Molecular dissection of germline chromothripsis in a developmental context using patient-derived iPS cells
Molecular dissection of germline chromothripsis in a developmental context using patient-derived iPS cells
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DOI：
10.1186/s13073-017-0399-z
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发表时间：
2017-01-26
期刊：
影响因子：
12.3
通讯作者：
Cuppen, Edwin
中科院分区：
文献类型：
--
作者：
Middelkamp, Sjors;van Heesch, Sebastiaan;Cuppen, Edwin
背景：生殖系染色体碎裂会导致复杂的基因组重排，这可能影响多个基因及其调控环境。复杂生殖系基因组重排患者中个体重排和受影响基因对表型的作用通常是未知的。
方法：为了在相关发育背景下剖析生殖系染色体碎裂的影响，我们对一名患有新生生殖系染色体碎裂的患者及其健康的双亲的血细胞、诱导多能干细胞（iPSCs）以及iPSC衍生的神经元细胞进行了基于三人组的RNA表达分析。此外，还进行了Hi - C和4C - seq实验以确定基因组重排对断点连接处附近基因转录调控的影响。
结果：67个基因位于涉及4条染色体上17个断点的复杂染色体碎裂重排的1 Mb范围内。我们发现其中3个基因（FOXP1、DPYD和TWIST1）均与发育障碍有关，且在患者中差异表达。有趣的是，对TWIST1表达的影响仅在患者的iPSC衍生的神经元细胞中可检测到，这强调了在生物学相关背景下研究发育障碍的必要性。染色体构象捕获分析表明，由于染色体碎裂事件，TWIST1失去了与几个增强子的基因组相互作用，这可能导致TWIST1表达失调，并促成了患者的颅缝早闭表型。
结论：我们证明，患者来源的iPSC分化与基于三人组的分子图谱分析相结合是一种有力的方法，可提高对致病性复杂基因组重排的解读。在此，我们应用这种方法确定了TWIST1、FOXP1和DPYD的错误表达是生殖系染色体碎裂重排导致的复杂先天性表型的关键因素。
Background: Germline chromothripsis causes complex genomic rearrangements that are likely to affect multiple genes and their regulatory contexts. The contribution of individual rearrangements and affected genes to the phenotypes of patients with complex germline genomic rearrangements is generally unknown.Methods: To dissect the impact of germline chromothripsis in a relevant developmental context, we performed trio-based RNA expression analysis on blood cells, induced pluripotent stem cells (iPSCs), and iPSC-derived neuronal cells from a patient with de novo germline chromothripsis and both healthy parents. In addition, Hi-C and 4C-seq experiments were performed to determine the effects of the genomic rearrangements on transcription regulation of genes in the proximity of the breakpoint junctions.Results: Sixty-seven genes are located within 1 Mb of the complex chromothripsis rearrangements involving 17 breakpoints on four chromosomes. We find that three of these genes (FOXP1, DPYD, and TWIST1) are both associated with developmental disorders and differentially expressed in the patient. Interestingly, the effect on TWIST1 expression was exclusively detectable in the patient's iPSC-derived neuronal cells, stressing the need for studying developmental disorders in the biologically relevant context. Chromosome conformation capture analyses show that TWIST1 lost genomic interactions with several enhancers due to the chromothripsis event, which likely led to deregulation of TWIST1 expression and contributed to the patient's craniosynostosis phenotype.Conclusions: We demonstrate that a combination of patient-derived iPSC differentiation and trio-based molecular profiling is a powerful approach to improve the interpretation of pathogenic complex genomic rearrangements. Here we have applied this approach to identify misexpression of TWIST1, FOXP1, and DPYD as key contributors to the complex congenital phenotype resulting from germline chromothripsis rearrangements.