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DYSTROPHINOPATHIES

Clinical syndromes   Becker   Cardiomyopathy   Cramps   Duchenne   Female carrier   Mental retardation   Microdeletion   Outliers Dystrophin gene   Mutations   Genotype vs Phenotype Dystrophin protein   Related glycoproteins Treatments Pathology   Duchenne   Becker Dystrophin: External link

From: A Kornberg Duchenne muscular dystrophy Standing from supine position

From Bramwell: Diseases of the Spinal Cord

Dystrophinopathies: Clinical

Clinical Syndromes   Becker   Cardiomyopathy   Cramps   Duchenne     Female carrier   Mental retardation   Microdeletion   Outliers Laboratory Treatments

From NLM Guillaume-Benjamin   Duchenne de Boulogne

De la Paralysie Musculaire Pseudo-hypertrophique... 1868

Clinical syndromes: General Weakness: Proximal > Distal; Symmetric Cardiomyopathy Serum CK: Usually very high Genotype-Phenotype correlation Duchenne muscular dystrophy Epidemiology Incidence: 1:3,600–9,300 live male births Newborn screening 36: CK levels at 12 days to 1 month = 1,800 to 5,400 Genotype: Dystrophin 96% with frameshift mutation 30% with new mutation 10% to 20% of new mutations are gonadal mosaic Clinical Weakness Onset age: 2 to 5 yrs Distribution Proximal > Distal Symmetric Legs & Arms Most involved muscles: Adductor magnus in legs Relatively spared muscles: Gracilis & Sartorius Course Reduced motor function by 2 to 3 years Steady decline in strength: After 6 to 11 years Gowers sign Standing up with the aid of hands pushing on knees External link: YouTube Loss of Ambulation Age: 9 - 13 years Later with: Steroid treatment External link: YouTube Muscle hypertrophy Especially calf May be generalized Increases with age Most commonly due to: Muscle replacement by fat & connective tissue Some relatively spared muscles may have true hypertrophy Musculoskeletal Contractures Especially ankles; Also hips & knees Treatment Non-surgical Night splints on ankles: More effective than passive stretch 3 Surgical Contracture release of ankles, knees or hips Early ambulation after surgery Scoliosis Onset: Most after loss of ambulation Partial relation to ambulation & growth status May be reduced if walking & standing are prolonged to ≥ 16 to 18 years Treatment: Surgical insertion of spinal rod Evaluation: X-ray; Sitting scoliosis series Timing of surgery: Usually before primary curve is 25% Vital capacity: Ahould be ≥ 40% of predicted Other clinical features Cardiomyopathy: Dilated; Especially > 15 years Mental retardation: Mean IQ ~ 88 Night blindness Altered response to flashes of light in dark adapted state ERG: b-wave, Reduced amplitude Dp260: Isoform of dystrophin in retina Gastrointestinal: Rare Type: Pseudo-obstruction; Gastric dilatation Upper GI tract Late in disease course Death Most common between 15 - 25 years Due to respiratory or cardiac failure Life prolonged by ~ 6 years to 25 years with respiratory support 8 Life shortened by 2 years with cardiomyopathy Laboratory Serum CK: Very high Highest: Ambulant patients; Up to 100x upper limit of normal Lower Increasing age: Less correlation between age & CK in Becker MD Lowest: Late stage disease; Severe muscle atrophy Birth: Increases over weeks after birth Troponin I: Elevated above normal but not to levels in cardiac ischemia Liver enzymes 18 High AST, ALT & LDH Up to 23x upper limit of normal Levels linearly correlate with CK level Normal: Alkaline phosphatase, γ-glutamyl transferase Muscle biopsy Endomysial fibrosis Variable fiber size: Small fibers rounded Muscle fiber necrosis & regeneration Myopathic grouping Hypercontracted (opaque) muscle fbers Muscle fiber internal archetecture: Normal or immature Dystrophin: Absent staining Other membrane proteins Sarcoglycans: Reduced Aquaporin 4: Reduced; Varied levels Diagnostic & Lab testing Muscle dystrophin Histochemical staining: Dystrophin protein absent in muscle Weatern blot: Dystrophin protein absent Genetic: Deletion, Duplication, Small mutation, Point mutation 10 Southern blot: Detects large deletions Sequencing of gene: Required to detect point & other small mutations MRI External link Treatments Prednisone 28 Doses Weekly dosing 6 Total dose: 5 to 10 mg/kg/week starting dose Dosing schedule: 2.5 to 5 mg/kg/day on Friday & Saturday evenings Side effects: Fewer than daily prednisone Weight gain: Some patients; Less than daily prednisone Cushingoid features: Mild Irritability: On day of dose Growth Not prominently impaired More growth (1" per year) than with daily prednisone Daily dosing Dose: 0.75 mg/kg/day starting dose Less optimal benefit:risk ratio than weekly regimen Effects of treatment Walking: Prolonged by 2 to 5 years Strength: Increased Falling: Reduced Pulmonary function: Improved Most beneficial While patient still ambulatory ? When started at early age (3 to 5 years) Scoliosis: May prolong walking long enough to reduce likelihood or severity Deflazacort Dose: 0.9 to 1.2 mg/kg/day starting dose Side effects Weight gain: Frequent; Less than daily prednisone Vertebral fractures: High frequency; Up to 100% by 9 years Short stature 27 Antisense oligonucleotides Exon 51 skip (Eteplirsen) Genetic effect: Change out-of-frame to in-frame deletion Dystrophin effect: Mild increase Frameshift deletions with possible benefit General: Must include exons 50-51 or 51-52 Specific deletions: Exon 50; 52; 43-50; 45-50; 47-50; 48-50; 49-50 External link: Skipping tool Clinical effects Slower decline in respiratory & arm function May not benefit cardiac function Trials 33 Other ASO: Exons 45; 53 (Vyondys) Gene replacement Minidystropphin Microdystrophin 34 Dystrophin expression: Present in muscle fibers Clinical: Functional improvement Serum CK: Reduced Transgene exons Sarepta/Roche: 1-17; 59-71 Genethon: 1-17; 59-70 Pfizer: 1-13; 50-51; 56-70 Solid: 1-11; 42-45; 57-71 Possible immunogenic Hinge 1 region: Exons 8 to 11 Utrophin: Up regulation Nonsense mutation read through: Gentamycin; Ataluren Myostatin or ActRIIB inhibition ? Oxandrolone: 0.1 mg/kg/day Cardiac Exon skippable mutations & Frequencies Exons: 8 (4%), 44 (8%), 45 (13%), 50 (5%), 51 (15%), 52 (3%), 53 (9%), 55 (2%) Exon 8 & 44 skippable mutations: Longer ambulation, especially 3 to 7 & 45 deletions Exon 44 skippable mutations: More revertant fibers in muscle Exon 51 skippable mutations: Shorter ambulation External link: Duchenne biography

