title: A cell system with targeted disruption of the SMN gene: functional conservation of the SMN protein and dependence of Gemin2 on SMN.
abstract: The motor neuron degenerative disease spinal muscular atrophy is caused by reduced expression of the survival motor neuron (SMN) protein. Here we report a genetic system developed in the chicken pre-B cell line DT40, in which the endogenous SMN gene is disrupted by homologous recombination, and SMN protein is expressed from a chicken SMN cDNA under control of a tetracycline (tet)-repressible promoter. Addition of tet results in depletion of SMN protein and consequent cell death, which directly demonstrates that SMN is required for cell viability. The tet-induced lethality can be rescued by expression of human SMN, indicating that the function of SMN is highly conserved between the two species. Cells expressing low levels of SMN display slow growth proportional to the amount of SMN they contain. Interestingly, the level of the SMN-interacting protein Gemin2 decreases significantly following depletion of SMN, supporting the conclusion that SMN and Gemin2 form a stable complex in vivo. This system provides a powerful setting for studying the function of SMN in vivo and for screening for potential therapeutics for spinal muscular atrophy.
authors: Howard Hughes Medical Institute and Department of Biochemistry and Biophysics, University of Pennsylvania, School of Medicine, Philadelphia 19104, USA.
authors: J
authors: J
authors: Wang
authors: G
authors: G
authors: Dreyfuss
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D015139:Blotting, Southern; D015153:Blotting, Western; D016923:Cell Death; D002460:Cell Line; D002470:Cell Survival; D002478:Cells, Cultured; D002645:Chickens; D017124:Conserved Sequence; D017362:Cyclic AMP Response Element-Binding Protein; D018076:DNA, Complementary / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D005091:Exons; D005434:Flow Cytometry; D015723:Gene Library; D006801:Humans; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009134:Muscular Atrophy, Spinal / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D010957:Plasmids; D011401:Promoter Regions, Genetic; D016601:RNA-Binding Proteins; D011995:Recombination, Genetic; D055532:SMN Complex Proteins; D013752:Tetracycline / Q000378:metabolism; D013997:Time Factors; D014162:Transfection
chemical_list: D017362:Cyclic AMP Response Element-Binding Protein; D018076:DNA, Complementary; C108381:GEMIN2 protein, human; D009419:Nerve Tissue Proteins; D016601:RNA-Binding Proteins; D055532:SMN Complex Proteins; D013752:Tetracycline