional link between CaV1.1 and RyR1. Moreover, these findings suggest that STAC3 could have two distinct functional interactions with RyR1, thereby having a highly dynamic role in the skeletal EC coupling machinery.|
|dc.title||The interactive role of STAC3 in the skeletal excitation-contraction coupling machinery|
|local.contributor.affiliation||John Curtin School of Medical Research, ANU College of Science, The Australian National University|
|Collections||Open Access Theses|
|Aditya PhD thesis.pdf||Thesis Material||23.14 MB||Adobe PDF|
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