DMD: Hypertrophic leg muscle Gowers sign Dystrophic muscle (Erb) DMD MRI: Adductor magnus involvement

Becker muscular dystrophy Genotype: Dystrophin mutations Deletion In-frame Frequency: 70% of patients Most common deletion locations: Exons 45-47 or 45-48 Milder muscle phenotype: Exons 45-55 38 Deletion location: Central rod domain Weakness: Less than average Other: May have Cardiomyopathy Cognitive Δ ↓ Life expectancy Other locations: 45-49; 45-53; 10-12; 48-49; 49-51 Frameshift mutation: 16% Point mutations > 70 identified Mutations in CpG All C to T; None G to A ? Related to direct or inverted gene repeats Nonsense mutations associated with milder BMD phenotype 21 Mutation Locations DMD exons prone to mutation-induced exon skipping Common: Exons 25, 27, 29, 31, 37, 38 Other: 43, 51, 56, 76 Mechanism: Altered splicing Worse course: Additional mutation in Myogenic factor 6 (MYF6) New mutations: Rare Clinical Myopathy Onset age: Usually > 7 yrs Weakness Proximal > Distal; Symmetric; Legs & Arms May be especially prominent in quadriceps or hamstrings Course of disability Slow progression Especially after adolescence Loss of ambulation: 4th to 6th decade Childhood & Adolescence: May be stable Relation to muscle dystrophin levels Severity Onset age Calf pain on exercise Muscle hypertrophy: Especially calves Failure to walk 16 - 80 years Systemic Joint contractures: Ankles & Other Cardiomyopathy: May occur before severe weakness Mental retardation Associated with deletion of Dp140 transcription unit Laboratory Serum CK Very high: 5,000 to 20,000 Lower levels: With increasing age & disability Muscle biopsy Myopathic Muscle fiber size: Varied; Small, Normal & Large Endomysial connective tissue: Increased Myopathic grouping Clustered necrotic or regenerating muscle fibers More common: Patients < 12 years of age Necrosis Regeneration: Immature muscle fibers Reduced dystrophin staining Rule out 2° change due to Sarcoglycanopathy External link: Becker biography

Becker MD Toe walking Enlarged calves From: A Kornberg MD From: Dejerine, J. Semiologie des affections du systeme nerveux.   Paris: Masson et Cie (pub.) 1914



• Outliers
  • Intermediate between Duchenne & Becker
  • Failure to walk 12 - 16 years
  
  
• Cramp & myalgia syndromes
  • Onset in childhood
  • Symptoms related to exercise
  • Normal or near-normal strength
  • Occasional rhabdomyolysis
  • CK: Persistently high CK (2,000 to 20,000) in most; Rarely normal
  
