Biallelic Variants in SNUPN Cause a Limb-Girdle Muscular Dystrophy with Myofibrillar Features

Academic Background

Muscular dystrophies are a complex and heterogeneous group of neuromuscular disorders characterized by progressive muscle weakness and atrophy due to loss of muscle fibers. Limb-Girdle Muscular Dystrophies (LGMD) are a subtype primarily affecting proximal muscles. Genetic variations are the main cause of these diseases, typically affecting proteins crucial for various muscle functions. Interestingly, genes associated with RNA splicing have also been identified as causes of certain types of muscular dystrophies. Despite the increasing number of associated genes and technological advancements, about half of LGMD patients still lack a definitive genetic diagnosis. Pre-mRNA splicing is a key step in generating mature mRNA transcripts, regulating gene expression through intron removal and exon ligation. Splicing is performed by a large dynamic ribonucleoprotein (RNP) complex—the spliceosome—mainly comprising U-rich small nuclear ribonucleoproteins (snRNPs). These include U1, U2, U4/U6, and U5 snRNPs.

Research Source

This paper was completed by an international research team, including authors Pablo Iruzubieta, Alberto Damborenea, Mihaela Ioghen, and others, affiliated with several top global research institutions. The study was published in the journal ‘Brain’, with online pre-publication on February 15, 2024.

Research Workflow

Research Subjects and Genetic Variant Analysis

This paper describes five patients from two unrelated families carrying biallelic variants in the SNUPN gene. SNUPN encodes the protein snurportin-1, which plays a crucial role in the nuclear transport of small nuclear ribonucleoproteins (snRNPs). Through whole-exome sequencing (WES) analysis, the study identified that patients from both families carried the c.926T>G; p.Ile309Ser allelic variant. This variant is extremely rare, with a frequency of only 0.000005472 in the gnomADv4 database.

Clinical and Imaging Features

Patients exhibited proximal weakness, restricted respiratory function, and significant contractures, although the severity varied among individuals within the same family. Muscle biopsies showed myofibrillar features, including myotilin deposits and Z-disc disorganization. MRI revealed severe involvement of the back muscles, sartorius, gracilis, peroneal muscles, and medial gastrocnemius.

Functional Experiments and Cellular Models

Further functional studies were conducted on patient-derived fibroblasts and muscle biopsies, revealing accumulation of snRNP complexes in the cytoplasm. RNA sequencing uncovered widespread RNA splicing dysregulation in patient muscles, particularly in genes involved in muscle development and splicing factors participating in early spliceosome assembly.

Drosophila Model Research

Additionally, the study used a Drosophila model for in vivo validation, further supporting the importance of snurportin-1 in muscle. Phagocytosis results showed that snurportin-1 downregulation affected the flies’ motor ability and lifespan.

Main Results

1. Clinical Presentation: Patients presented with childhood-onset proximal muscle weakness, restrictive respiratory dysfunction, and frequent contractures. MRI showed a characteristic pattern of muscle damage.
2. Pathological Features: Muscle biopsies showed myofibrillar features such as myotilin and p62 aggregation, and Z-disc disorganization. Electron microscopy also revealed focal myofibrillar fragmentation and mitochondrial depletion.
3. Molecular Biology Validation: In patient-derived fibroblasts and muscles, cytoplasmic accumulation of snRNP complexes was observed, although total snRNP and snurportin-1 expression levels remained unchanged.
4. RNA Splicing Dysregulation: RNA sequencing revealed widespread splicing dysregulation in genes related to muscle development, particularly prominent in genes encoding spliceosome components.
5. In Vivo Validation: Experiments in Drosophila further validated the critical impact of snurportin-1 on muscle development and function.

Research Conclusions

The study demonstrates that SNUPN gene variants are a new cause of a novel limb-girdle muscular dystrophy with specific clinical, pathological, and imaging features. This research enhances our understanding of the importance of splicing-related proteins in muscle diseases. This discovery provides a new target for genetic testing in patients and reveals the crucial role of snRNP transport and RNA splicing in muscle pathology.

Research Highlights

1. Confirmation of a New Transport Protein Variant: First confirmation of snurportin-1 as a pathogenic gene for limb-girdle muscular dystrophy, enriching the molecular genetic spectrum of muscular dystrophies.
2. Specific Pathological and Imaging Features: The study describes in detail the unique pathological and imaging features of this new classification, aiding in early clinical detection and diagnosis.
3. Cellular and Drosophila Model Validation: Comprehensive validation of snurportin-1’s function in muscle and the pathological changes caused by its variants through a combination of in vitro cellular experiments and in vivo Drosophila models.

Significance of the Research

This study not only discovers a new type of limb-girdle muscular dystrophy but also further reveals the important role of RNA splicing in muscle diseases, providing new directions and tools for future basic research and clinical diagnosis. As more family cases are reported, we will gain a more comprehensive understanding of the clinical phenotype and molecular mechanisms of SNUPN-related LGMD. Therefore, the SNUPN gene should be included in genetic testing programs for patients with myopathies.