Calpains, Cleaved Mini-Dysferlin_C72, and L-Type Channels Underpin Calcium-Dependent Muscle Membrane Repair

Abstract

Dysferlin is proposed as a key mediator of calcium-dependent muscle membrane repair, although its precise role has remained elusive. Dysferlin interacts with a new membrane repair protein, mitsugumin 53 (MG53), an E3 ubiquitin ligase that shows rapid recruitment to injury sites. Using a novel ballistics assay in primary human myotubes, we show it is not full-length dysferlin recruited to sites of membrane injury but an injury-specific calpain-cleavage product, mini-dysferlin C72 . Mini-dysferlin C72 -rich vesicles are rapidly recruited to injury sites and fuse with plasma membrane compartments decorated by MG53 in a process coordinated by L-type calcium channels. Collective interplay between activated calpains, dysferlin, and L-type channels explains how muscle cells sense a membrane injury and mount a specialized response in the unique local environment of a membrane injury. Mini-dysferlin C72 and MG53 form an intricate lattice that intensely labels exposed phospholipids of injury sites, then infiltrates and stabilizes the membrane lesion during repair. Our results extend functional parallels between ferlins and synaptotagmins. Whereas otoferlin exists as long and short splice isoforms, dysferlin is subject to enzymatic cleavage releasing a synaptotagmin-like fragment with a specialized protein-or phospholipid-binding role for muscle membrane repair.

Proper Voltage-Dependent Ion Channel Function in Dysferlin-Deficient Cardiomyocytes

“…Thus, it is unknown if dysferlin-deficiency impairs cardiac ion channel function, a potential pathophysiological mechanism in the dysferlinopathies. The following lines of evidence point to a potential regulatory effect of dysferlin on Ca v 1.2 channels: First, in skeletal muscle, dysferlin colocalizes with the L-type calcium channel [16,17], and muscle membrane repair requires an interplay between these two proteins [18]. Secondly, the dysferlin interaction partner AHNAK [19,20] associates with regulatory β-subunits of Ca v 1.2 channels [21,22], and thirdly, the α 2 δ 1 calcium channel subunit, which modulates the channel's function [23], is significantly downregulated in the dysferlin-deficient heart (NCBI GEO profile: http://www.ncbi.nlm.nih.gov/geoprofiles?term=GDS1247[ACCN]+cacna2d1, ref.…”

A fragmented form of annexin A1 is secreted from C2C12 myotubes by electric pulse-induced contraction

“…In similar examples, it was reported that dysferlin was also fragmented by calpain into ''mini-dysferlin'' and acted as a valid molecule for membrane repair [34]. We realized that it will be important in future analyses to test whether the culture medium after contraction possesses this healing effect.…”

Plasma membrane and cytoskeleton dynamics during single-cell wound healing

“…Indeed, fusion of dysferlin-rich vesicles to the plasmalemma necessitates prior decoration of the injury site by MG53. However, dysferlin recruitment to the membrane have been shown to occur before that of MG53-rich vesicle [43,114,119]. This is particularly puzzling, since dysferlin is also enriched in the sarcolemma and transverse tubules of uninjured muscle fibers [120].…”

“…Dysferlin-mediated fusion also necessitates processing of dysferlin by calpains into minidysferlinC72 [43,119,126]. Dysferlin processing do not involve the muscle-specific calpain-3, but rather µ-calpain and m-calpain, as Caspn3-null mice do not show signs of loss of sarcolemma resealing after needle scratch or laser damage [127].…”

“…[43,97,119,126,127,195,196] Ferlins Dysferlin -Muscle-cells of dysferlin-null mice fail to undergo Ca2+-dependent membrane resealing after laser-mediated injury.…”