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首页> 外文期刊>International Journal of Molecular Sciences >Altered Pre-mRNA Splicing Caused by a Novel Intronic Mutation c.1443+5GA in the Dihydropyrimidinase (DPYS) Gene
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Altered Pre-mRNA Splicing Caused by a Novel Intronic Mutation c.1443+5GA in the Dihydropyrimidinase (DPYS) Gene

机译：二氢嘧啶酶（DPYS）基因中的新型内含子突变c.1443 + 5G> A引起的改变的前mRNA剪接。

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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+5GA in intron 8 and a previously described missense mutation c.1001AG (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+5GA 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+5GA mutation results in aberrant splicing of the pre-mRNA encoding DHP, underlying the DHP deficiency in two unrelated Chinese patients.
机译：二氢嘧啶酶（DHP）缺乏症是由DPYS基因突变引起的常染色体隐性遗传疾病。患者的尿液，血液和脑脊液中的二氢尿嘧啶和二氢胸腺嘧啶含量很高。 DHP主要在肝和肾细胞中表达这一事实妨碍了对DPYS突变对mRNA前剪接的影响的分析。小基因方法可以使用不表达内源性mRNA的细胞检测mRNA剪接畸变。我们已经使用基于微基因的方法来分析推定的pre-mRNA剪接突变对两名新发现的中国小儿DHP缺乏症患者的影响。 DPYS的突变分析表明，这两名患者都是内含子8中新的内含子突变c.1443 + 5G> A和外显子6中先前描述的错义突变c.1001A> G（p.Q334R）的复合杂合。含有DPYS外显子7、8和9的突变小基因构建体在HEK293细胞中表达后产生不同的剪接产物。 c.1443 + 5G> A突变导致DPYS小基因构建体的前mRNA剪接发生改变，外显子8被完全跳过。对DHP晶体结构的分析表明，外显子8的缺失严重影响了其折叠，稳定性和均聚。酶以及催化位点的破坏。因此，分析表明c.1443 + 5G> A突变导致编码DHP的前mRNA的异常剪接，这是两名无关中国患者中DHP缺乏的基础。

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