Point mutations in the coding regions of genes are commonly assumed to exert their effects by altering single amino acids in the encoded proteins. However, there is increasing evidence that many human disease genes harbour exonic mutations that affect pre-mRNA splicing. Nonsense, missense and even translationally silent mutations can inactivate genes by inducing the splicing machinery to skip the mutant exons. Similarly, coding-region single-nucleotide polymorphisms might cause phenotypic variability by influencing splicing accuracy or efficiency. As the splicing mechanisms that depend on exonic signals are elucidated, new therapeutic approaches to treating certain genetic diseases can begin to be explored.

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Listening to silence and understanding nonsense: exonic mutations that affect splicing

http://sites.utoronto.ca/intron/blencowecell2006.pdf

Alternative Splicing: New Insights from Global Analyses

Alteration of correct splicing patterns by disruption of an exonic splicing enhancer may be a frequent mechanism by which point mutations cause genetic diseases. Spinal muscular atrophy results from the lack of functional survival of motor neuron 1 gene (SMN1), even though all affected individuals carry a nearly identical, normal SMN2 gene. SMN2 is only partially active because a translationally silent, single-nucleotide difference in exon 7 causes exon skipping. Using ESE motif-prediction tools, mutational analysis and in vivo and in vitro splicing assays, we show that this single-nucleotide change occurs within a heptamer motif of an exonic splicing enhancer, which in SMN1 is recognized directly by SF2/ASF. The abrogation of the SF2/ASF-dependent ESE is the basis for inefficient inclusion of exon 7 in SMN2, resulting in the spinal muscular atrophy phenotype.

Disruption of an SF2/ASF-dependent exonic splicing enhancer in SMN2 causes spinal muscular atrophy in the absence of SMN1

https://www.cell.com/trends/genetics/pdf/S0168-9525(01)02626-9.pdf

Alternative splicing: multiple control mechanisms and involvement in human disease

When genome variants are identified in genomic DNA, especially during routine analysis of disease-associated genes, their functional implications might not be immediately evident. Distinguishing between a genomic variant that changes the phenotype and one that does not is a difficult task. An increasing amount of evidence indicates that genomic variants in both coding and non-coding sequences can have unexpected deleterious effects on the splicing of the gene transcript. So how can benign polymorphisms be distinguished from disease-associated splicing mutations?

Genomic variants in exons and introns: identifying the splicing spoilers

Neurofibromatosis type 1 (NF1) is one of the mostcommon inherited disorders in humans and is causedby mutations in the NF1gene. To date, the majority ofthe reported NF1mutations are predicted to result inprotein truncation, but veryfew studieshavecorrelatedthe causative NF1mutation with its effect at the mRNAlevel. We have applied a whole NF1cDNA screeningmethodology to the study of 80 unrelated NF1 patientsand have identified 44 different mutations, 32 beingnovel, in 52 of these patients. Mutations were detectedin 87% of the familial cases, but in 51% of the sporadicones. At least 15 of the 80 NF1 patients (19%) hadrecurrent mutations. The study shows that in 50% of thepatients in whom the mutations were identified, theseresulted in splicing alterations. Most of the splicingmutations did not involve the conserved AG/GTdinucleotides of the splice sites. One frameshift, twononsense and two missense mutations were alsoresponsible for alterations in mRNA splicing. Thelocation and type of mutation within the NF1gene, andits putative effect at the protein level, do not indicateany relationship to any specific clinical feature of NF1.The high proportion of aberrant spliced transcriptsdetected in NF1 patients stresses the importance ofstudying mutations at both the genomic and RNA level.It is possible that part of the clinical variability in NF1could be due to mutations affecting mRNA splicing,which is the most common molecular defect in NF1.INTRODUCTIONNeurofibromatosis type 1 (NF1) (MIM 162200) is one of themost common inherited disorders in humans with a prevalenceof 1 in 3000 individuals. NF1 is an autosomal dominantdisorder fully penetrant at the age of 5 years, but with a vari-able clinical expression, even among members of the samefamily. The main clinical features of NF1 are cafe au lait spots(CLSs), cutaneous neurofibromas and Lisch nodules (1,2). TheNF1 gene was mapped to chromosome 17q11.2 and was posi-tionally cloned in 1990 (3–5). It spans 350 kb of genomicDNA, contains 60 exons (6,7) and is transcribed ubiquitouslyto an mRNA of 11–13 kb that encodes for a protein, neuro-fibromin, of 2818 amino acids (8). The only region of neuro-fibromin with a well-defined function and structure is a centraldomain with a high similarity to ras-specific GTPase-activating proteins (GAPs), called the GAP-related domain(GRD), which downregulates ras activity (9–11).The NF1 gene has one of the highest mutation ratesdescribed for any human disorder ( 1 10

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Judit García

Papers

Mutations affecting mRNA splicing are the most common molecular defects in patients with neurofibromatosis type 1