Missed threads

Baralle, Diana; Lucassen, Anneke; Buratti, Emanuele

doi:10.1038/embor.2009.170

Cited by 105 publications

(84 citation statements)

References 25 publications

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“…As genetic testing improves and becomes more widely requested, RT-PCR analysis will also be important to interpret the clinical significance of unclassified sequence variants. 12 To perform RT-PCR analysis, an appropriate RNA source has to be available. Peripheral blood is an easily accessible source.…”

Section: Discussionmentioning

confidence: 99%

Analysis of LDLR mRNA in patients with familial hypercholesterolemia revealed a novel mutation in intron 14, which activates a cryptic splice site

2010

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Familial hypercholesterolemia (FH) is caused by a defective low-density lipoprotein receptor (LDLR), and 41000 mutations in LDLR have been identified. However, in some patients with clinically defined FH, no mutation can be detected within the exons and adjacent intronic segments of the LDLR. We have analyzed RNA extracted from blood samples of patients with clinically defined FH and identified an aberrantly spliced mRNA containing an 81-bp insert from intron 14. The aberrant splicing was caused by a novel intronic mutation, c.2140+86C4G, which activated a cryptic splice site. Although the cryptic splice site does not completely surpass the normal splice site, the mutation was found to cosegregate with high cholesterol levels in a family, which supports the notion that c.2140+86C4G causes FH. The insertion of 81 bp in LDLR mRNA encodes an in-frame insertion of 27 amino acids in the LDLR. However, the insertion was found to hamper LDLR activity by preventing the receptor from leaving the endoplasmic reticulum, probably because of misfolding of the protein. In patients with clinically defined hypercholesterolemia, despite normal results from sequencing of exonic regions of the LDLR gene, characterization of the LDLR mRNA might identify the underlying genetic defect.

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Section: Discussionmentioning

confidence: 99%

Analysis of LDLR mRNA in patients with familial hypercholesterolemia revealed a novel mutation in intron 14, which activates a cryptic splice site

2010

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“…The best chance of recognizing a splicing pathology caused by a synonymous mutation relies on bioinformatics analysis using a combination of various predictive algorithms. 24 However, also the bioinformatics approach lacks specificity and, indeed, in our patient, studies at the mRNA levels were at first performed because the consequences of the synonymous change did not fit with the disease phenotype. These studies confirmed a major effect on splicing and thus, retrospectively, attested to the presence of exonic regulatory sequences in the region spanning the mutation site.…”

Section: Spink5 Mutation Identificationmentioning

confidence: 99%

“…Controversial models are not unusual and reflect the nature of ESE and ESS sequences, which are redundant and degenerate, and the complexity of the splicing process, which is finely regulated by an interplay between different regulatory elements. [16][17][18][19]24 Mutation c.5080G4T in BRCA1 provides a precedent. This variant causes exon 18 skipping and was initially thought to disrupt an ESE for SF2/ASF by in silico analysis.…”

Section: Spink5 Mutation Identificationmentioning

confidence: 99%

“…23,28 Further studies will allow to better decipher the mechanism underlying splicing regulation of SPINK5 exon 11 and to explore possible therapeutic strategies. 24 Up to know, the c.891C4T variant represents the first example of an ESE/ESS mutation identified in the SPINK5 gene. In addition to SPINK5, several other genes have been targeted by mutations in ESE/ESS motifs, including BRCA1, 26 SMN1/2, 29,30 PDHA1, 31 GH, 32 DMD 33 and COL7A1.…”

Section: Spink5 Mutation Identificationmentioning

confidence: 99%

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A synonymous mutation in SPINK5 exon 11 causes Netherton syndrome by altering exonic splicing regulatory elements

et al. 2012

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Netherton syndrome (NS) is a rare, life-threatening ichthyosiform syndrome caused by recessive loss-of-function mutations in SPINK5 gene encoding lymphoepithelial Kazal-type-related inhibitor (LEKTI), a serine protease inhibitor expressed in the most differentiated epidermal layers and crucial for skin barrier function. We report the functional characterization of a previously unrecognized synonymous variant, c.891C4T (p.Cys297Cys), identified in the SPINK5 exon 11 of an NS patient. We demonstrated that the c.891C4T mutation is associated with abnormal pre-mRNA splicing and residual LEKTI expression in the patient's keratinocytes. Subsequent minigene splicing assays and in silico predictions confirmed the direct role of the synonymous mutation in inhibiting exon 11 inclusion by a mechanism that involves the activity of exonic regulatory sequences, namely splicing enhancer and silencer. However, this deleterious effect was not complete and a residual amount of normal mRNA and LEKTI protein could be detected, correlating with the relatively mild patient's phenotype. Our study represents the first identification of a disease-causing SPINK5 mutation that alters splicing without affecting canonical splice sites.

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“…Recently, several methods have been developed to evaluate the clinical effect of mutations that may cause splicing defects. 12 As is intuitively obvious, direct analysis of the mature mRNA from the patient remains the most reliable method to determine whether or not a genetic variation affects splicing. However, cells/RNA from the patient might not be available or the transcript may be expressed only in highly selected tissues, making this approach not always possible.…”

Section: Introductionmentioning

confidence: 99%

Splicing mutations in glycogen-storage disease type II: evaluation of the full spectrum of mutations and their relation to patients’ phenotypes

et al. 2010

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Glycogen-storage disease type II is an autosomal recessive-inherited disorder due to the deficiency of acid a-glucosidase. A large number of mutations in the acid a-glucosidase gene have been described to date. Among them, B15% are variations that may affect mRNA splicing process. In this study, we have for the first time comprehensively reviewed the available information on splicing mutations of the acid a-glucosidase gene and we have evaluated their possible impact on the splicing process using different in silico approaches. Out of the 39 different GAA-sequence variations described, an in silico analysis using seven different programs showed that 97% of them are predicted to have an impact on the splicing process. Moreover, this analysis showed a quite good correlation between the impact of the mutation on the splicing process and the clinical phenotype. In addition, we have performed the functional characterization of three novel sequence variants found in Italian patients and still uncharacterized. Using a minigene system, we have confirmed their pathogenic nature. In conclusion, this study has shown that in silico analysis represents a useful tool to select mutations that affect the splicing process of the acid a-glucosidase gene and provides an updated picture of all this kind of mutations reported till now.

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Missed threads

Cited by 105 publications

References 25 publications

Analysis of LDLR mRNA in patients with familial hypercholesterolemia revealed a novel mutation in intron 14, which activates a cryptic splice site

Analysis of LDLR mRNA in patients with familial hypercholesterolemia revealed a novel mutation in intron 14, which activates a cryptic splice site

A synonymous mutation in SPINK5 exon 11 causes Netherton syndrome by altering exonic splicing regulatory elements

Splicing mutations in glycogen-storage disease type II: evaluation of the full spectrum of mutations and their relation to patients’ phenotypes

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