PDF(Pubmed)
Abstract：
UNASSIGNED: Genetic diagnosis of Alport syndrome (AS), which results from pathogenic variants in COL4A3, COL4A4, or COL4A5 genes, is hindered by large numbers of unclassified variants detected using next-generation sequencing (NGS). We examined the impact on splicing of variants of uncertain significance in COL4A3 to COL4A5.
UNASSIGNED: Nine unrelated patients with clinical diagnosis or suspicion of AS were enrolled according to the criteria. Their clinical and genetic data were collected. Blood and urine samples were obtained from the patients and their family members. Sanger sequencing was used to confirm the 9 COL4A3 to COL4A5 unclassified variants identified by NGS. COL4A3 to COL4A5 mRNAs from urine were analyzed using targeted reverse transcription polymerase chain reaction and direct sequencing.
UNASSIGNED: Nine COL4A3 to COL4A5 unclassified variants were found to alter mRNAs splicing. Skipping of an exon or an exon fragment was induced by variants COL4A3 c.828+5G>A; COL4A4 c.3506-13_3528del; and COL4A5 c.451A>G (p. [Ile151Val]), c.2042-9 T>G, c.2689 G>C (p. [Glu897Gln]) and c.1033-10_1033-2delGGTAATAAA. Retention of an intron fragment was caused by variants COL4A3 c.3211-30G＞T, and COL4A5 c.4316-20T>A and c.1033-10 G>A, respectively. The 9 families in this study obtained genetic diagnosis of AS, including 3 with autosomal recessive AS and 6 with X-linked AS.
UNASSIGNED: Our findings demonstrate that urine mRNA analysis facilitates the identification of abnormal splicing of unclassified variants in Alport genes, which provides evidence of routine use of RNA analysis to improve genetic diagnosis of AS.
UNASSIGNED: Nine unrelated patients with clinical diagnosis or suspicion of AS were enrolled according to the criteria. Their clinical and genetic data were collected. Blood and urine samples were obtained from the patients and their family members. Sanger sequencing was used to confirm the 9 COL4A3 to COL4A5 unclassified variants identified by NGS. COL4A3 to COL4A5 mRNAs from urine were analyzed using targeted reverse transcription polymerase chain reaction and direct sequencing.
UNASSIGNED: Nine COL4A3 to COL4A5 unclassified variants were found to alter mRNAs splicing. Skipping of an exon or an exon fragment was induced by variants COL4A3 c.828+5G>A; COL4A4 c.3506-13_3528del; and COL4A5 c.451A>G (p. [Ile151Val]), c.2042-9 T>G, c.2689 G>C (p. [Glu897Gln]) and c.1033-10_1033-2delGGTAATAAA. Retention of an intron fragment was caused by variants COL4A3 c.3211-30G＞T, and COL4A5 c.4316-20T>A and c.1033-10 G>A, respectively. The 9 families in this study obtained genetic diagnosis of AS, including 3 with autosomal recessive AS and 6 with X-linked AS.
UNASSIGNED: Our findings demonstrate that urine mRNA analysis facilitates the identification of abnormal splicing of unclassified variants in Alport genes, which provides evidence of routine use of RNA analysis to improve genetic diagnosis of AS.
摘要：
■Alport综合征（AS）的基因诊断，由COL4A3，COL4A4或COL4A5基因的致病变异产生，受到使用下一代测序(NGS)检测到的大量未分类变体的阻碍。我们检查了COL4A3至COL4A5中不确定意义的变体对剪接的影响。
■根据标准纳入9名临床诊断或怀疑为AS的无关患者。收集他们的临床和遗传数据。从患者及其家庭成员获得血液和尿液样本。Sanger测序用于确认NGS鉴定的9个COL4A3至COL4A5未分类变体。使用靶向逆转录聚合酶链反应和直接测序分析来自尿液的COL4A3至COL4A5mRNA。
■发现九种COL4A3至COL4A5未分类变体改变mRNA剪接。通过变体COL4A3c.8285G>A诱导外显子或外显子片段的跳跃；COL4A4c.3506-13_3528del；和COL4A5c.41A>G（p。[Ile151Val]），c.2042-9T>G，c.2689G>C（p。[Glu897Gln]）和c.1033-10_1033-2delGGTAATAAA。内含子片段的保留是由变异COL4A3c.3211-30G＞T引起的，和COL4A5c.4316-20T>A和c.1033-10G>A，分别。本研究中的9个家庭获得了AS的遗传诊断，包括3例常染色体隐性遗传AS和6例X连锁AS。
■我们的研究结果表明，尿液mRNA分析有助于鉴定Alport基因中未分类变体的异常剪接，这提供了常规使用RNA分析来改善AS的遗传诊断的证据。
■根据标准纳入9名临床诊断或怀疑为AS的无关患者。收集他们的临床和遗传数据。从患者及其家庭成员获得血液和尿液样本。Sanger测序用于确认NGS鉴定的9个COL4A3至COL4A5未分类变体。使用靶向逆转录聚合酶链反应和直接测序分析来自尿液的COL4A3至COL4A5mRNA。
■发现九种COL4A3至COL4A5未分类变体改变mRNA剪接。通过变体COL4A3c.8285G>A诱导外显子或外显子片段的跳跃；COL4A4c.3506-13_3528del；和COL4A5c.41A>G（p。[Ile151Val]），c.2042-9T>G，c.2689G>C（p。[Glu897Gln]）和c.1033-10_1033-2delGGTAATAAA。内含子片段的保留是由变异COL4A3c.3211-30G＞T引起的，和COL4A5c.4316-20T>A和c.1033-10G>A，分别。本研究中的9个家庭获得了AS的遗传诊断，包括3例常染色体隐性遗传AS和6例X连锁AS。
■我们的研究结果表明，尿液mRNA分析有助于鉴定Alport基因中未分类变体的异常剪接，这提供了常规使用RNA分析来改善AS的遗传诊断的证据。
