C-terminal residues of skeletal muscle calsequestrin are essential for calcium binding and for skeletal ryanodine receptor inhibition.

Nicole A Beard,Angela F Dulhunty

doi:10.1186/s13395-015-0029-7

Nicole A Beard, Angela F Dulhunty

Open Access

https://doi.org/10.1186/s13395-015-0029-7

Copy DOI

Abstract

BackgroundSkeletal muscle function depends on calcium signaling proteins in the sarcoplasmic reticulum (SR), including the calcium-binding protein calsequestrin (CSQ), the ryanodine receptor (RyR) calcium release channel, and skeletal triadin 95 kDa (trisk95) and junctin, proteins that bind to calsequestrin type 1 (CSQ1) and ryanodine receptor type 1 (RyR1). CSQ1 inhibits RyR1 and communicates store calcium load to RyR1 channels via trisk95 and/or junctin.MethodsIn this manuscript, we test predictions that CSQ1’s acidic C-terminus contains binding sites for trisk95 and junctin, the major calcium binding domain, and that it determines CSQ1’s ability to regulate RyR1 activity.ResultsProgressive alanine substitution of C-terminal acidic residues of CSQ1 caused a parallel reduction in the calcium binding capacity but did not significantly alter CSQ1’s association with trisk95/junctin or influence its inhibition of RyR1 activity. Deletion of the final seven residues in the C-terminus significantly hampered calcium binding, significantly reduced CSQ’s association with trisk95/junctin and decreased its inhibition of RyR1. Deletion of the full C-terminus further reduced calcium binding to CSQ1 altered its association with trisk95 and junctin and abolished its inhibition of RyR1.ConclusionsThe correlation between the number of residues mutated/deleted and binding of calcium, trisk95, and junctin suggests that binding of each depends on diffuse ionic interactions with several C-terminal residues and that these interactions may be required for CSQ1 to maintain normal muscle function.

Highlights

Skeletal muscle function depends on calcium signaling proteins in the sarcoplasmic reticulum (SR), including the calcium-binding protein calsequestrin (CSQ), the ryanodine receptor (RyR) calcium release channel, and skeletal triadin 95 kDa and junctin, proteins that bind to calsequestrin type 1 (CSQ1) and ryanodine receptor type 1 (RyR1)
We have previously suggested that despite CSQ1 causing a modest activation of purified RyR1 in the bilayer, it is the overwhelming inhibition of native RyR1 which drives the overall effect of CSQ1 in the cell, which would acts as a brake on SR Ca2+ release
CSQ1 is both an inhibitor of native RyR1 under resting conditions and a luminal Ca2+ sensor for the channel, so that it may act as a brake on RyR1 Ca2+ release in times of low store load [7,9]

Summary

Introduction

Skeletal muscle function depends on calcium signaling proteins in the sarcoplasmic reticulum (SR), including the calcium-binding protein calsequestrin (CSQ), the ryanodine receptor (RyR) calcium release channel, and skeletal triadin 95 kDa (trisk95) and junctin, proteins that bind to calsequestrin type 1 (CSQ1) and ryanodine receptor type 1 (RyR1). Ryanodine receptor type 1 (RyR1) is central to excitation-contraction coupling, and its activity and ability to release Ca2+ is refined by the level of Ca2+ load inside the SR and by a luminal SR protein complex, including calsequestrin (CSQ), skeletal triadin. Calsequestrin type 1 (CSQ1) is the only isoform expressed in fast twitch muscle fibers, while equal amounts of CSQ1 and the so-called cardiac CSQ2 isoform are expressed in slow twitch fibers [2] Both isoforms display a high degree of homology, with the C-terminal tail extended in CSQ2. CSQ1 is a low affinity, moderate to high-capacity Ca2+-binding protein, binding between

Methods

Results

Discussion

Conclusion