Modelling the developmental spliceosomal craniofacial disorder Burn-McKeown syndrome using induced pluripotent stem cells
Modelling the developmental spliceosomal craniofacial disorder Burn-McKeown syndrome using induced pluripotent stem cells
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DOI：
10.1371/journal.pone.0233582
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发表时间：
2020-05
期刊：
影响因子：
3.7
通讯作者：
Katherine A. Wood;Charlie F. Rowlands;Huw B. Thomas;Steven Woods;Julieta O’Flaherty;S. Douzgou;S. Kimber;W. Newman;R. O’Keefe
中科院分区：
文献类型：
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作者：
Katherine A. Wood;Charlie F. Rowlands;Huw B. Thomas;Steven Woods;Julieta O’Flaherty;S. Douzgou;S. Kimber;W. Newman;R. O’Keefe
头面部发育障碍Burn-McKeown综合征(BMKS)是由前信使RNA剪接因子基因TXNL4A/DIB1的双等位变异引起的。大多数BMKS患者中，TXNL4A的一个等位基因的启动子区域有34个碱基对缺失，另一个等位基因的功能缺失变异，导致TXNL4A的表达减少。然而，目前尚不清楚这种普遍表达的剪接体蛋白的表达减少如何导致发育过程中的头面部缺陷。在这里，我们将一名BMKS患者和她未受影响的母亲的外周单个核血细胞重新编程为诱导多能干细胞(IPSCs)，并将IPSCs分化为诱导神经脊细胞(INCCs)，这是正常颅面发育所需的关键细胞类型。BMKS患者来源的IPSCs的增殖速度慢于母亲来源和无关对照来源的IPSCs，RNA-Seq分析显示在基因表达和选择性剪接方面存在显著差异。与母体和无关的对照IPSCs相比，患者IPSCs表现出向iNCC分化的缺陷，特别是在经历上皮向间充质转化(EMT)方面的延迟。对分化的iNCC的RNA-Seq分析显示，与颅面和胚胎发育相关的基因普遍存在表达变化和错误剪接，这突显了对WNT信号的抑制反应，WNT信号是iNCC分化过程中激活的关键途径。此外，我们发现WNT途径中的关键基因TCF7L2外显子4的错误剪接是患者细胞中WNT反应下调的潜在原因。此外，错误剪接的基因具有共同的序列属性，如长度、剪接位点强度和序列基序，这表明特定基因亚集的剪接对TXNL4A表达的变化特别敏感。综上所述，这些数据首次揭示了BMKS患者TXNL4A表达减少如何影响剪接和NCC功能，导致胚胎颅面发育缺陷。
The craniofacial developmental disorder Burn-McKeown Syndrome (BMKS) is caused by biallelic variants in the pre-messenger RNA splicing factor gene TXNL4A/DIB1. The majority of affected individuals with BMKS have a 34 base pair deletion in the promoter region of one allele of TXNL4A combined with a loss-of-function variant on the other allele, resulting in reduced TXNL4A expression. However, it is unclear how reduced expression of this ubiquitously expressed spliceosome protein results in craniofacial defects during development. Here we reprogrammed peripheral mononuclear blood cells from a BMKS patient and her unaffected mother into induced pluripotent stem cells (iPSCs) and differentiated the iPSCs into induced neural crest cells (iNCCs), the key cell type required for correct craniofacial development. BMKS patient-derived iPSCs proliferated more slowly than both mother- and unrelated control-derived iPSCs, and RNA-Seq analysis revealed significant differences in gene expression and alternative splicing. Patient iPSCs displayed defective differentiation into iNCCs compared to maternal and unrelated control iPSCs, in particular a delay in undergoing an epithelial-to-mesenchymal transition (EMT). RNA-Seq analysis of differentiated iNCCs revealed widespread gene expression changes and mis-splicing in genes relevant to craniofacial and embryonic development that highlight a dampened response to WNT signalling, the key pathway activated during iNCC differentiation. Furthermore, we identified the mis-splicing of TCF7L2 exon 4, a key gene in the WNT pathway, as a potential cause of the downregulated WNT response in patient cells. Additionally, mis-spliced genes shared common sequence properties such as length, splice site strengths and sequence motifs, suggesting that splicing of particular subsets of genes is particularly sensitive to changes in TXNL4A expression. Together, these data provide the first insight into how reduced TXNL4A expression in BMKS patients might compromise splicing and NCC function, resulting in defective craniofacial development in the embryo.