  
• Dystrophinopathies: Cardiomyopathy ⁵
  • General features
    • Nosology: X-linked Dilated Cardiomyopathy
    • Onset: Commonly in teenage years
    • Types: Dilated cardiomyopathy > Hypertrophic > Conduction defects
    • Possible biomarker: ST2 ³²
    • Probable pathophysiology: Dystrophin deficiency
      • Dystrophin deficient cardiomyocytes have reduced threshold for development of sarcolemmal injury under mechanical stress
      • Important location: Cardiomyocytes rather than cardiac vasculature ⁹
    • Also see: Cardiomyopathies
  • Clinical types
    • Cardiomyopathy with Becker or Duchenne MD syndromes: Common
      • Common mutations
        • Deletions in Exons 45-55 region (Spectrin-like region)
        • Mutations within exons 45 to 55 with dystrophin protein lacking hinge 3: Later-onset DCM
        • Out of phase mutations affecting exons 45 to 49: Risk of earlier DCM onset (2nd or 3rd decades)
      • Clinical features
        • Tachycardia: > 100; Common even < 10 years of age
        • Dilated cardiomyopathy: May develop after period of hypertrophy; Increased frequency with age
        • Conduction system abnormalities: Occasional patients
        • Symptomatic: 57% by age 18
        • Progression: Variable
        • Weakness: Typical Becker or Duchenne syndrome
        • May be associated with only: Myalgias after exercise & High CK
        • Treatments ¹⁹
          • Effects
            • 10 year survival: 2x Increased
            • Hospitalizations: 5x Reduced
          • Drugs
            • ACE inhibitors
            • Type 1 angiotensin II–receptor blockers (ARBs)
            • β-blockers
            • Corticosteroids
      • Laboratory
        • EKG
          • Changes due to atrophy of posterobasal region & lateral wall of left ventricle
          • Features
            • Anterior shift of the QRS
            • Tall right precordial R waves
            • Moderately deep Q waves in lead aVL
          • Age association
            • Age 10: EKG changes in 60%; No clinical manifestations
            • Age 18: EKG or ECHO changes in most
        • ECHO indications: Test Duchenne MD patients at age 13 to 14
        • Cardiovascular magnetic resonance with late gadolinium enhancement
          • May detect cardiac fibrosis before ECHO
        • Pattern of cardiac involvement
          • Early: Left ventricular free wall
          • Late: Right ventricular free wall; Atria
        • Cardiac pathology (Becker patients): Dystrophin reduced but not
    • Cardiomyopathy: Becker MD with deletions affecting exons 2 to 9
      • Dilated cardiomyopathy
      • Onset age (Ejection fraction < 55%): Earlier; Mean 23 years; Range 10 to 40 years
    • Selective Cardiomyopathy (CMD3B ): Minimal or mild weakness
      • Mutations
        • General features
          • Effects: Often affect splicing
          • Location: Most at 5' end of dystrophin gene
        • Muscle promoter & exon 1: Deletions & Point mutations
          • Cardiac muscle dystrophin
            • Loss of full length dystrophin M isoform
            • No up regulation of other dystrophin isoforms
          • Skeletal muscle dystrophin
            • Absent dystrophin muscle (M) isoform expression
            • Increased expression of brain (B) & cerebellar Purkinje (CP) isoforms
              • Upregulated by dystrophin muscle enhancer 1 (DME1) ⁴
                    Muscle-specific enhancer in dystrophin gene
              • B & CP isoforms are full length (427 kDa) like the M isoform
        • Intron 11: Alu-like sequence rearrangement
          • Alu-like sequence: 136 bp
          • Contains many stop codons
          • Effects of mutation
            • Cardiac muscle: Absent dystrophin splicing
            • Skeletal muscle: Residual dystrophin splicing
        • Exon 29
          • Nonsense mutation
          • Exon skipping
          • Affects dystrophin expression only in heart
        • Other mutations
          • Exon 2-7: Duplication
          • Exon 9: Hinge 1 region
      • Clinical features
        • Weakness: Minimal or Mild
        • Cardiomyopathy: Males
          • Onset: Late teens
          • Congestive heart failure
          • Rapidly progressive: 1 to 2 years
          • More severe cardiomyopathy with mutations at 5' end of dystrophin gene
        • Cardiomyopathy: Manifesting females
          • Onset: 5th decade
          • Congestive heart failure
          • Chest pain: Atypical
          • Slowly progressive over > 10 years
      • Differential diagnosis: X-linked Dilated Cardiomyopathy with Neutropenia
  
