Subject(s)
Horses/genetics , Animals , Base Sequence , Calcium-Transporting ATPases/genetics , Chromosome Mapping , DNA Primers/genetics , DNA-Binding Proteins/genetics , Female , Lod Score , Male , Microphthalmia-Associated Transcription Factor , Microsatellite Repeats , Molecular Sequence Data , Nerve Tissue Proteins/genetics , Polymerase Chain Reaction , Polymorphism, Genetic , Proto-Oncogene Proteins c-kit/genetics , Sarcoplasmic Reticulum Calcium-Transporting ATPases , Transcription Factors/geneticsABSTRACT
Polymyositis is regarded as an autoimmune inflammatory muscle disease. A major subgroup of patients have autoantibodies to cellular histidyl-transfer RNA synthetase (HRS). We have analyzed the role of the autoantigen HRS in the induction of murine myositis in a comparative study of inoculation of BALB/c mice with recombinant HRS protein versus naked DNA coding for HRS. Adult BALB/c mice produced antibodies to human HRS following inoculation with HRS protein and adjuvant, but myositis was not observed. Alternatively, expression plasmid DNA constructs encoding full-length and truncated human HRS were inoculated intramuscularly in gene transfer studies. DNA-inoculated mice produced relatively low anti-HRS antibody titers. However, in contrast to recombinant HRS protein-inoculated mice, HRS gene transfer induced pathology with evidence of cellular infiltration of perivascular and endomysial regions of the inoculated muscle. Multiple inoculations of a plasmid construct encoding a hybrid molecule consisting of HRS and the transferrin receptor cytoplasmic tail induced the highest levels of antibodies and persisting cellular infiltration. Unlike HRS, expression of influenza virus hemagglutinin (HA) following inoculation of an HA plasmid did not induce myositis. Transfer of naked DNA constructs expressing HRS is likely to provide valuable information on the autoimmune response to this protein and its role in the development of myositis.
Subject(s)
Histidine-tRNA Ligase/genetics , Histidine-tRNA Ligase/immunology , Immunization , Myositis/immunology , Animals , Disease Models, Animal , Drug Administration Schedule , Female , Gene Transfer Techniques , Hemagglutinin Glycoproteins, Influenza Virus/genetics , Hemagglutinin Glycoproteins, Influenza Virus/pharmacology , Histidine-tRNA Ligase/pharmacology , Humans , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Inbred Strains , Molecular Sequence Data , Muscle, Skeletal/immunology , Muscle, Skeletal/pathology , Myositis/chemically induced , Polymyositis/immunology , Recombinant Proteins/genetics , Recombinant Proteins/immunology , Recombinant Proteins/pharmacologyABSTRACT
The mdx mouse, an animal model used to study Duchenne muscular dystrophy (DMD), has a nonsense mutation in exon 23 of the dystrophin gene which should result in a truncated protein that cannot be correctly localized at the sarcolemma of the muscle fibres. Immunohistochemical staining with anti-dystrophin antibodies had shown that while most of the muscle tissue was dystrophin-negative, a small percentage of muscle fibres were clearly dystrophin-positive and had somehow by-passed the primary nonsense mutation. A nested PCR-based examination of dystrophin gene transcripts around the mdx mutation revealed several alternatively processed transcripts, of which four mRNA species skipped the mutation in exon 23, were in-frame and could be translated into a shorter, but still functional dystrophin protein. Specific tests for these transcripts demonstrated these were also present in normal adult and embryonic mouse muscle tissue.
Subject(s)
Dystrophin/metabolism , Genetic Therapy , Muscle Fibers, Skeletal/metabolism , Muscular Dystrophy, Animal/metabolism , Muscular Dystrophy, Animal/therapy , Animals , Dystrophin/genetics , Exons , Humans , Mice , Mice, Inbred C57BL , Mice, Inbred mdx/genetics , Muscular Dystrophy, Animal/genetics , Mutation , Polymerase Chain Reaction , Sensitivity and Specificity , Transcription, GeneticABSTRACT
The murine histidyl-tRNA synthetase-encoding gene (MMHRS) coding region has been cloned and sequenced. The 1527-bp transcript shows a strikingly similar structural organization to that of its human counterpart, particularly within the three class II aminoacyl-tRNA synthetase structural motifs and the two histidyl-tRNA synthetase signature regions. It is predicted, as in humans, to have a coiled-coil alpha-helical structure that is characteristic of many autoantigens. MMHRS shows some degree of polymorphism at both the DNA and amino-acid levels, although its sequence is well conserved amongst the commonly used laboratory mouse strains.
