DNA damage response (DDR) genes orchestrating the network of DNA repair, cell cycle control, are essential for the rapid proliferation of neural progenitor cells. To date, the potential association between specific DDR genes and the risk of human neural tube defects (NTDs) has not been investigated. Using whole-genome sequencing and targeted sequencing, we identified significant enrichment of rare deleterious RAD9B variants in spina bifida cases compared to controls (8/409 vs. 0/298; p = .0241). Among the eight identified variants, the two frameshift mutants and p.Gln146Glu affected RAD9B nuclear localization. The two frameshift mutants also decreased the protein level of RAD9B. p.Ser354Gly, as well as the two frameshifts, affected the cell proliferation rate. Finally, p.Ser354Gly, p.Ser10Gly, p.Ile112Met, p.Gln146Glu, and the two frameshift variants showed a decreased ability for activating JNK phosphorylation. RAD9B knockdowns in human embryonic stem cells profoundly affected early differentiation through impairing PAX6 and OCT4 expression. RAD9B deficiency impeded in vitro formation of neural organoids, a 3D cell culture model for human neural development. Furthermore, the RNA-seq data revealed that loss of RAD9B dysregulates cell adhesion genes during organoid formation. These results represent the first demonstration of a DDR gene as an NTD risk factor in humans.

中文翻译：

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RAD9B 的缺失会损害早期神经发育并增加人类脊柱裂的风险。

协调 DNA 修复网络、细胞周期控制的 DNA 损伤反应 (DDR) 基因对于神经祖细胞的快速增殖至关重要。迄今为止，尚未研究特定 DDR 基因与人类神经管缺陷 (NTD) 风险之间的潜在关联。使用全基因组测序和靶向测序，我们发现与对照组相比，脊柱裂病例中罕见的有害 RAD9B 变体显着富集（8/409 对 0/298；p = .0241）。在八个已识别的变体中，两个移码突变体和 p.Gln146Glu 影响 RAD9B 核定位。这两个移码突变体也降低了 RAD9B 的蛋白质水平。p.Ser354Gly，以及两个移码，影响细胞增殖率。最后，p.Ser354Gly、p.Ser10Gly、p.Ile112Met、p.Gln146Glu、并且这两种移码变体显示出激活 JNK 磷酸化的能力降低。人类胚胎干细胞中的 RAD9B 敲低通过损害 PAX6 和 OCT4 表达而深刻影响早期分化。RAD9B 缺陷阻碍了神经类器官的体外形成，这是一种用于人类神经发育的 3D 细胞培养模型。此外，RNA-seq 数据显示，RAD9B 的缺失会在类器官形成过程中失调细胞粘附基因。这些结果代表了 DDR 基因作为人类 NTD 风险因素的首次证明。用于人类神经发育的 3D 细胞培养模型。此外，RNA-seq 数据显示，RAD9B 的缺失会在类器官形成过程中失调细胞粘附基因。这些结果代表了 DDR 基因作为人类 NTD 风险因素的首次证明。用于人类神经发育的 3D 细胞培养模型。此外，RNA-seq 数据显示，RAD9B 的缺失会在类器官形成过程中失调细胞粘附基因。这些结果代表了 DDR 基因作为人类 NTD 风险因素的首次证明。