Dysferlin-mediated phosphatidylserine sorting engages macrophages in sarcolemma repair

Abstract

Failure to repair the sarcolemma leads to muscle cell death, depletion of stem cells and myopathy. Hence, membrane lesions are instantly sealed by a repair patch consisting of lipids and proteins. It has remained elusive how this patch is removed to restore cell membrane integrity. Here we examine sarcolemmal repair in live zebrafish embryos by real-time imaging. Macrophages remove the patch. Phosphatidylserine (PS), an ‘eat-me' signal for macrophages, is rapidly sorted from adjacent sarcolemma to the repair patch in a Dysferlin (Dysf) dependent process in zebrafish and human cells. A previously unrecognized arginine-rich motif in Dysf is crucial for PS accumulation. It carries mutations in patients presenting with limb-girdle muscular dystrophy 2B. This underscores the relevance of this sequence and uncovers a novel pathophysiological mechanism underlying this class of myopathies. Our data show that membrane repair is a multi-tiered process involving immediate, cell-intrinsic mechanisms as well as myofiber/macrophage interactions.

Muscle cell communication in development and repair

“…PS is normally found within the inner lipid bilayer and flips to the outer bilayer with plasma membrane disruption during injury, and this configuration serves as a signal to invading macrophages to remove the damaged myofiber [6]. However, external PS need not always signaling injury, as PS also localizes to the site of cell-cell contact during myoblast fusion and at the site of membrane repair [7-11]. Blocking PS exposure with an anti-PS antibody inhibits muscle cell fusion [9,12] indicating the necessity of external PS in cell-cell communication during membrane fusion.…”

“…With high-resolution live-cell imaging of multiple components mediating the repair process, dysferlin was observed to localize in the membrane immediately adjacent to sites of disruption [10]. This position may indicate that dysferlin regulates membrane incorporation at the site of damage through its interaction with phospholipids and other repair proteins [11]. …”

Cellular mechanisms and signals that coordinate plasma membrane repair

“…Following plasma membrane injury in Xenopus oocytes and zebrafish skeletal muscle, PS localizes directly adjacent to the wound site [86]. PS translocation in zebrafish muscle is guided by the arginine-rich PS binding region of dysferlin [133]. However, the details of how dysferlin itself translocates to the injury site and how it helps localize PS require further investigation.…”

Dysferlin mediates membrane tubulation and links T-tubule biogenesis to muscular dystrophy

“…C2-domain proteins often promote Ca 2+ -dependent membrane interaction and fusion (Lemmon, 2008), and the C2A domain of dysferlin binds to phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate [PI(4,5)P 2 ] (Davis et al, 2002;Fuson et al, 2014;Therrien et al, 2009), which is an important regulator of cellular trafficking, ion channels and Ca 2+ homeostasis (Balla, 2013;Hilgemann et al, 2001). The first C2 domain, C2A, is essential for phosphatidylserine accumulation at the membrane repair site (Middel et al, 2016). Dysferlin localizes to the plasma membrane and to the T-tubule membrane in striated muscle (Ampong et al, 2005;Anderson et al, 1999;Klinge et al, 2008), and transits to late endosomes, similar to myoferlin (Redpath et al, 2016).…”