Linked Life Data

15979033

Source:http://linkedlifedata.com/resource/pubmed/id/15979033

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
3
pubmed:dateCreated
2005-6-27
pubmed:abstractText
The splicing pattern of pre-mRNA is unpredictable in genes harboring a single-nucleotide change within the consensus sequence of a splice-donor site. In the dystrophin gene, a transition from G to A at the fifth position of intron-32 (4518+5G > A) has been reported as a polymorphism within the consensus sequence or a mutation identified in Duchenne muscular dystrophy (DMD). Here, we report both in vivo and in vitro evidence that shows inactivation of the splice-donor site caused by this mutation. In one Japanese DMD case, two novel dystrophin mRNAs were identified in the patient's lymphocytes, one with a 98 bp deletion of the 3' end of exon-32 (dys32-98) and the other with a 28 bp intron retained between exons 32 and 33 (dys32 + 28). Genomic sequencing disclosed a single-nucleotide change from G to A at the fifth position of intron-32 (4518+5G > A). To demonstrate in vitro the inactivation of this splice-donor site by this nucleotide change, mini-dystrophin genes comprising three exons harboring either normal or mutant intron-32 sequences were expressed in HeLa cells, and the splicing products were analyzed by reverse-transcription PCR amplification. A normal transcript consisting of three exons was obtained from the normal construct. From the mutant, we obtained one product containing a 98 bp deletion at the 3' end of exon-32, indicating complete inactivation of the native splice-donor site. Thus, both in vivo and in vitro experiments demonstrate that 4518+5G > A causes a splicing error leading to transcript termination; it did not behave like a silent polymorphism. Our results indicate that the in vitro splicing system is a powerful tool for determining the underlying mechanism of a disease-causing mutation in a splicing consensus sequence.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jul
pubmed:issn
1096-7192
pubmed:author
pubmed:issnType
Print
pubmed:volume
85
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
213-9
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
2005
pubmed:articleTitle
A G-to-A transition at the fifth position of intron-32 of the dystrophin gene inactivates a splice-donor site both in vivo and in vitro.
pubmed:affiliation
Department of Pediatrics, Kobe University Graduate School of Medicine, 7-5-1 Kusunokicho, Chuo, Kobe 650-0017, Japan.
pubmed:publicationType
Journal Article, Case Reports, Research Support, Non-U.S. Gov't