EFTUD2 gene deficiency disrupts osteoblast maturation and inhibits chondrocyte differentiation via activation of the p53 signaling pathway

Background: Mandibulofacial dysostosis with microcephaly (MFDM) is characteristic of multiple skeletal anomalies comprising craniofacial anomalies/dysplasia, microcephaly, dysplastic ears, choanal atresia, and short stature. Heterozygous loss of function variants of EFTUD2 was previously reported in MFDM; however, the mechanism underlying EFTUD2-associated skeletal dysplasia remains unclear.Results: We identified a novel frameshift variant of EFTUD2 (c.1030_1031delTG, p.Trp344fs*2) in an MFDM Chinese patient with craniofacial dysmorphism including ear canal structures and microcephaly, mild intellectual disability, and developmental delay. We generated a zebrafish model of eftud2 deficiency, and a consistent phenotype consisting of mandibular bone dysplasia and otolith loss was observed. We also showed that EFTUD2 deficiency significantly inhibited proliferation, differentiation, and maturation in human calvarial osteoblast (HCO) and human articular chondrocyte (HC-a) cells. RNA-Seq analysis uncovered activated TP53 signaling with increased phosphorylation of the TP53 protein and upregulation of five TP53 downstream target genes (FAS, STEAP3, CASP3, P21, and SESN1) both in HCO and in eftud2-/- zebrafish. Additionally, inhibition of p53 by morpholino significantly reduced the mortality of eftud2-/- larvae.Conclusions: Our results confirm a novel de novo variant of the EFTUD2 gene and suggest that EFTUD2 may participate in the maturation and differentiation of osteoblasts and chondrocytes, possibly via activation of the TP53 signaling pathway. Thus, mutations in this gene may lead to skeletal anomalies in vertebrates.

背景：小头畸形的下颌面骨发育不全（MFDM）具有多种骨骼异常的特征，包括颅面异常/发育不良、小头畸形、耳朵发育不良、后鼻孔闭锁以及身材矮小。此前有报道称EFTUD2的杂合性功能缺失变异存在于MFDM中；然而，EFTUD2相关骨骼发育不良的潜在机制仍不清楚。 结果：我们在一名患有颅面畸形（包括耳道结构异常和小头畸形）、轻度智力障碍以及发育迟缓的中国MFDM患者中鉴定出一种新的EFTUD2移码变异（c.1030_1031delTG，p.Trp344fs*2）。我们构建了eftud2缺陷的斑马鱼模型，并观察到包括下颌骨发育不良和耳石缺失在内的一致表型。我们还表明，EFTUD2缺陷显著抑制人颅骨成骨细胞（HCO）和人关节软骨细胞（HC - a）的增殖、分化和成熟。RNA - Seq分析发现，在HCO以及eftud2 - / - 斑马鱼中，TP53信号通路被激活，TP53蛋白磷酸化增加，五个TP53下游靶基因（FAS、STEAP3、CASP3、P21和SESN1）上调。此外，吗啉代寡核苷酸对p53的抑制显著降低了eftud2 - / - 幼鱼的死亡率。 结论：我们的研究结果证实了EFTUD2基因的一种新的从头变异，并表明EFTUD2可能参与成骨细胞和软骨细胞的成熟和分化，可能是通过激活TP53信号通路实现的。因此，该基因的突变可能导致脊椎动物的骨骼异常。