MYL1-Related Congenital Myopathy: Clinical, Genetic and Pathological Insights

Abstract

Congenital myopathies and congenital muscular dystrophies encompass heterogeneous clinical and genetic groups of disorders characterised by muscle weakness with antenatal or early postnatal onset. These conditions are categorised according to distinctive myopathological features and causative genes. Despite advances in diagnosis through massive parallel sequencing and progress in understanding the underlying pathogenesis, many aspects of these disorders remain poorly understood. MYL1-related congenital myopathy is an ultra-rare and severe condition, associated with a deficiency of essential/alkali light myosin and impaired development of fast-twitch type II muscle fibres. This study aims to advance the understanding of the phenotype and pathogenesis of MYL1-congenital myopathy. We analysed the clinical characteristics of two individuals harbouring three novel variants in the MYL1 gene. We conducted detailed genomic analysis and extensive studies on their muscles using histological, immunohistochemical, immunofluorescence, Western Blot and electron microscopy. Both individuals showed a very severe congenital myopathy, characterised by congenital hypotonia and weakness, requiring ventilatory and nutritional assistance. Muscle biopsy revealed dystrophic-like or myopathic changes, with notable smallness of fast-twitch type II fibres, often arranged around larger type I fibres, drawing a floret pattern. These fibres expressed developmental myosin and exhibited features of aberrant myofibrillogenesis. Type I myofibres exhibited correct sarcomere alignment, but like the small fast-twitch fibres, both showed distorted cell organelles, vacuolar aggregates and membranous debris, indicating autophagic impairment. Our findings confirm that bi-allelic MYL1 variants are associated with a severe congenital myopathy, characterised by a distinctive clinical and histopathological phenotype involving impaired type II fibre development. Additionally, our study reveals a role for MYL1 in the organisation and trophism of all muscle fibre types. SUMMARY: MYL1 biallelic variants cause severe congenital myopathy with early hypotonia and type II fibre hypotrophy. Muscle biopsy shows a distinct pattern, including floret-like fibre arrangement. Findings suggest a broader role for MYL1 in fibre organisation and autophagy across muscle fibre types.

Keywords: congenital muscular dystrophy; congenital myopathy; essential/alkali myosin light chain; fast‐twitch type II muscle fibres; muscle cell autophagy; myosin heavy chain.

FIGURE 1

Clinical images of the two individuals with MYL1‐related congenital myopathy. (A) Thoraco‐abdominal radiograph of Individual 1 at birth, revealing a bilateral humerus fracture. The fracture is displaced on the left side and nondisplaced on the right side. (B) Lateral photograph of Individual 2 at 2 years and 4 months, demonstrating head control and the ability to elevate upper limbs against gravity. (C) Frontal photograph of Individual 2 at 2 years and 9 months, highlighting preserved strength in the upper limbs and hands compared to the lower limbs, as she is even able to propel her wheelchair.

FIGURE 2

In silico evaluation of MYL1 genetic variants found in Individuals 1 and 2. (A) The upper panel shows the MYL1 gene structure, which codifies the skeletal muscle fast‐twitch specific myosin essential light chain (ELC). ELC domain structure (Uniprot: P05976) is represented in the middle panel. Genetic variants found in Individuals 1 and 2 are indicated with red symbols, and previously reported variants are represented in black (in the upper panel if they are splicing variants and in the middle panel those that are missense or truncating). Previously reported variants (according to the Human Genome Mutation Database) are associated with severe congenital myopathy. The lower panel shows the tolerance landscape of ELC according to the MetaDome web server, indicating the positions of the truncating variants relative to regions of varying tolerance to variation. (B) Schematic drawing of protein interactions in striated muscle, highlighting the major myosin regulatory proteins. Abbreviations: ELC: myosin essential light chain; RLC: myosin regulatory light chain‐2; MyBP‐C: myosin‐binding protein.

