Expression of recombinant dystrophin and its localization to the cell membrane.
Duchenne's muscular dystrophy (DMD) is an X-linked progressive myopathy caused by a defect in the DMD gene locus. The gene corresponding to the DMD locus produces a 14-kilobase (kb) messenger RNA that codes for a large cytoskeletal membrane protein, dystrophin. DMD and Becker's muscular dystrophy are the consequences of dystrophin mutations. The exact biological function of dystrophin remains unknown but it has been demonstrated that it is localized to the cytoplasmic face of the cell membrane and has direct interaction with several other membrane proteins. We report here the synthesis of a 14-kb full-length complementary DNA for the mouse muscle dystrophin mRNA and the expression of this cDNA in COS cells. The recombinant dystrophin is indistinguishable from mouse muscle dystrophin by western blot analysis with anti-dystrophin antibodies and was shown by an immunofluorescent technique to be localized in the cell membrane. Our successful construction of a functional full-length cDNA opens opportunities for the study of structure and function of dystrophin and provides an opportunity to initiate gene therapy studies.
Lee
C C
CC
Institute for Molecular Genetics, Baylor College of Medicine, Houston, Texas 77030.
Pearlman
J A
JA
Chamberlain
J S
JS
Caskey
C T
CT
D000818:Animals; D010582:Bacteriophage lambda / Q000235:genetics; D001483:Base Sequence; D015153:Blotting, Western; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D016189:Dystrophin / Q000235:genetics*; D005455:Fluorescent Antibody Technique; D015870:Gene Expression; D005822:Genetic Vectors; D051379:Mice; D008969:Molecular Sequence Data; D009132:Muscles / Q000737:chemistry; D012333:RNA, Messenger / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics; D015183:Restriction Mapping; D014162:Transfection
D016189:Dystrophin; D012333:RNA, Messenger; D011994:Recombinant Proteins; D004247:DNA