Abstract
Spinal muscular atrophy is a neurodegenerative disorder caused by the deletion or mutation of the survival-of-motor-neuron gene,
SMN1.
An
SMN1
paralog,
SMN2,
differs by a C→T transition in exon 7 that causes substantial skipping of this exon, such that
SMN2
expresses only low levels of functional protein. A better understanding of
SMN
splicing mechanisms should facilitate the development of drugs that increase survival motor neuron (SMN) protein levels by improving
SMN2
exon 7 inclusion. In addition, exonic mutations that cause defective splicing give rise to many genetic diseases, and the
SMN1/2
system is a useful paradigm for understanding exon-identity determinants and alternative-splicing mechanisms. Skipping of
SMN2
exon 7 was previously attributed either to the loss of an SF2/ASF–dependent exonic splicing enhancer or to the creation of an hnRNP A/B–dependent exonic splicing silencer, as a result of the C→T transition. We report the extensive testing of the enhancer-loss and silencer-gain models by mutagenesis, RNA interference, overexpression, RNA splicing, and RNA-protein interaction experiments. Our results support the enhancer-loss model but also demonstrate that hnRNP A/B proteins antagonize SF2/ASF–dependent ESE activity and promote exon 7 skipping by a mechanism that is independent of the C→T transition and is, therefore, common to both
SMN1
and
SMN2.
Our findings explain the basis of defective
SMN2
splicing, illustrate the fine balance between positive and negative determinants of exon identity and alternative splicing, and underscore the importance of antagonistic splicing factors and exonic elements in a disease context.