  
• Dystrophinopathies: Manifesting female carriers ³¹
  • Epidemiology
    • Symptomatic female carriers: 0.43 per 100,000 inhabitants in UK
  • Genetics
    • Mechanism
      • Normal X chromosome: Non-random inactivation
      • Detected by: Methylation status of the androgen receptor (AR) gene
    • Frequency in carriers
      • Duchenne-like 3% to 19%
      • Becker 14%
      • Cardiomyopathy 5% to 19%
      • Serum CK high 45%
    • Mutation types
      • Deletions: Less common (47%) than in dystrophinopathy males (66%)
      • No relation between: Site of mutation & Presence of weakness
      • Out of frame deletion: 91% of symptomatic carriers
  • Clinical
    • Weakness
      • Onset age: 1 to 62 years; Median 6 years
      • Course: Progressive; Mild to Severe
      • Distribution
        • Proximal
        • Asymmetric
        • Often
          • Arms & Legs
          • Legs > Arms
        • Shoulder abduction; Elbow flexion; Knee extension
        • Other occasional: Camptocormia ²³
      • Female Duchenne phenotype
        • Onset age: 1 to 5 years; Median 3 years
        • Severe weakness
        • Muscle biopsy: Absent dystrophin
        • X chromosome with normal DMD gene
          • Total inactivation
          • Usually associated with X-autosome translocation
      • Female Becker phenotype
        • Onset age: 5 to 54 years; Median 38 years
        • Weakness: Legs > Arms; Asymmetry (35%)
        • Gasctocnemius hypertrophy
        • Myalgia (25%)
        • Serum CK: 300 to 20,000
    • Cramps & Myalgia: 70% have weakness
    • Cardiomyopathy: Adult onset ²⁰
      • Onset age: 10 to 54 years; Median 40 years
      • Dilated: Duchenne 8%-10%; Becker 0%-1%
      • Left ventricle dilation (Echocardiography): Duchenne 19%; Becker 16%
      • May occur without manifesting skeletal myopathy
      • Rare in non-manifesting children: Normal EKG & Echo ⁷
      • Recommended screening
        • Echocardiography
        • Every 5 years
    • Joint contractures
      • Earlier onset patients
      • Elbows
  • Laboratory
    • Serum testing in carriers
      • CK high
        • Frequency: Duchenne 53% to 100%; Becker 30%
        • Range: 600 to 2,000
        • Asymptomatic carriers: High in 26%
      • Troponin I & T: Rarely detectable; Not related to disease-specific cardiac disorders
    • Muscle pathology
      • Myopathic changes: Varied fiber size
      • Dystrophin staining: Mosaic
        • Variable degrees of staining in different individual, or groups of, muscle fibers
        • More likely to be abnormal: Younger patients
    • Muscle MRI ²²
      • General
        • Proximal more involved than Distal
        • Asymmetric
      • Muscle patterns
        • Frequent involvement: Quadratus femoris, Gluteus maximus, Biceps femoris long head, Adductor magnus
        • Intermediate involvement: Gluteus medius, Vasti, Paraspinal, Semimembranosus, Gastrocnemius medialis
        • Spared, relative: Popliteus, Iliopsoas, Rectus abdominis, Sartorius, Gracilis, Tibialis posterior
      • Mosaicism
        • Frequency: 80% of patients; 12% of muscles
        • Patchy, asymmetric involvement of muscles
        • May be predominantly one side
  
  
• Mental retardation (Several types) ²⁵
  • General: More neuropsychiatric disturbance with mutations at 3' end of dystrophin gene
  • Reduced verbal IQ
    • Type: Selective defects
      • Shorter digit span memory
      • Developmental delay
    • Present in most DMD patients
    • No clear correlations with location of deletions
  • Autism spectrum disorder
  • Reduced total IQ < 80: Mainly with 5' end of deletion between exons 35 to 79
    • Exon 52 deletions associated with retardation
      • Disrupts or deletes Dp140 coding sequence
      • Typical DMD cognitive change
    • S-dystrophin (116 kDa) promoter (intron 55) & exon 56
      • Severe mental retardation
      • Motor: Variable; DMD & BMD phenotype
      • Nerve conduction testing: Normal
    • C-terminal mutations
      • Disrupt Dp71 (Brain dystrophin) coding sequence: Altered Dp71 transcripts
      • In dystroglycan binding domain
      • Severe retardation
      • Absent ERG b-wave
    • Copy number variation: Some mutations of uncertain significance
      • Developmental delay or Intellectual disability
      • May occur with normal strength & muscle biopsy
      • Serum CK: Normal or mildly high
      • May also be present in asymptomatic adults
  • Contiguous gene syndromes: MR often severe
  
