Abstract
Dysferlin is a multi-C2 domain transmembrane protein involved in a plethora of cellular functions, most notably in skeletal muscle membrane repair, but also in myogenesis, cellular adhesion and intercellular calcium signaling. We previously showed that dysferlin interacts with alpha-tubulin and microtubules in muscle cells. Microtubules are heavily reorganized during myogenesis to sustain growth and elongation of the nascent muscle fiber. Microtubule function is regulated by post-translational modifications, such as acetylation of its alpha-tubulin subunit, which is modulated by the histone deacetylase 6 (HDAC6) enzyme. In this study, we identified HDAC6 as a novel dysferlin-binding partner. Dysferlin prevents HDAC6 from deacetylating alpha-tubulin by physically binding to both the enzyme, via its C2D domain, and to the substrate, alpha-tubulin, via its C2A and C2B domains. We further show that dysferlin expression promotes alpha-tubulin acetylation, as well as increased microtubule resistance to, and recovery from, Nocodazole- and cold-induced depolymerization. By selectively inhibiting HDAC6 using Tubastatin A, we demonstrate that myotube formation was impaired when alpha-tubulin was hyperacetylated early in the myogenic process; however, myotube elongation occurred when alpha-tubulin was hyperacetylated in myotubes. This study suggests a novel role for dysferlin in myogenesis and identifies HDAC6 as a novel dysferlin-interacting protein.
Highlights
Recessive mutations in the DYSF gene cause Limb girdle muscular dystrophy type 2B (LGMD2B) [1], Miyoshi Myopathy [1] and Distal anterior compartment myopathy [2]
Recombinant dysferlin was able to bind either to recombinant FLAG-histone deacetylase 6 (HDAC6) expressed in HEK293T cells (Figure 1A) or to native HDAC6 from homogenized murine testes (Figure 1B), which are a rich source of the enzyme
Dysferlin is a multi-C2 domain transmembrane protein involved in skeletal muscle membrane repair, and has been implicated in myogenesis, cellular adhesion and intercellular calcium signaling
Summary
Recessive mutations in the DYSF gene cause Limb girdle muscular dystrophy type 2B (LGMD2B) [1], Miyoshi Myopathy [1] and Distal anterior compartment myopathy [2]. Dysferlin is a large type II transmembrane protein composed of two DysF domains and seven C2 domains that mediate lipid [3,4] and protein binding interactions [5,6,7,8,9]. Dysferlin is a critical component of the calcium-dependent sarcolemmal repair complex, but recent studies have proposed additional roles for dysferlin in myogenesis [15,16,17], intercellular calcium signaling [18] and cellular adhesion [19]. Our recent work identified alpha-tubulin and microtubules as novel binding partners of dysferlin [6], suggesting a possible role for dysferlin in microtubule dynamics or stability
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