23 research outputs found
Muscle Chloride Channel Dysfunction in Two Mouse Models of Myotonic Dystrophy
Muscle degeneration and myotonia are clinical hallmarks of myotonic dystrophy type 1 (DM1), a multisystemic disorder caused by a CTG repeat expansion in the 3′ untranslated region of the myotonic dystrophy protein kinase (DMPK) gene. Transgenic mice engineered to express mRNA with expanded (CUG)250 repeats (HSALR mice) exhibit prominent myotonia and altered splicing of muscle chloride channel gene (Clcn1) transcripts. We used whole-cell patch clamp recordings and nonstationary noise analysis to compare and biophysically characterize the magnitude, kinetics, voltage dependence, and single channel properties of the skeletal muscle chloride channel (ClC-1) in individual flexor digitorum brevis (FDB) muscle fibers isolated from 1–3-wk-old wild-type and HSALR mice. The results indicate that peak ClC-1 current density at −140 mV is reduced >70% (−48.5 ± 3.6 and −14.0 ± 1.6 pA/pF, respectively) and the kinetics of channel deactivation increased in FDB fibers obtained from 18–20- d-old HSALR mice. Nonstationary noise analysis revealed that the reduction in ClC-1 current density in HSALR FDB fibers results from a large reduction in ClC-1 channel density (170 ± 21 and 58 ± 11 channels/pF in control and HSALR fibers, respectively) and a modest decrease in maximal channel open probability(0.91 ± 0.01 and 0.75 ± 0.03, respectively). Qualitatively similar results were observed for ClC-1 channel activity in knockout mice for muscleblind-like 1 (Mbnl1ΔE3/ΔE3), a second murine model of DM1 that exhibits prominent myotonia and altered Clcn1 splicing (Kanadia et al., 2003). These results support a molecular mechanism for myotonia in DM1 in which a reduction in both the number of functional sarcolemmal ClC-1 and maximal channel open probability, as well as an acceleration in the kinetics of channel deactivation, results from CUG repeat–containing mRNA molecules sequestering Mbnl1 proteins required for proper CLCN1 pre-mRNA splicing and chloride channel function
Lysosomal abnormalities in hereditary spastic paraplegia types SPG15 and SPG11
Objective
Hereditary spastic paraplegias (HSPs) are among the most genetically diverse inherited neurological disorders, with over 70 disease loci identified (SPG1-71) to date. SPG15 and SPG11 are clinically similar, autosomal recessive disorders characterized by progressive spastic paraplegia along with thin corpus callosum, white matter abnormalities, cognitive impairment, and ophthalmologic abnormalities. Furthermore, both have been linked to early-onset parkinsonism. Methods
We describe two new cases of SPG15 and investigate cellular changes in SPG15 and SPG11 patient-derived fibroblasts, seeking to identify shared pathogenic themes. Cells were evaluated for any abnormalities in cell division, DNA repair, endoplasmic reticulum, endosomes, and lysosomes. Results
Fibroblasts prepared from patients with SPG15 have selective enlargement of LAMP1-positive structures, and they consistently exhibited abnormal lysosomal storage by electron microscopy. A similar enlargement of LAMP1-positive structures was also observed in cells from multiple SPG11 patients, though prominent abnormal lysosomal storage was not evident. The stabilities of the SPG15 protein spastizin/ZFYVE26 and the SPG11 protein spatacsin were interdependent. Interpretation
Emerging studies implicating these two proteins in interactions with the late endosomal/lysosomal adaptor protein complex AP-5 are consistent with shared abnormalities in lysosomes, supporting a converging mechanism for these two disorders. Recent work withZfyve26−/− mice revealed a similar phenotype to human SPG15, and cells in these mice had endolysosomal abnormalities. SPG15 and SPG11 are particularly notable among HSPs because they can also present with juvenile parkinsonism, and this lysosomal trafficking or storage defect may be relevant for other forms of parkinsonism associated with lysosomal dysfunction
Early onset and novel features in a spinal and bulbar muscular atrophy patient with a 68 CAG repeat
AbstractSpinal and bulbar muscular atrophy (SBMA) is an X-linked neuromuscular disease caused by a trinucleotide (CAG) repeat expansion in the androgen receptor gene. Patients with SBMA have weakness, atrophy, and fasciculations in the bulbar and extremity muscles. Individuals with CAG repeat lengths greater than 62 have not previously been reported. We evaluated a 29year old SBMA patient with 68 CAGs who had unusually early onset and findings not seen in others with the disease. Analysis of the androgen receptor gene confirmed the repeat length of 68 CAGs in both peripheral blood and fibroblasts. Evaluation of muscle and sensory function showed deficits typical of SBMA, and in addition the patient had manifestations of autonomic dysfunction and abnormal sexual development. These findings extend the known phenotype associated with SBMA and shed new insight into the effects of the mutated androgen receptor
Consensus-based care recommendations for adults with myotonic dystrophy type 1
Purpose of review
Myotonic dystrophy type 1 (DM1) is a severe, progressive genetic disease that affects between 1 in 3,000 and 8,000 individuals globally. No evidence-based guideline exists to inform the care of these patients, and most do not have access to multidisciplinary care centers staffed by experienced professionals, creating a clinical care deficit.
