Zebrafish models for nemaline myopathy reveal a spectrum of nemaline bodies contributing to reduced muscle function.

Tamar E Sztal,Phillip I Bird,Nigel G Laing,Michaela Yuen,Sandra T Cooper,Derek L Stemple,Elisabeth M Busch-Nentwich,Aminah Giousoh,Caitlin Williams,Adam C Parslow,Mo Zhao,Robert J Bryson-Richardson,Adam Costin,Viola Oorschot,Georg Ramm,Thomas E Hall,Peter D Currie,Kristen J Nowak

doi:10.1007/s00401-015-1430-3

Abstract

Nemaline myopathy is characterized by muscle weakness and the presence of rod-like (nemaline) bodies. The genetic etiology of nemaline myopathy is becoming increasingly understood with mutations in ten genes now known to cause the disease. Despite this, the mechanism by which skeletal muscle weakness occurs remains elusive, with previous studies showing no correlation between the frequency of nemaline bodies and disease severity. To investigate the formation of nemaline bodies and their role in pathogenesis, we generated overexpression and loss-of-function zebrafish models for skeletal muscle α-actin (ACTA1) and nebulin (NEB). We identify three distinct types of nemaline bodies and visualize their formation in vivo, demonstrating these nemaline bodies not only exhibit different subcellular origins, but also have distinct pathological consequences within the skeletal muscle. One subtype is highly dynamic and upon breakdown leads to the accumulation of cytoplasmic actin contributing to muscle weakness. Examination of a Neb-deficient model suggests this mechanism may be common in nemaline myopathy. Another subtype results from a reduction of actin and forms a more stable cytoplasmic body. In contrast, the final type originates at the Z-disk and is associated with myofibrillar disorganization. Analysis of zebrafish and muscle biopsies from ACTA1 nemaline myopathy patients demonstrates that nemaline bodies also possess a different protein signature. In addition, we show that the ACTA1D286G mutation causes impaired actin incorporation and localization in the sarcomere. Together these data provide a novel examination of nemaline body origins and dynamics in vivo and identifies pathological changes that correlate with muscle weakness.Electronic supplementary materialThe online version of this article (doi:10.1007/s00401-015-1430-3) contains supplementary material, which is available to authorized users.

Highlights

Nemaline myopathy is a congenital skeletal muscle disease characterized by muscle weakness and low muscle tone, except in a few rare patients with hypertonia [20]
We describe the generation of zebrafish ACTA1 nemaline myopathy models allowing us to investigate disease pathogenesis in vivo
Through the overexpression of an eGFP-tagged ACTA1 variant or knockdown of α-actin in the skeletal muscle, we observed the formation of three distinct types of nemaline bodies

Summary

Introduction

Nemaline myopathy is a congenital skeletal muscle disease characterized by muscle weakness and low muscle tone, except in a few rare patients with hypertonia [20]. Mutations in ten different genes cause nemaline myopathy: skeletal muscle α-actin (ACTA1) [31], nebulin (NEB) [40], α-tropomyosin [24], β-tropomyosin [9], troponin T1 [22], cofilin 2 (CFL2) [1], KBTBD13 [45], KLHL40 [43], KLHL41 [13] and leiomodin 3 (LMOD3) [58]. All of these genes either encode components of the thin filament or regulate thin filament organization and stability. Deletion of exon 55, causing a common form of autosomal recessive nemaline myopathy [26], results in shortened thin filaments, alterations in cross-bridge cycling kinetics, and reduced calcium sensitivity [37, 38]

Methods

Results

Conclusion