bromochloroacetic-acid and Muscular-Dystrophies

Spectacular success has recently been made towards elucidation of the molecular basis of various forms of epidermolysis bullosa (EB), a group of heritable blistering skin diseases. The information derived from these studies has already had a profound impact in terms of precise diagnosis and classification, early prenatal prediction of the phenotype and genetic counseling in families at risk for recurrence. This review highlights recent progress made in defining the molecular basis of junctional and dystrophic forms of EB and the genotype/phenotype relationships established from these studies. Extensive molecular studies, such as the ones captured in this review, form a foundation for the rational design of gene therapies to counteract these conditions in the future.

Topics: Amino Acid Sequence; Animals; Antigens, CD; Autoantigens; Basement Membrane; Carrier Proteins; Codon; Collagen; Collagen Type XVII; Cytoskeletal Proteins; Desmosomes; DNA Mutational Analysis; Dystonin; Epidermolysis Bullosa; Genetic Heterogeneity; Genetic Therapy; Genotype; Humans; Integrin beta4; Keratins; Laminin; Mice; Mice, Knockout; Molecular Sequence Data; Muscular Dystrophies; Mutation; Nerve Tissue Proteins; Non-Fibrillar Collagens; Sequence Deletion; Skin

Plectin, a cytolinker of the plakin family, anchors the intermediate filament (IF) network formed by keratins 5 and 14 (K5/K14) to hemidesmosomes, junctional adhesion complexes in basal keratinocytes. Genetic alterations of these proteins cause epidermolysis bullosa simplex (EBS) characterized by disturbed cytoarchitecture and cell fragility. The mechanisms through which mutations located after the documented plectin IF-binding site, composed of the plakin-repeat domain (PRD) B5 and the linker, as well as mutations in K5 or K14, lead to EBS remain unclear. We investigated the interaction of plectin C terminus, encompassing four domains, the PRD B5, the linker, the PRD C, and the C extremity, with K5/K14 using different approaches, including a rapid and sensitive fluorescent protein-binding assay, based on enhanced green fluorescent protein-tagged proteins (FluoBACE). Our results demonstrate that all four plectin C-terminal domains contribute to its association with K5/K14 and act synergistically to ensure efficient IF binding. The plectin C terminus predominantly interacted with the K5/K14 coil 1 domain and bound more extensively to K5/K14 filaments compared with monomeric keratins or IF assembly intermediates. These findings indicate a multimodular association of plectin with K5/K14 filaments and give insights into the molecular basis of EBS associated with pathogenic mutations in plectin, K5, or K14 genes.

Topics: Binding Sites; Cell Line, Tumor; Epidermolysis Bullosa Simplex; HEK293 Cells; Humans; Keratin-14; Keratin-5; Keratins; Muscular Dystrophies; Mutation; Plectin; Protein Binding; Protein Multimerization; Protein Structure, Quaternary; Protein Structure, Tertiary; Recombinant Proteins; Two-Hybrid System Techniques

Plectin is a widely expressed cytomatrix component involved in the attachment of the cytoskeleton to the plasma membrane. We have recently reported that the skin and muscles of three patients affected by epidermolysis bullosa simplex with muscular dystrophy (MD-EBS), a genetic disorder characterized by skin blistering associated with muscle involvement, are not reactive with antibodies specific to plectin. We demonstrated that in the skin, lack of plectin leads to failure of keratin filaments to connect to the plasma membrane via the hemidesmosomes, whereas in the muscle the deficient expression of the molecule correlates with an aberrant localization of desmin in the muscle fibers. In this study we demonstrate that in a MD-EBS kindred with two affected members, the disease results from a homozygous nonsense mutation in the plectin (PLEC1) gene leading to a premature stop codon (CGA to TGA) and decay of the aberrant plectin messenger RNA. The segregation of the mutated allele implicates the mutation in the pathology of the disorder. These results confirm the critical role of plectin in providing cell resistance to mechanical stresses both in the skin and the muscle.

Topics: Alleles; Blotting, Northern; Codon, Nonsense; Codon, Terminator; Desmin; Epidermolysis Bullosa Simplex; Humans; Intermediate Filament Proteins; Keratins; Molecular Sequence Data; Muscle Fibers, Skeletal; Muscular Dystrophies; Pedigree; Plectin; Point Mutation; Skin