Nemaline myopathy (NM) is a genetically heterogeneous congenital myopathy defined by muscle weakness and the presence of nemaline bodies—rod-like structures derived from Z-disk abnormalities in skeletal muscle fibers. While mutations in five genes encoding thin filament components have been established as causes of NM, this study identifies a sixth: CFL2, which encodes cofilin-2, a key regulator of actin dynamics in skeletal muscle. A homozygous c.103G>A mutation in CFL2 was identified in two affected siblings from a consanguineous Middle Eastern family. The mutation results in an alanine-to-threonine substitution at residue 35 (A35T), located near a nuclear localization signal within a highly conserved region of the protein.
Clinical presentation in both patients included neonatal hypotonia, delayed motor development, recurrent falls, and progressive ambulatory impairment. The elder sister, aged 16, walks short distances but uses a wheelchair outdoors. The younger sibling, the proband, was diagnosed with NM at age 2 after a muscle biopsy revealed nemaline bodies, occasional minicores, and concentric laminated bodies. Notably, no significant facial weakness or foot drop was observed, distinguishing the phenotype from some other forms of NM.
Histopathological analysis confirmed the presence of nemaline bodies on Gomori trichrome staining and minicores on NADH-tetrazolium reductase staining. Electron microscopy revealed unstructured minicores and concentric laminated bodies, suggesting distinct ultrastructural abnormalities. Immunofluorescence using anti-a-actinin-2 antibodies showed that nemaline bodies were positive for this Z-disk protein. Phalloidin staining demonstrated abnormal F-actin accumulations in 4% of myofibers, colocalizing with sarcomeric actin, indicating a primary defect in actin organization.
Despite normal ACTA1 sequencing, the presence of actin filament aggregates pointed toward a broader actinopathy. Quantitative analysis revealed that cofilin-2 levels were markedly reduced in the proband’s muscle tissue. Two-dimensional SDS-PAGE and immunoblotting confirmed a significant decrease in unphosphorylated cofilin-2, with no detectable phosphorylated form. Paradoxically, CFL2 mRNA levels were elevated 4- to 20-fold compared to controls, implicating post-transcriptional mechanisms such as protein instability or degradation.
To assess the functional impact of the A35T mutation, wild-type and mutant cofilin-2 proteins were expressed in both mammalian and bacterial systems. In eukaryotic cells, both proteins localized normally and formed cytoplasmic and nuclear rods upon overexpression. However, in E. coli, the A35T variant showed severe solubility defects—while equal amounts of protein were present in total lysates, the mutant failed to purify under standard conditions. Only after treatment with 6 M urea was the mutant protein recovered, indicating aggregation due to misfolding.
Structural modeling based on chicken cofilin-2 revealed that the T35 side chain clashed sterically with I55, disrupting the β-sheet structure essential for stability.Tributyltetradecyl phosphonium Biochemical Assay Reagents This predicted conformational change supports the hypothesis that the A35T mutation destabilizes the tertiary structure, leading to premature degradation and functional deficiency.Copper(II) perchlorate hexahydrate Biological Activity
The accumulation of F-actin in nemaline bodies, minicores, and laminated bodies likely stems from impaired actin depolymerization due to cofilin-2 deficiency.PMID:34634487 This study provides strong genetic and biochemical evidence linking CFL2 mutations to a distinct form of congenital myopathy characterized by multiple pathological features. It also highlights the importance of cofilin-2 in maintaining sarcomeric integrity and suggests that its dysfunction may underlie a subset of NM cases previously attributed to nonspecific pathology. Future research should focus on identifying additional CFL2 mutations and exploring targeted therapies aimed at restoring actin homeostasis.MedChemExpress (MCE) offers a wide range of high-quality research chemicals and biochemicals (novel life-science reagents, reference compounds and natural compounds) for scientific use. We have professionally experienced and friendly staff to meet your needs. We are a competent and trustworthy partner for your research and scientific projects.Related websites: https://www.medchemexpress.com