  
• Chromosome Xp21 contiguous gene syndromes may include
  • Microdeletion: Possible associated disorders
    • Duchenne Muscular Dystrophy
    • Aland Island eye disease
    • Adrenal hypoplasia
    • Glycerol kinase deficiency
    • Retinitis Pigmentosa (RP3)
    • Mental retardation (MRX1) : IL1 receptor accessory-like protein (IL1RAPL)
    • Ornithine transcarbamylase deficiency
    • Chronic Granulomatous Disease, X-linked (CDGX)
    • McLeod red cell phenotype
  • Inversion: Pericentric Xq22
    • Disruption of RLGP (RAB40AL) gene : Ras-like GTPase in mitochondria
    • Clinical
      • Mental retardation Profound
      • Athetosis
      • Nystagmus
      • Congenital hypotonia: Severe
      • Duchenne MD

Dystrophin gene ¹

• Structure
  • 79 Exons: Only 0.6% of gene
  • 8 Promoters
  • Introns: Large; 99.4% of gene
  • Genomic DNA: 2.2 million base pairs
• Dystrophin mRNA
  • Size: 14kb
  • Encodes 3685 amino acid 427kDa protein
• 7 different dystrophin transcripts with
  • Different promoter regions & initial exons
    • N-terminus regions excluded in smaller dystrophins
    • C-terminus region is part of all sizes of dystrophin
  • Cell type specificities: Determined by promoters
    • Muscle: 427 kDa molecular weight
      • Expressed in Skeletal, Cardiac & Smooth Muscle + Retina
    • Cortical: 427 kDa; B isoform
      • Cortical post-synaptic densities, Retina, ? Skeletal Muscle
      • Different first exon from muscle isoform
      • Functionally homologous to muscle isoform
    • Purkinje Cell: 427 kDa; Cerebellar; CP isoform
      • Different first exon from muscle isoform
      • Functionally homologous to muscle isoform
      • ? Present in adult muscle
    • Retinal: 260 kDa
      • Retinal exon 1 spliced to exon 30
      • Mouse retina: outer plexiform layer
    • Brain (Fetal) & Kidney: 140 kDa
      • Promoter & first exon: In large intron between exon 44 and 45
    • Schwann cell (S-dystrophin): 116 kDa
      • Onset exon 56
      • Schwann Cells: Submembrane of external (abaxonal) layer; Nodes of Ranvier
      • Mouse model: Reduced dystrophin in peripheral nerve causes demyelinating neuropathy
    • Glial: 71 kDa
      • Onset exon 63
      • Brain (Glia), Viscera (Lung, Liver, Kidney), Cardiac Muscle
  • Alternative splicing
    • Exon 78: Muscle & brain developmental switch
    • Exons 71 to 74: Brain, heart & Purkinje
    • Exon 68: Smooth muscle
  • External link: Dystrophin isoforms

Dystrophin protein ¹

• Dystrophin Localization: Subsarcolemmal region in cardiac and skeletal muscle
  • Organized periodically in costameres
  • Enriched at myotendinous and neuromuscular junctions
  • Cardiac muscle: Associated with T-tubules also
  • Smooth muscle
    • Discontinuous along membranes
    • Alternates with vinculin
• Dystrophin functions
  • Mechanical
    • ? Stabilizaton of membrane during contraction & relaxation
    • Part of link between intracellular cytoskeleton & extracellular matrix
  • Functional
    • Plays role in ability of muscle fibers to differentiate into fast glycolytic type
    • May play a role in organization of postsynaptic membrane & AChRs
• Dystrophin: Structural Domains ²
  

  From: HPRD Dystrophin

  • NH₂-terminus
    • 1st 240 amino acids
    • α-actinin homology
    • Cytoskeleton binding: via F-actin
      • 3 distinct regions
      • Links dystrophin to F-actin subsarcolemmal cytoskeleton
      • "Calponin homology domain" (CH)
      • External link: SMART
  • Rod: Coiled-coiled
    • 2400 amino acids
      
