Aberrantly spliced α-dystrobrevin alters α-syntrophin binding in myotonic dystrophy type 1

Abstract

Background: Myotonic dystrophy type 1 (DM1) is a multisystemic disorder caused by a CTG repeat expansion in the DMPK gene. Aberrant messenger RNA (mRNA) splicing of several genes has been reported to explain some of the symptoms in DM1, but the cause of muscle wasting is still unknown. By contrast, many forms of muscular dystrophy are caused by abnormalities of the dystrophin–glycoprotein complex (DGC). α-Dystrobrevin is a key component of the DGC in striated muscle and plays important roles in maturation and signal transduction by interacting with α-syntrophin. The goal of this study was to investigate alternative splicing of α-dystrobrevin in DM1 and examine α-syntrophin binding of different α-dystrobrevin splice isoforms.

Methods: Splicing patterns of α-dystrobrevin in DM1 muscle were studied by reverse-transcriptase PCR. Expression of the variant splice isoform was examined by immunoblotting and immunohistochemistry. Alternatively spliced isoforms were expressed in cultured cells to investigate interaction with α-syntrophin. α-Syntrophin expression was examined by immunoblotting.

Results: α-Dystrobrevin mRNA including exons 11A and 12 was increased in both skeletal and cardiac muscle of DM1 patients. The aberrantly spliced α-dystrobrevin isoform was localized to the sarcolemma, and showed increased binding with α-syntrophin. Furthermore, levels of α-syntrophin associated with the DGC were increased in DM1 muscle.

Conclusion: Alternative splicing of α-dystrobrevin is dysregulated in myotonic dystrophy type 1 (DM1) muscle, resulting in changes in α-syntrophin binding. These results raise the possibility that effects on α-dystrobrevin splicing may influence signaling in DM1 muscle cells.

GLOSSARY: α-DB = α-dystrobrevin; α-syn = α-syntrophin; ALS = amyotrophic lateral sclerosis; β-DG = β-dystroglycan; CC = coiled-coil domain; cDM = congenital myotonic dystrophy type 1; cDNA = complementary DNA; Cont = control; DAPC = dystrophin-associated protein complex; DBS = dystrophin binding site; DGC = dystrophin–glycoprotein complex; DM1 = myotonic dystrophy type 1; EF = EF hand region; GAPDH = glyceraldehyde 3-phosphate dehydrogenase; IP = immunoprecipitation; LGMD = limb-girdle muscular dystrophy; MDRS = muscular disability rating scale; mRNA = messenger RNA; NA = not available; NMJ = neuromuscular junction; NT = not tested; P = postnatal day; PM = polymyositis; RT-PCR = reverse-transcriptase PCR; SBS = syntrophin binding site; SDS-PAGE = sodium dodecyl sulfate polyacrylamide gel electrophoresis; TBS = Tris-buffered saline; WCL = whole-cell lysate; vr = variable region; Y = unique tyrosine kinase substrate domain; ZZ = zinc-binding domain.