title: Survival of motor neuron gene downregulation by RNAi: towards a cell culture model of spinal muscular atrophy.
abstract: Gene silencing with double-stranded RNA (RNAi) has proved useful for gene function studies, and should be especially well suited to studying diseases resulting in embryonal lethality where transgenic animal models are difficult to generate. We are applying this approach to the autosomal recessive disease spinal muscular atrophy (SMA). SMA is caused by mutations in the survival of motor neuron gene (SMN). The SMN protein is ubiquitously expressed and plays a role in RNA processing and its reduction in SMA ultimately leads to motor neuron degeneration in the spinal cord. The reasons for this motor neuron selectivity, however, are still unclear. SMN is essential for the viability of most eukaryotic organisms and this has made the generation of animal models of SMA extremely difficult. Here we describe a different approach to study SMN function using RNAi to silence SMN expression in cells. We designed double-stranded small interfering RNA (siRNA) targeted against murine Smn and transfected the murine embryonal terato-carcinoma cell line P19. The siRNAs reduced both Smn RNA and protein levels in the P19 cells compared to controls. These results illustrate that double-stranded RNA can be an effective gene silencing approach even in a protein that is essential for survival and highly expressed, and it could therefore be a valuable tool to study SMN function.
authors.affiliation: Department of Human Anatomy and Genetics, Oxford University, South Parks Rd., Oxford OX1 3QX, UK. Barbara.Trulzch@anat.ox.ac.uk
authors.forename: Barbara
authors.initials: B
authors.lastname: Trülzsch
authors.forename: Kay
authors.initials: K
authors.lastname: Davies
authors.forename: Matthew
authors.initials: M
authors.lastname: Wood
mesh_terms: D000818:Animals; D015153:Blotting, Western / Q000379:methods; D002277:Carcinoma; D045744:Cell Line, Tumor; D017362:Cyclic AMP Response Element-Binding Protein; D004195:Disease Models, Animal; D015536:Down-Regulation; D004622:Embryo, Mammalian; D020868:Gene Silencing / Q000187:drug effects*; D051379:Mice; D009046:Motor Neurons / Q000187:drug effects* / Q000473:pathology; D009134:Muscular Atrophy, Spinal / Q000235:genetics* / Q000473:pathology; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000378:metabolism; D011419:Propidium / Q000378:metabolism; D012330:RNA, Double-Stranded / Q000494:pharmacology*; D012333:RNA, Messenger / Q000096:biosynthesis; D034741:RNA, Small Interfering / Q000737:chemistry / Q000494:pharmacology*; D016601:RNA-Binding Proteins; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D055532:SMN Complex Proteins; D013997:Time Factors; D014162:Transfection
chemical_list: D017362:Cyclic AMP Response Element-Binding Protein; D009419:Nerve Tissue Proteins; D012330:RNA, Double-Stranded; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; D055532:SMN Complex Proteins; D011419:Propidium