    • 3 helix segments: 109 amino acids
      • Contain 2 proline-rich turns
      • Repeated 24 times
    • Helix segments disrupted by non-helical regions
      • 4 Proline-rich regions
      • ? Act as hinges
    • Spectrin (SPEC) homology
      • Repeated domain 21 times
      • External link: Pfam
    • Function
      • Confers flexibility & elasticity to dystrophin
      • Actin binding
    • Deletion of rod region (Exons 17 to 48): Human disease & Mouse knockouts
      • Normal localization of dystrophin-associated proteins to membrane
      • Mild or no myopathy
  • WW domain (AA 3056-3088)
    • Overlaps rod & cysteine rich domains
    • Contains 2 conserved tryptophan domains
    • Binds to proteins with
      • Particular proline-motifs, [AP]-P-P-[AP]-Y
      • Four conserved aromatic positions: Usually Trp
    • External link: SMART
  • Cysteine-rich
    • Amino acids 3080 to 3360
    • Some homology to α-actinin carboxy-terminus
    • Contains 2 incomplete EF-hand calcium binding motifs
    • Contains ZnF_ZZ domain (AA 3307 to 3352)
      • Putative zinc-binding
      • Appears to bind calmodulin
      • Missense mutation of conserved cysteine: Duchenne muscular dystrophy phenotype
      • External link: SMART
    • Functions: Required for membrane attachment to cytoskeleton via binding
      • Directly to: β-Dystroglycan
        • via WW domain (AA 3055–3092)
        • Necessary but not sufficient
      • Indirectly to: Dystrophin-associated glycoproteins
  • Carboxy-terminal domain
    • Last 420 amino acids
    • Homology to: Utrophin; Dystrobrevin
    • Contains many potential phosphorylation sites
    • Binds to
      • Dystrobrevins: Via coiled-coil motifs
      • Syntrophins (Exon 74)
      • ± DAGs
      • β-Dystroglycan: ZZ domain (AA 3307–3354), Cys 3340 ¹³
• Dystrophin forms
  • Full-length dystrophin (427 kDa)
    • Expressed in skeletal muscle, cardiac muscle & brain
    • Each form is initiated from a different promoter
  • Short dystrophins: Share same C-terminal domain
• Dystrophin phosphorylation
  • Phosphorylated by
    • Kinase types: Proline-directed & Serine/threonine
    • Examples
      • Calmodulin-dependent protein kinase II
      • p44^erk1 mitogen-activated protein kinase (MAPK3)
      • p34^cdc2 kinase
      • Casein kinase
  • Dephosphorylated by: Calcineurin
  • Phosphorylation alters dystrophin binding to: Actin; Syntrophins
• Dystrophin protein family: Other proteins with structural similarities
  • Utrophin (UTRN)
    • Structure: Contains same 4 major domains as dystrophin
    • Widely expressed
      • Muscle: Localized to neuromuscular junctions
      • Myelin
    • Binds to: F-actin; β-Dystroglycan
    • Disorders: ? 6q duplication syndromes
  • Dystrobrevins: α ; β
    • Homology to C-terminal region of dystrophin
    • α: Highly expressed in brain, skeletal & cardiac muscles; Concentrated at NMJs
    • β: Highly expressed in brain, kidney & pancreas
    • Subellular localization: Cytoplasmic
  • Dystrophin-related protein 2 (DRP2)
    • High expression in brain & spinal cord
    • Also in kidney, ovary, epididymus
    • Homology to C-terminal region of dystrophin
  • Other proteins with sequence homologies with dystrophin: Spectrins & α-actinins

Dystrophin: Genotypes & Phenotype Correlations

• Dystrophin gene mutations: Types & Size
  • Point mutations, dystrophin ¹⁰
    • Locations
      • Along entire gene
      • > 40% 3' of exon 55
      • Many in splice acceptor/donor sites or regulatory domains
      • Promoter region: Associated with dilated cardiomyopathy
    • Type
      • Stop
        • Action: Often cause premature truncation of translation
        • Types
          • Nonsense
          • Frameshift: Small (1 base pair) deletion or insertion
        • Phenotype
          • Most commonly occurs with Duchenne phenotype
            • Loss of ambulation: 10 to 11 years
            • Stop codons in in-frame exons, especially rod domain: May be ambulatory to ~16 years
          • May also occur with Becker phenotype ¹⁶
            • Mutations in certain exons
              • Flanked by exons allowing exon-skipping
              • Rod domain location
            • Locations of stop codons with common Becker phenotype: 1, 27, 29, 37, 38, 51
            • Locations of stop codons with occasional Becker phenotype: 25, 40,43,56,75
        • Also see: Out-of-frame mutations
      • Missense mutations, dystrophin
        • Frequency: Some
        • May occur with Duchenne (Cys3313Phe) or Becker (Asp165Val) phenotype
        • L54R: Mutation in actin binding domain
        • ZZ-cysteine-rich: β-dystroglycan-binding region
        • Critical domain for dystrophin function
        • Missense mutations often result in Duchenne (severe) phenotype
          • Mutations: Cys3313Phe; D3335H; C3340Y
          • D3335H & Cys3313Phe do not prevent β-dystroglycan-binding
          • Glu3367Del, adjacent to ZZ region, also produces severe MD phenotype
      • Splice mutations, dystrophin
        • Frequency: Up to 34% of dystrophin point mutations
        • Disease phenotype: Duchenne (IVS11-9G to A) or Becker (IVS25+1G to C)
        • Becker dystrophy: Often affect cononical splice site sequences
          • Cryptic splice site activation
            • May produce insertion of more amino acids or pseudoexon ¹¹
              • IVS25+2036A>G in intron 25; IVS62–285A>G in intron 62
            • Pseudo-exons ¹⁵
              • Intronic mutations
                • Cause novel splice sites & splicing changes
                • Effect: Intronic sequence included in mRNA
                • Produce: Out-of-frame or stop effects
              • Detected by: Analysis of mRNA from muscle
              • Associated pohenotype: Duchenne or Becker
              • Therapy: Pseudoexon skipping may increase amount of wild-type dystrophin
          • In frame skipping of exon
          • Inefficient function of mutant splice site
            • Single base substitution: Disrupts the invariant GT dinucleotide at 5' end of intron 54
            • Produces small amount of normal dystrophin
      • Common in CpG sequences: All C-to-T transitions
      • Many private mutations: Unique to family
    • Frequency: Detected in ~73% of patients without deletions or duplications
    • External link: Leiden database
  • Deletion mutations, Dystrophin ¹⁴
    • Location
      • Majority of deletions found at the 3' end region
      • 5' end deletions in 18%
      • Most common in large introns in NH₂-terminus & Rod domains
      • Hotspots
        • Most prominent: Around exons 45-52
          • Starting breakpoint (5' end): Intron 44 involved most frequently (15%)
            • Intron 44 features
              • Largest intron of the DMD gene
              • Contains a hotspot for meiotic recombination
            • Intron 44 deletion breakpoints: Distributed relatively evenly within intron 44
          • 3' end: Introns 50-55
        • Other: Exons 3-19
        • Population specific patterns of breakpoints in the dystrophin gene
    • Size: More than one exon usually deleted
    • Clinical-Genetic correlations
      • Out of frame deletions
        • General
          • Severe (Duchenne): Most common
            