FIGURE 3

Histopathological findings in the muscle samples of individuals with MYL1‐related congenital myopathy. Upper row pictures correspond to Propositus 1 and lower row pictures to Propositus 2. Haematoxylin‐eosin in Individuals 1 (A) and 2 (F) shows marked variability in the size of muscle fibres, with a population of hypertrophic fibres surrounded by small fibres. Fibrosis and fatty infiltration were more prominent in Individual 2, as observed in (F). Modified Gömöri trichrome staining in Individuals 1 (B) and 2 (G) shows an increase in endomysial and perimysial connective tissue. Dark purple granular inclusions (arrows) were identified in Individual 2 (G). SDH in Individual 1 (C) and NADH‐TR histochemistry in Individual 2 (H) showed uniform staining in larger fibres and a mixed pattern of dark and light in the smaller fibres. Fast myosin immunohistochemical staining of Individual 1 (D) showed that small fibres were fast fibres, and slow myosin immunohistochemical staining showed that all hypertrophic fibres of Individual 1 (E) were slow fibres. Similarly, ATPase 9.4 (I) and ATPase 4.6 (not shown) in Individual 2 showed that all small fibres corresponded to IIA or IIC with no presence of IIB/IIX, whereas the large fibres were all type I.

FIGURE 4

Immunofluorescence assays on muscle samples from individuals with MYL1‐related congenital myopathy The upper row (A‑D) shows immunofluorescence images showing the expression of slow myosin (MYH7) (A), fast myosin (MYH2A) (B), embryonic myosin (MYH3/MYHe) (C) and neonatal myosin (MYH8/MYHn) (D) in Individual 2. Note that MYH7 is mainly expressed in large fibres and a few scattered small fibres (A). MYH2A expression is restricted to small fibres (B), with a high proportion of these small fibres co‐expressing MYH8 (C), while only a small proportion of them also co‐expressed MYH3 (D). The lower row presents double immunofluorescence staining of MYL1 (green) and b‐laminin (red) in Individual 1 (E) and Individual 2 (F), compared to control muscles (G). In Individual 1 (E), a reduction in MYL1 expression is observed, while in Individual 2 (F), MYL1 expression is absent. In contrast, the control muscle (G) showed two intensities of immunostaining with a checkerboard distribution, with low or very low expression in type I fibres.

FIGURE 5

Electron microscopy of muscle samples from individuals with MYL1‐related congenital myopathy. The upper row (A‑C) shows electron micrographs of muscle samples from Individual 1. Small fibres (type II) show a range of myofibrillar abnormalities, with diversity in frequency and size, as well as disrupted sarcomeric organisation (A, B). The sarcoplasm of these small fibres contained areas of electron‐dense aggregates whose subcellular structure is indistinct. Some small fibres also showed centralised nuclei, without evidence of redundant basement membrane folding. Micrograph (C) shows a large fibre (type I) with well‐preserved myofibrils and aligned sarcomeric Z‐bands. Additionally, intermyofibrillar aggregates are visible, including elements with the shape of mitochondria. The middle and lower rows (D‑J) show electron micrographs from Individual 2. Micrograph (D) shows a collection of small muscle fibres with immature features embedded in a fibrous background. Micrographs (E) and (F) provide detailed views of a small muscle fibre, where the sarcoplasm contains sparse filamentous bundles attached to miniature Z‐disks, suggestive of myofibril precursors (black arrow). Additionally, membranous aggregates are visible, including vacuoles, complex membrane folding and myelin‐like features (white arrow). Micrograph (G) corresponds to a large type I myofibre with an invagination of the sarcolemma, suggestive of incipient fibre splitting. The fibre has well‐preserved myofilaments, proper sarcomere organisation and aligned Z bands. This large fibre depicts aggregates of vacuoles, degraded mitochondria and membranous structures of diverse sizes within the intermyofibrillar (H) and subsarcolemmal (I) spaces. In micrograph (J), proliferation of basal lamina folds (star) appears in the extracellular space spanning between a large type I fibre (large black arrow) and adjacent small myofibres with immature features (small black arrow). Finally, note the nuclear abnormalities in both small and large fibres, consisting of irregular shapes with infoldings and segmentations, abnormal chromatin distribution and nuclear envelope abnormalities.