Repair of pre-mRNA splicing

Nlend, Rachel Nlend; Meyer, Kathrin; Schümperli, Daniel

doi:10.4161/rna.7.4.12206

Cited by 28 publications

(14 citation statements)

References 153 publications

(174 reference statements)

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“…SF2/ASF recognizes a splicing enhancer that includes C in the +6 position at the 5′ end of exon 7 in SMN1 to promote exon 7 inclusion. 25 This interaction is antagonized by hnRNPA1, which binds at an overlapping splicing silencer that is created by the C to T transition at the +6 position in exon 7 in the SMN2 transcripts. 25 Little to no increase in luciferase activity or change in RNA was observed in the SMN2-luciferase reporter cell-line (Fig.…”

Section: Resultsmentioning

confidence: 99%

Identification of Novel Compounds That Increase SMN Protein Levels Using an Improved SMN2 Reporter Cell Assay

Cherry

Evans

et al. 2012

SLAS Discovery

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Spinal muscular atrophy (SMA) is a neurodegenerative disorder that is characterized by progressive loss of motor neuron function. It is caused by the homozygous loss of the SMN1 (survival of motor neuron 1) gene and a decrease in full-length SMN protein. SMN2 is a nearly identical homolog of SMN1 that, due to alternative splicing, expresses predominantly truncated SMN protein. SMN2 represents an enticing therapeutic target. Increasing expression of full-length SMN from the SMN2 gene might represent a treatment for SMA. We describe a newly designed cell-based reporter assay that faithfully and reproducibly measures full-length SMN expression from the SMN2 gene. This reporter can detect increases of SMN protein by an array of compounds previously shown to regulate SMN2 expression and by the overexpression of proteins that modulate SMN2 splicing. It also can be used to evaluate changes at both the transcriptional and splicing level. This assay can be a valuable tool for the identification of novel compounds that increase SMN2 protein levels and the optimization of compounds already known to modulate SMN2 expression. We present here preliminary data from a high-throughput screen using this assay to identify novel compounds that increase expression of SMN2.

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Section: Resultsmentioning

confidence: 99%

Identification of Novel Compounds That Increase SMN Protein Levels Using an Improved SMN2 Reporter Cell Assay

Cherry

Evans

et al. 2012

SLAS Discovery

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“…[29][30][31] The exon definition model requires simultaneous recognition of correction in SMA. [20][21][22][23][24] Here we focus on emerging novel aspects of splicing regulation of SMN2 exon 7 that happens to be the last coding as well as the last internal exon. The 54-nt long human SMN exon 7 is flanked by a 5.76 kb intron 6 and a 444 nt intron 7 (Fig.…”

Section: O N O T D I S T R I B U T Ementioning

confidence: 99%

Alternative splicing in spinal muscular atrophy underscores the role of an intron definition model

Singh

2011

RNA Biology

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H umans have two nearly identical copies of the Survival MotorNeuron (SMN) gene: SMN1 and SMN2. The two SMN genes code for identical proteins; however, SMN2 predominantly generates a shorter transcript due to skipping of exon 7, the last coding exon. Skipping of SMN2 exon 7 leads to production of a truncated SMN protein that is highly unstable. The inability of SMN2 to compensate for the loss of SMN1 results in spinal muscular atrophy (SMA), the second most prevalent genetic cause of infant mortality. Since SMN2 is almost universally present in SMA patients, correction of SMN2 exon 7 splicing holds the promise for cure. Consistently, SMN2 exon 7 splicing has emerged as one of the best studied splicing systems in humans. The vast amount of recent literature provides a clue that SMN2 exon 7 splicing is regulated by an intron definition mechanism, which does not require cross-exon communication as prerequisite for exon inclusion. Our conclusion is based on the prominent role of intronic cis-elements, some of them have emerged as the frontrunners among potential therapeutic targets of SMA. Further, the widely expressed T-cellrestricted intracellular antigen-1 (TIA1), a member of the glutamine rich domain containing RNA-binding proteins, has recently been found to regulate SMN exon 7 splicing by binding to intron 7 sequences away from the 5' splice site (ss). These findings make a strong argument for an "intron definition model," according to which regulatory sequences within a downstream intron are capable of enforcing exon inclusion even in the

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“…Effective protein restoration in DMD via exon skipping has been shown in patient derived cell cultures, animal models, (reviewed in [9] and even in clinical trials [13,14]. Similar results have been obtained in SMA via the related mechanism of exon inclusion [10,15-17]. …”

Section: Introductionmentioning

confidence: 62%

Antisense-mediated isoform switching of steroid receptor coactivator-1 in the central nucleus of the amygdala of the mouse brain

et al. 2013

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BackgroundAntisense oligonucleotide (AON)-mediated exon skipping is a powerful tool to manipulate gene expression. In the present study we investigated the potential of exon skipping by local injection in the central nucleus of the amygdala (CeA) of the mouse brain. As proof of principle we targeted the splicing of steroid receptor coactivator-1 (SRC-1), a protein involved in nuclear receptor function. This nuclear receptor coregulator exists in two splice variants (SRC-1a and SRC-1e) which display differential distribution and opposing activities in the brain, and whose mRNAs differ in a single SRC-1e specific exon.MethodsFor proof of principle of feasibility, we used immunofluorescent stainings to study uptake by different cell types, translocation to the nucleus and potential immunostimulatory effects at different time points after a local injection in the CeA of the mouse brain of a control AON targeting human dystrophin with no targets in the murine brain. To evaluate efficacy we designed an AON targeting the SRC-1e-specific exon and with qPCR analysis we measured the expression ratio of the two splice variants.ResultsWe found that AONs were taken up by corticotropin releasing hormone expressing neurons and other cells in the CeA, and translocated into the cell nucleus. Immune responses after AON injection were comparable to those after sterile saline injection. A successful shift of the naturally occurring SRC-1a:SRC-1e expression ratio in favor of SRC-1a was observed, without changes in total SRC-1 expression.ConclusionsWe provide a proof of concept for local neuropharmacological use of exon skipping by manipulating the expression ratio of the two splice variants of SRC-1, which may be used to study nuclear receptor function in specific brain circuits. We established that exon skipping after local injection in the brain is a versatile and useful tool for the manipulation of splice variants for numerous genes that are relevant for brain function.

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Repair of pre-mRNA splicing

Cited by 28 publications

References 153 publications

Identification of Novel Compounds That Increase SMN Protein Levels Using an Improved SMN2 Reporter Cell Assay

Identification of Novel Compounds That Increase SMN Protein Levels Using an Improved SMN2 Reporter Cell Assay

Alternative splicing in spinal muscular atrophy underscores the role of an intron definition model

Antisense-mediated isoform switching of steroid receptor coactivator-1 in the central nucleus of the amygdala of the mouse brain

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