          • Moderately severe (Outliers): Exon 3 to 7, or Exon 44 or 45 deletions
          • Mild with cardiomyopathy: Muscle promoter & exon 1 deletion
            
          • Rate of strength decline: May correlate with location of deletion
        • Specific DMD associations: "Skippable" deletion locations (On steroid treatment) ³⁵
          • Slower decline in respiratory function (FVC%)
            • Skip44: Maximum at 8.5 years; 2.7% per year; Also slower motor decline
            • Skip53: Maximum at 8.5 years; Decline starts late at 14 years; More motor decline
          • Faster decline in respiratory function (FVC%)
            • Skip51: Decline onset 5 years; 5.9% decline per year;
            • Skip45: Decline onset 5 years; 3.7% decline per year
      • In frame deletions
        • General: Milder phenotype
        • More severe disease than expected
          • Deletions of ≥ 34 exons
          • Exon 3 (In-frame): May present with Duchenne phenotype
          • Largest in-frame deletion: Exons 10 to 60 (Contains entire rod domain)
            • Severe, early-onset disease
            • Utrophin protein: Absent from sarcolemma
        • N-terminal (5') (N-terminal 240 AAs): Actin-binding region
          • Critical domain
          • Mutations often result in Duchenne or severe Becker phenotype
          • Exon 3 to 9 deletion: Mild phenotype or Asymptomatic
        • Proximal third of rod domain: Cramps & Myalgias
          • Exons 13 to 17
          • Dystrophin staining of muscle is often normal
        • Central rod domain: In frame deletions
          • General
            • Mild or Moderate phenotype
          • Exon 45 to 55 ³⁷
            • Mild phenotype: Some patients asymptomatic in 6th decade
            • CNS: 90% with cognitive impairment
            • Cardiopathy (19%)
          • Exon 48: Mild involvement
          • Deletions bordering exon 51 (Hinge domain): Mild involvement ²⁶
        • WW-cysteine-rich (Exons 63 to 69): β-dystroglycan-binding
        • Critical domain
        • Dystrophin may not bind to β-dystroglycan Loss of β-dystroglycan and sarcoglycan complex from sarcolemma
        • Mutations often result in Duchenne phenotype
        • C-terminal
          • Frequency: Rare
          • Phenotype: May be varied, mild or severe even in same family
        • Double deletions in one gene ¹²
          • Mutation locations: Hot spot exons 3 to 19 & 45 to 52
          • Clinical phenotype: Milder
          • Muscle
            • Often some dystrophin expressed
            • Partial expression at rim of fibers or Revertants
      • N-Terminus region
        • Exon 1 & Promoter region: Mild weakness ± Cardiomyopathy, severe
        • Exon 3 to 7: Exceptions to frameshift rule
          • Out of frame deletion
          • May produce Duchenne or Becker phenotype
          • Dystrophin may be produced with start at exon 8
          • Loss of ambulation: Mean 4 to 5 years later than other out of frame deletions
          • Effects partially related to endogenous exon skipping
        • Exon 5: In-frame; More severe phenotype than expected Becker
        • Exon 19: May be prone to small mutations
      • Central rod domain: Out of frame mutations in this region in 60% of DMD
        • Exon 45: Deletions can present as either Duchenne or Becker phenotype
        • Exon 45-46: Duchenne phenotype
        • Exon 45-47 or 45-48: Becker, moderate phenotype (97%)
        • Exon 52: Early loss of ambulation, Mean 10 years
      • Also see: Frame shifting
    • Disease frequency of deletions or duplications of ≥ 1 exon
      • Duchenne
        • Deletions: 60%
        • Duplications: 5%
      • Becker
        • 55% to 85% of patients have gene deletions or duplications
        • Many Becker patients have in-frame mutations in the dystrophin rod domain
          • Effect: Reduce or increase dystrophin length
          • Location: Especially frequent between exons 45 and 53
          • Mutant dystrophins retain actin- & β-dystroglycan-binding sites
    • External link: Leiden database
  • Very large chromosomal deletions
    • "Xp21 chromosomal microdeletion syndromes"
    • Multisystem disorders
    • Duchenne muscular dystrophy phenotype
      
