Stimulating full-length SMN2 expression by delivering bifunctional RNAs via a viral vector.
Spinal muscular atrophy (SMA) is an autosomal recessive neuromuscular disorder that is the leading genetic cause of infant mortality. SMA is caused by the loss of survival motor neuron-1 (SMN1). In humans, a nearly identical copy gene is present, called SMN2. SMN2 is retained in all SMA patients and encodes an identical protein compared to SMN1. However, a single silent nucleotide difference in SMN2 exon 7 results in the production of a spliced isoform (called SMNDelta7) that encodes a nonfunctional protein. The presence of SMN2 represents a unique therapeutic target since SMN2 has the capacity to encode a fully functional protein. Here we describe an in vivo delivery system for short bifunctional RNAs that modulate SMN2 splicing. Bifunctional RNAs derive their name from the presence of two domains: an antisense RNA sequence specific to a target RNA and an untethered RNA segment that serves as a binding platform for splicing factors. Plasmid-based and recombinant adeno-associated virus vectors were developed that expressed bifunctional RNAs that stimulated SMN2 exon 7 inclusion and full-length SMN protein in patient fibroblasts. These experiments provide a mechanism to modulate splicing from a variety of genetic contexts and demonstrate directly a novel therapeutic approach for SMA.
Baughan
Travis
T
Department of Molecular Microbiology and Immunology, Life Sciences Center, University of Missouri, Columbia, MO 65211, USA.
Shababi
Monir
M
Coady
Tristan H
TH
Dickson
Alexa M
AM
Tullis
Gregory E
GE
Lorson
Christian L
CL
D015153:Blotting, Western; D002478:Cells, Cultured; D017362:Cyclic AMP Response Element-Binding Protein / Q000235:genetics* / Q000378:metabolism; D000229:Dependovirus / Q000235:genetics*; D005091:Exons / Q000235:genetics; D005347:Fibroblasts / Q000378:metabolism / Q000473:pathology; D005822:Genetic Vectors / Q000235:genetics*; D006367:HeLa Cells; D006801:Humans; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008957:Models, Genetic; D009134:Muscular Atrophy, Spinal / Q000235:genetics / Q000473:pathology / Q000628:therapy; D009419:Nerve Tissue Proteins / Q000235:genetics* / Q000378:metabolism; D012313:RNA / Q000235:genetics* / Q000378:metabolism; D012322:RNA Precursors / Q000235:genetics / Q000378:metabolism; D012326:RNA Splicing / Q000235:genetics; D016601:RNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D055532:SMN Complex Proteins; D055533:Survival of Motor Neuron 1 Protein; D055540:Survival of Motor Neuron 2 Protein; D014162:Transfection / Q000379:methods
D017362:Cyclic AMP Response Element-Binding Protein; D009419:Nerve Tissue Proteins; D012322:RNA Precursors; D016601:RNA-Binding Proteins; D055532:SMN Complex Proteins; C526471:SMN1 protein, human; C526477:SMN2 protein, human; D055533:Survival of Motor Neuron 1 Protein; D055540:Survival of Motor Neuron 2 Protein; D012313:RNA; D008156:Luciferases