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Altered Pre-mRNA Splicing Caused by a Novel Intronic Mutation c.1443+5G>A in the Dihydropyrimidinase (DPYS) Gene

Dihydropyrimidinase (DHP) deficiency is an autosomal recessive disease caused by mutations in the DPYS gene. Patients present with highly elevated levels of dihydrouracil and dihydrothymine in their urine, blood and cerebrospinal fluid. The analysis of the effect of mutations in DPYS on pre-mRNA spl...

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Autores principales: Nakajima, Yoko, Meijer, Judith, Zhang, Chunhua, Wang, Xu, Kondo, Tomomi, Ito, Tetsuya, Dobritzsch, Doreen, Van Kuilenburg, André B. P.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4730329/
https://www.ncbi.nlm.nih.gov/pubmed/26771602
http://dx.doi.org/10.3390/ijms17010086
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author Nakajima, Yoko
Meijer, Judith
Zhang, Chunhua
Wang, Xu
Kondo, Tomomi
Ito, Tetsuya
Dobritzsch, Doreen
Van Kuilenburg, André B. P.
author_facet Nakajima, Yoko
Meijer, Judith
Zhang, Chunhua
Wang, Xu
Kondo, Tomomi
Ito, Tetsuya
Dobritzsch, Doreen
Van Kuilenburg, André B. P.
author_sort Nakajima, Yoko
collection PubMed
description Dihydropyrimidinase (DHP) deficiency is an autosomal recessive disease caused by mutations in the DPYS gene. Patients present with highly elevated levels of dihydrouracil and dihydrothymine in their urine, blood and cerebrospinal fluid. The analysis of the effect of mutations in DPYS on pre-mRNA splicing is hampered by the fact that DHP is primarily expressed in liver and kidney cells. The minigene approach can detect mRNA splicing aberrations using cells that do not express the endogenous mRNA. We have used a minigene-based approach to analyze the effects of a presumptive pre-mRNA splicing mutation in two newly identified Chinese pediatric patients with DHP deficiency. Mutation analysis of DPYS showed that both patients were compound heterozygous for a novel intronic mutation c.1443+5G>A in intron 8 and a previously described missense mutation c.1001A>G (p.Q334R) in exon 6. Wild-type and the mutated minigene constructs, containing exons 7, 8 and 9 of DPYS, yielded different splicing products after expression in HEK293 cells. The c.1443+5G>A mutation resulted in altered pre-mRNA splicing of the DPYS minigene construct with full skipping of exon 8. Analysis of the DHP crystal structure showed that the deletion of exon 8 severely affects folding, stability and homooligomerization of the enzyme as well as disruption of the catalytic site. Thus, the analysis suggests that the c.1443+5G>A mutation results in aberrant splicing of the pre-mRNA encoding DHP, underlying the DHP deficiency in two unrelated Chinese patients.
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spelling pubmed-47303292016-02-11 Altered Pre-mRNA Splicing Caused by a Novel Intronic Mutation c.1443+5G>A in the Dihydropyrimidinase (DPYS) Gene Nakajima, Yoko Meijer, Judith Zhang, Chunhua Wang, Xu Kondo, Tomomi Ito, Tetsuya Dobritzsch, Doreen Van Kuilenburg, André B. P. Int J Mol Sci Article Dihydropyrimidinase (DHP) deficiency is an autosomal recessive disease caused by mutations in the DPYS gene. Patients present with highly elevated levels of dihydrouracil and dihydrothymine in their urine, blood and cerebrospinal fluid. The analysis of the effect of mutations in DPYS on pre-mRNA splicing is hampered by the fact that DHP is primarily expressed in liver and kidney cells. The minigene approach can detect mRNA splicing aberrations using cells that do not express the endogenous mRNA. We have used a minigene-based approach to analyze the effects of a presumptive pre-mRNA splicing mutation in two newly identified Chinese pediatric patients with DHP deficiency. Mutation analysis of DPYS showed that both patients were compound heterozygous for a novel intronic mutation c.1443+5G>A in intron 8 and a previously described missense mutation c.1001A>G (p.Q334R) in exon 6. Wild-type and the mutated minigene constructs, containing exons 7, 8 and 9 of DPYS, yielded different splicing products after expression in HEK293 cells. The c.1443+5G>A mutation resulted in altered pre-mRNA splicing of the DPYS minigene construct with full skipping of exon 8. Analysis of the DHP crystal structure showed that the deletion of exon 8 severely affects folding, stability and homooligomerization of the enzyme as well as disruption of the catalytic site. Thus, the analysis suggests that the c.1443+5G>A mutation results in aberrant splicing of the pre-mRNA encoding DHP, underlying the DHP deficiency in two unrelated Chinese patients. MDPI 2016-01-12 /pmc/articles/PMC4730329/ /pubmed/26771602 http://dx.doi.org/10.3390/ijms17010086 Text en © 2016 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons by Attribution (CC-BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Nakajima, Yoko
Meijer, Judith
Zhang, Chunhua
Wang, Xu
Kondo, Tomomi
Ito, Tetsuya
Dobritzsch, Doreen
Van Kuilenburg, André B. P.
Altered Pre-mRNA Splicing Caused by a Novel Intronic Mutation c.1443+5G>A in the Dihydropyrimidinase (DPYS) Gene
title Altered Pre-mRNA Splicing Caused by a Novel Intronic Mutation c.1443+5G>A in the Dihydropyrimidinase (DPYS) Gene
title_full Altered Pre-mRNA Splicing Caused by a Novel Intronic Mutation c.1443+5G>A in the Dihydropyrimidinase (DPYS) Gene
title_fullStr Altered Pre-mRNA Splicing Caused by a Novel Intronic Mutation c.1443+5G>A in the Dihydropyrimidinase (DPYS) Gene
title_full_unstemmed Altered Pre-mRNA Splicing Caused by a Novel Intronic Mutation c.1443+5G>A in the Dihydropyrimidinase (DPYS) Gene
title_short Altered Pre-mRNA Splicing Caused by a Novel Intronic Mutation c.1443+5G>A in the Dihydropyrimidinase (DPYS) Gene
title_sort altered pre-mrna splicing caused by a novel intronic mutation c.1443+5g>a in the dihydropyrimidinase (dpys) gene
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4730329/
https://www.ncbi.nlm.nih.gov/pubmed/26771602
http://dx.doi.org/10.3390/ijms17010086
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