  • Duplications ¹⁴
    • Locations
      • Mostly near 5' end of dystropin gene: Exons 6 and 7 duplicated most frequently (22% of cases)
      • Exon 2 alone
        • Common single exon duplication
        • Duplications including the promoter region don't disturb the reading frame
      • Duplication frequency gradually decreases to 1% towards 3' end of dystrophin gene (exons 66–79)
      • Small local increases at exons 18, 22, 38 & 51
      • Few duplications in exon 45–55 deletion hotspot region
    • Length: Mean: 7 exons
    • Duplication effects may be complex: Reading frame rule not always applicable
    • Duchenne dystrophy: Frequency of duplications 5%
  • Mutation Database: HGMD; Utah
• Effects of mutation on gene reading frame
  • Reading frame shifted ("Out-of-frame")
    • Results in stop codons
    • Produces small, unstable protein, ? with impaired membrane attachment
    • Severely reduced or absent levels of dystrophin
      • Some dystrophin often synthesized with:
        • Single exon deletions (e.g. 44 or 45)
        • Deletions starting or finishing with exons 3 or 51
      • Dystrophin often absent with:
        
        • Deletions starting or finishing with exons 46 or 52
      • Weakness: Genetic correlations
      • Mental retardation (Several types)
  • Dystrophin reading frame not shifted ("In-frame")
    • No stop codons
    • Produces internal deletions or duplicatons in protein
    • Levels of dystrophin:
      • High (40% to 70%): Distal rod deletions (exons 45 - 48)
      • Low (10% of normal): N-terminus deletions
    • Clinical Phenotype: Correlates with amount & function of dystrophin
• Effects of dystrophin mutations: Other
  • Dystrophin absence
    • DMD phenotype: Severe
    • Reduction in sarcoglycans & other proteins in dystrophin-glycoprotein complex
    • Dysferlin increased in cytoplasm
  • Functional consequences of loss of dystrophin on muscle fibers
    • Increased movement of membrane impermeant molecules into and out of muscle cells
    • Force production: Decreased; Hypersensitive to lengthening, or eccentric contraction
    • Force decrement with eccentric contraction correlates with acutely increased sarcolemmal permeability
    • Disorganized costameres
• Modifiers: Other genes
  • Secreted phosphoprotein 1 (SPP1; Osteopontin) ¹⁷
    • Polymorphism (rs28357094): Dominant G allele in promoter region
    • Polymorphism effect: Lower SPP1 mRNA production
    • TGFβ pathway
    • Clinical effects of G allele
      • Reduced grip strength: Especially in older steroid treated patients
      • Fewer patients ambulatory after 13 years
  • Latent transforming growth factor-beta-binding protein 4 (LTBP4) ²⁴
    • Polymorphism: IAAM haplotype
    • Clinical effect of IAAM homozygotes: Ambulatory to older age
    • IAAM effect mechanism: May relate to changes in TGFβ-related phospho-SMAD signaling
  • α-actinin-3 (ACTN3) ²⁹
    • Mutation: Null polymorphism (R577X); Heterozygous
    • Effects
      • Reduced: Strength & Walking speed
      • Less: Stretch-induced eccentric damage in mouse model
  • CD40 (TNFRSF5) ³⁰
    • Pathways: NF-κB and TGFβ
    • Mutation: Minor allele at rs1883832, in 5'-untranslated region of CD40
    • Effect: Earlier loss of ambulation
  • Thrombospondin 1 (THBS1)
    • SNPs in regulatory region of THBS1
    • THBS1: Activator of TGFβ
  • TCTEX1D1 (DYNLT5)
    • Polymorphisms associated with earlier age of loss of ambulation

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  Duchenne Headstone

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  Becker MD: Rising from the Floor - Supine to Standing

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  Return to Myopathy & NMJ Index
  Return to Neuromuscular Syndromes
  
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  10/3/2024