Dataset Information

Defects in G-Actin Incorporation into Filaments in Myoblasts Derived from Dysferlinopathy Patients Are Restored by Dysferlin C2 Domains.

ABSTRACT: Dysferlin is a transmembrane C-2 domain-containing protein involved in vesicle trafficking and membrane remodeling in skeletal muscle cells. However, the mechanism by which dysferlin regulates these cellular processes remains unclear. Since actin dynamics is critical for vesicle trafficking and membrane remodeling, we studied the role of dysferlin in Ca²⁺-induced G-actin incorporation into filaments in four different immortalized myoblast cell lines (DYSF2, DYSF3, AB320, and ER) derived from patients harboring mutations in the dysferlin gene. As compared with immortalized myoblasts obtained from a control subject, dysferlin expression and G-actin incorporation were significantly decreased in myoblasts from dysferlinopathy patients. Stable knockdown of dysferlin with specific shRNA in control myoblasts also significantly reduced G-actin incorporation. The impaired G-actin incorporation was restored by the expression of full-length dysferlin as well as dysferlin N-terminal or C-terminal regions, both of which contain three C2 domains. DYSF3 myoblasts also exhibited altered distribution of annexin A2, a dysferlin partner involved in actin remodeling. However, dysferlin N-terminal and C-terminal regions appeared to not fully restore such annexin A2 mislocation. Then, our results suggest that dysferlin regulates actin remodeling by a mechanism that does to not involve annexin A2.

SUBMITTER: Baez-Matus X

PROVIDER: S-EPMC6981584 | biostudies-literature | 2019 Dec

REPOSITORIES: biostudies-literature

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Defects in G-Actin Incorporation into Filaments in Myoblasts Derived from Dysferlinopathy Patients Are Restored by Dysferlin C2 Domains.

Báez-Matus Ximena X Figueroa-Cares Cindel C Gónzalez-Jamett Arlek M AM Almarza-Salazar Hugo H Arriagada Christian C Maldifassi María Constanza MC Guerra María José MJ Mouly Vincent V Bigot Anne A Caviedes Pablo P Cárdenas Ana M AM

International journal of molecular sciences 20191219 1

Dysferlin is a transmembrane C-2 domain-containing protein involved in vesicle trafficking and membrane remodeling in skeletal muscle cells. However, the mechanism by which dysferlin regulates these cellular processes remains unclear. Since actin dynamics is critical for vesicle trafficking and membrane remodeling, we studied the role of dysferlin in Ca<sup>2+</sup>-induced G-actin incorporation into filaments in four different immortalized myoblast cell lines (DYSF2, DYSF3, AB320, and ER) deriv ...[more]

PMID: 31861684

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Project description:Dysferlin is an integral membrane protein of the transverse tubules of skeletal muscle that is mutated or absent in limb girdle muscular dystrophy 2B and Miyoshi myopathy. Here we examine the role of dysferlin's seven C2 domains, C2A through C2G, in membrane repair and Ca2+ release, as well as in targeting dysferlin to the transverse tubules of skeletal muscle. We report that deletion of either domain C2A or C2B inhibits membrane repair completely, whereas deletion of C2C, C2D, C2E, C2F or C2G causes partial loss of membrane repair that is exacerbated in the absence of extracellular Ca2+ . Deletion of C2C, C2D, C2E, C2F or C2G also causes significant changes in Ca2+ release, measured as the amplitude of the Ca2+ transient before or after hypo-osmotic shock and the appearance of Ca2+ waves. Most deletants accumulate in endoplasmic reticulum. Only the C2A domain can be deleted without affecting dysferlin trafficking to transverse tubules, but Dysf-ΔC2A fails to support normal Ca2+ signalling after hypo-osmotic shock. Our data suggest that (i) every C2 domain contributes to repair; (ii) all C2 domains except C2B regulate Ca2+ signalling; (iii) transverse tubule localization is insufficient for normal Ca2+ signalling; and (iv) Ca2+ dependence of repair is mediated by C2C through C2G. Thus, dysferlin's C2 domains have distinct functions in Ca2+ signalling and sarcolemmal membrane repair and may play distinct roles in skeletal muscle. KEY POINTS: Dysferlin, a transmembrane protein containing seven C2 domains, C2A through C2G, concentrates in transverse tubules of skeletal muscle, where it stabilizes voltage-induced Ca2+ transients and participates in sarcolemmal membrane repair. Each of dysferlin's C2 domains except C2B regulate Ca2+ signalling. Localization of dysferlin variants to the transverse tubules is not sufficient to support normal Ca2+ signalling or membrane repair. Each of dysferlin's C2 domains contributes to sarcolemmal membrane repair. The Ca2+ dependence of membrane repair is mediated by C2C through C2G. Dysferlin's C2 domains therefore have distinct functions in Ca2+ signalling and sarcolemmal membrane repair.

Dataset Information

Defects in G-Actin Incorporation into Filaments in Myoblasts Derived from Dysferlinopathy Patients Are Restored by Dysferlin C2 Domains.

Publications

Defects in G-Actin Incorporation into Filaments in Myoblasts Derived from Dysferlinopathy Patients Are Restored by Dysferlin C2 Domains.

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