Impaired Intracellular Ca2+ Dynamics, M-Band and Sarcomere Fragility in Skeletal Muscles of Obscurin KO Mice.

Enrico Pierantozzi,Beatrix Dienes,Barbara Colombini,Carlo Reggiani,Dilson E Rassier,Tamás Oláh,László Csernoch,Cecilia Paolini,Stephan Lange,Maria Angela Bagni,Vincenzo Sorrentino,János Fodor,Egidio Maria Rubino,Péter Szentesi,Daniela Rossi

doi:10.3390/ijms23031319

Abstract

Obscurin is a giant sarcomeric protein expressed in striated muscles known to establish several interactions with other proteins of the sarcomere, but also with proteins of the sarcoplasmic reticulum and costameres. Here, we report experiments aiming to better understand the contribution of obscurin to skeletal muscle fibers, starting with a detailed characterization of the diaphragm muscle function, which we previously reported to be the most affected muscle in obscurin (Obscn) KO mice. Twitch and tetanus tension were not significantly different in the diaphragm of WT and Obscn KO mice, while the time to peak (TTP) and half relaxation time (HRT) were prolonged. Differences in force-frequency and force-velocity relationships and an enhanced fatigability are observed in an Obscn KO diaphragm with respect to WT controls. Voltage clamp experiments show that a sarcoplasmic reticulum’s Ca2+ release and SERCA reuptake rates were decreased in muscle fibers from Obscn KO mice, suggesting that an impairment in intracellular Ca2+ dynamics could explain the observed differences in the TTP and HRT in the diaphragm. In partial contrast with previous observations, Obscn KO mice show a normal exercise tolerance, but fiber damage, the altered sarcomere ultrastructure and M-band disarray are still observed after intense exercise.

Highlights

The OBSCN gene encodes a giant sarcomeric protein, obscurin, originally identified as a titin-interacting protein [1]
We reported experiments aimed at determining the effects of obscurin deletion on the contractile properties and Ca2+ kinetics in ex vivo muscle fibers, along with additional experiments where we re-evaluated the running ability of Obscn KO mice and the effects of intense exercise on the stability of the contractile apparatus in diaphragm muscle
We reported that Obscn KO mice displayed a reduced sarcoplasmic reticulum volume, decreased localization of dystrophin at costameres, reduced exercise tolerance and sarcolemma and M-band fragility following intense exercise [16,23,27,35]

Summary

Introduction

The OBSCN gene encodes a giant sarcomeric protein, obscurin, originally identified as a titin-interacting protein [1]. Two high-molecular-weight obscurin variants, known as obscurin A (~720 kDa) and obscurin B (970–870 kDa), are present. Both proteins are characterized by a modular architecture consisting of tandem immunoglobulin (Ig) and fibronectin III (Fn3)-like domains, followed by some signaling motifs, including a calmodulin IQ-binding motif, a src homology (SH3) domain, a RhoGEF and a pleckstrin homology (PH) domain. These two variants differ at the C-terminal region, with obscurin. In addition to the two giant obscurin A and B, additional obscurin isoforms of a smaller molecular weight have been detected in non-muscle tissues, the physiological role of these non-muscular obscurin isoforms is not yet known [9]

Methods

Results

Conclusion