Recent findings
The Myotonic Dystrophy Foundation (MDF) recruited 66 international clinicians experienced in DM1 patient care to develop consensus-based care recommendations. MDF created a 2-step methodology for the project using elements of the Single Text Procedure and the Nominal Group Technique. The process generated a 4-page Quick Reference Guide and a comprehensive, 55-page document that provides clinical
care recommendations for 19 discrete body systems and/or care considerations.
Summary
The resulting recommendations are intended to help standardize and elevate care for this patient population and reduce variability in clinical trial and study environments. Described as “one of the more variable diseases found in medicine,” myotonic dystrophy type
1 (DM1) is an autosomal dominant, triplet-repeat expansion disorder that affects somewhere between 1:3,000 and 1:8,000 individuals worldwide.1 There is a modest association between increased repeat expansion and disease severity, as evidenced by the average age of onset and overall morbidity of the condition. An expansion of over 35 repeats typically indicates an unstable and expanding mutation. An expansion of 50 repeats or higher is consistent with a diagnosis of DM1. DM1 is a multisystem and heterogeneous disease characterized by distal weakness, atrophy, and myotonia, as well as symptoms in the heart, brain, gastrointestinal tract, endocrine, and respiratory systems. Symptoms may occur at any age. The severity of the condition varies widely among affected individuals, even among members of the same family.
Comprehensive evidence-based guidelines do not currently
exist to guide the treatment of DM1 patients. As a result, the international patient community reports varied levels of care and care quality, and difficulty accessing care adequate to manage their symptoms, unless they have access to multidisciplinary neuromuscular clinics.
Consensus-based care recommendations can help standardize
and improve the quality of care received by DM1 patients
and assist clinicians who may not be familiar with the significant variability, range of symptoms, and severity of the disease. Care recommendations can also improve the landscape for clinical trial success by eliminating some of the inconsistencies in patient care to allow more accurate understanding of the benefit of potential therapies
Myotonic disorders
Myotonia reflects a state of muscle fiber hyperexcitability. Impaired
transmembrane conductance of either chloride or sodium ions results in
myotonia. Myotonic disorders include the myotonic dystrophies and
nondystrophic myotonias. Mutations in the genes encoding chloride
(ClC-1) or sodium (SCN4A) channels expressed exclusively in skeletal
muscle cause nondystrophic myotonias. Genetic defects in the myotonic
dystrophies do not involve ion channel or its regulator proteins.
Recent research supports a novel RNA-mediated disease mechanism of
myotonia in the myotonic dystrophies. Myotonic dystrophy Type 1 is
caused by CTG repeat expansion in the 3′ untranslated region in
the Dystrophia Myotonica Protein Kinase (DMPK) gene. Myotonic dystrophy
Type 2 is caused by CCTG repeat expansion in the first intron in Zinc
Finger Protein 9 (ZNF9) gene. The expanded repeat is transcribed in RNA
and forms discrete inclusions in nucleus in both types of myotonic
dystrophies. Mutant RNA sequesters MBNL1, a splice regulator protein
and depletes MBNL1 from the nucleoplasm. Loss of MBNL1 results in
altered splicing of ClC-1 mRNA. Altered splice products do not encode
functional ClC-1 protein. Subsequent loss of chloride conductance in
muscle membrane causes myotonia in the myotonic dystrophies. The
purpose of this review is to discuss the clinical presentation, recent
advances in understanding the disease mechanism with particular
emphasis on myotonic dystrophies and potential therapy options in
myotonic disorders
Myotonic dystrophy type 1 is associated with nuclear foci of mutant RNA, sequestration of muscleblind proteins and deregulated alternative splicing in neurons
\u27Double trouble\u27: Diagnostic challenges in Duchenne muscular dystrophy in patients with an additional hereditary skeletal dysplasia
Expression and Purification of ZASP Subdomains and Clinically Important Isoforms: High-Affinity Binding to G-Actin
Z-disc-associated, alternatively spliced, PDZ motif-containing protein (ZASP) is a principal component of the sarcomere. The three prevalent isoforms of ZASP in skeletal muscle are generated by alternative splicing of exons 9 and 10. The long isoforms, either having (ZASP-L) or lacking exon 10 (ZASP-L\u394ex10), include an N-terminal PDZ domain, an actin-binding region (ABR) with a conserved motif (ZM), and three C-terminal LIM domains. The short isoform (ZASP-S) lacks the LIM domains. Mutations, A147T and A165V, within the ZM of ZASP-L\u394ex10 cause myofibrillar myopathy, but the mechanism is unknown. We have prepared these proteins, their ABR, and the respective mutant variants in recombinant form, characterized them biophysically, and analyzed their actin-binding properties by surface plasmon resonance and electron microscopy. All the proteins were physically homogeneous and monomeric and had circular dichroic spectra consistent with partially folded conformations. Comparison of the NMR HSQC spectra of ZASP-S and the PDZ domain showed that the ABR is unstructured. ZASP-S and its mutant variants and ZASP-L\u394ex10 all bound to immobilized G-actin with high affinity (Kd 48 10(-8) to 10(-9) M). Constructs of the isolated actin-binding region missing exon 10 (ABR\u39410) bound with lower affinity (Kd 48 10(-7) M), but those retaining exon 10 (ABR+10) did so only weakly (Kd 48 10(-5) M). ZASP-S, and the ABR\u39410, also induced F-actin and array formation, even in conditions of low ionic strength and in the absence of KCl and Mg(2+) ions. Interestingly, the ZM mutations A147T and A165V did not affect any of the results described above