Please use this identifier to cite or link to this item: https://hdl.handle.net/2440/7212
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Type: Journal article
Title: Preferential pre-mRNA utilisation of an upstream cryptic 5' splice site created by a single base deletion mutation in exon 37 of the FBN-1 gene
Author: Gibson, M.
Ellis, S.
Ades, L.
Haan, E.
Cleary, E.
Citation: The Federation of European Biochemical Societies (FEBS) Journal, 1998; 256(1):221-228
Publisher: SPRINGER VERLAG
Issue Date: 1998
ISSN: 0014-2956
1432-1033
Abstract: A heterozygous deletion of a single base (A4704) from exon 37 of the fibrillin-1 gene was defined in a patient with Marfan syndrome and subsequently in his previously undiagnosed father. The deletion created a cryptic 5' splice site in exon 37 which was utilised in preference to the normal 5' splice site during pre-mRNA processing in skin fibroblasts cultured from the proband. The mutant mRNA showed a 48-bp deletion from the 3' end of exon 37 which was predicted to restore the reading frame in the mutant mRNA and result in the deletion of a 16-amino-acid sequence from a central eight-cysteine repeat motif of the fibrillin-1 molecule. Interestingly, the cryptic 5' splice site in exon 37 and the normal 5' splice site had equally strong consensuses for splice-site selection. The preferential utilisation of the cryptic site is discussed in relation to current theories on the mechanisms involved in pre-mRNA splicing. Analysis by reverse-transcription PCR indicated that, in the patients skin fibroblasts, the steady-state level of the mis-spliced mutant mRNA was close to that from the normal allele. In addition, evidence from immunoblotting and pulse-chase biosynthetic labelling indicated that close to normal amounts of fibrillin-1 were being synthesised and secreted by the cells. However, in contrast to control cells cultured from an unaffected individual, little fibrillin-1 was detected, either biosynthetically or by immunofluorescence, in the extracellular matrix produced by the proband's fibroblasts. Thus, the slightly shorter mutant fibrillin-1 molecules appeared to be exerting a powerful dominant-negative effect on the incorporation of normal fibrillin-1 molecules into microfibrils in this culture system. This severe inhibition of microfibril synthesis in cell culture contrasts with the 'classic' phenotype of the proband, suggesting that factors influencing microfibril formation may differ greatly between in vivo and in vitro environments.
Keywords: Fibroblasts
Humans
Marfan Syndrome
Microfilament Proteins
RNA Precursors
RNA, Messenger
Gene Amplification
RNA Splicing
Sequence Deletion
Amino Acid Sequence
Base Sequence
Sequence Homology, Amino Acid
Mutation
Exons
Molecular Sequence Data
Child
Male
Fibrillin-1
Fibrillins
Description: Article first published online: 25 DEC 2001
Rights: Copyright © 1998 John Wiley & Sons, Inc. All Rights Reserved.
DOI: 10.1046/j.1432-1327.1998.2560221.x
Appears in Collections:Aurora harvest 5
Paediatrics publications

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