4-Phenylbutyrate restores localization and membrane repair to human dysferlin mutations

Kana Tominaga,Naoomi Tominaga,Eric O Williams,Laura Rufibach,Verena Schöwel,Simone Spuler,Mohan Viswanathan,Leonard P Guarente

doi:10.1016/j.isci.2021.103667

Abstract

SummaryDysferlinopathies are muscular dystrophies caused by recessive loss-of-function mutations in dysferlin (DYSF), a membrane protein involved in skeletal muscle membrane repair. We describe a cell-based assay in which human DYSF proteins bearing missense mutations are quantitatively assayed for membrane localization by flow cytometry and identified 64 localization-defective DYSF mutations. Using this platform, we show that the clinically approved drug 4-phenylbutryric acid (4-PBA) partially restores membrane localization to 25 mutations, as well as membrane repair to cultured myotubes expressing 2 different mutations. Two-day oral administration of 4-PBA to mice homozygous for one of these mutations restored myofiber membrane repair. 4-PBA may hold therapeutic potential for treating a subset of humans with muscular dystrophy due to dysferlin deficiency.

Highlights

Dysferlinopathy is an adult-onset, progressive, rare form of muscular dystrophy caused by recessive loss-offunction mutations in the gene encoding dysferlin, DYSF, and includes the clinical diagnoses of limb-girdle muscular dystrophy 2B/R2, Miyoshi myopathy type 1, and distal anterior compartment myopathy (Illa et al, 2001; Liu et al, 1998)
SUMMARY Dysferlinopathies are muscular dystrophies caused by recessive loss-of-function mutations in dysferlin (DYSF), a membrane protein involved in skeletal muscle membrane repair
We describe a cell-based assay in which human DYSF proteins bearing missense mutations are quantitatively assayed for membrane localization by flow cytometry and identified 64 localization-defective DYSF mutations

Summary

Introduction

Dysferlinopathy is an adult-onset, progressive, rare form of muscular dystrophy caused by recessive loss-offunction mutations in the gene encoding dysferlin, DYSF, and includes the clinical diagnoses of limb-girdle muscular dystrophy 2B/R2, Miyoshi myopathy type 1, and distal anterior compartment myopathy (Illa et al, 2001; Liu et al, 1998). A diagnosis of dysferlinopathy is made when a patient is shown to have >80% reduction in DYSF protein by western blot (Cacciottolo et al, 2011) and is confirmed by sequencing of the DYSF gene to identify the causative mutation(s). A tail-anchored type-2 integral plasma membrane (PM) protein, DYSF has a very short extracellular domain and a large intracellular domain containing 7 calcium-binding C2-domains (Abdullah et al, 2014), as well as FerA and DysF domains that together mediate intracellular membrane fusion events (Bansal and Campbell, 2004; Lek et al, 2012), calcium homeostasis (Kerr et al, 2014), and lipid metabolism (Haynes et al, 2019). Other reported roles for dysferlin include calcium regulation at the t-tubule triad junction (Kerr et al, 2014), vesicular trafficking (Bansal et al, 2003; Lek et al, 2012), and monocyte adhesion (de Morree et al, 2013)

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