Lan Yu, Zhang Nian, Liu Han, Xu Jingyue, Jiang Rulang
Division of Plastic Surgery, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA Division of Developmental Biology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA
Division of Developmental Biology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA.
Development. 2016 Jul 1;143(13):2344-55. doi: 10.1242/dev.134577. Epub 2016 May 25.
Cleft palate is a common major birth defect for which currently known causes account for less than 30% of pathology in humans. In this study, we carried out mutagenesis screening in mice to identify new regulators of palatogenesis. Through genetic linkage mapping and whole-exome sequencing, we identified a loss-of-function mutation in the Golgb1 gene that co-segregated with cleft palate in a new mutant mouse line. Golgb1 is a ubiquitously expressed large coiled-coil protein, also known as giantin, that is localized at the Golgi membrane. Using CRISPR/Cas9-mediated genome editing, we generated and analyzed developmental defects in mice carrying additional Golgb1 loss-of-function mutations, which supported a crucial requirement for Golgb1 in palate development. Through maxillary explant culture assays, we demonstrate that the Golgb1 mutant embryos have intrinsic defects in palatal shelf elevation. Just prior to the developmental stage of palatal shelf elevation in wild-type littermates, Golgb1 mutant embryos exhibit increased cell density, reduced hyaluronan accumulation and impaired protein glycosylation in the palatal mesenchyme. Together, these results demonstrate that, although it is a ubiquitously expressed Golgi-associated protein, Golgb1 has specific functions in protein glycosylation and tissue morphogenesis.
腭裂是一种常见的严重出生缺陷,目前已知的病因在人类病理中所占比例不到30%。在本研究中,我们在小鼠中进行了诱变筛选,以鉴定腭发育的新调节因子。通过遗传连锁图谱分析和全外显子组测序,我们在一个新的突变小鼠品系中鉴定出Golgb1基因的功能缺失突变,该突变与腭裂共分离。Golgb1是一种广泛表达的大型卷曲螺旋蛋白,也称为巨蛋白,定位于高尔基体膜。使用CRISPR/Cas9介导的基因组编辑,我们生成并分析了携带额外Golgb1功能缺失突变的小鼠的发育缺陷,这支持了Golgb1在腭发育中的关键需求。通过上颌外植体培养试验,我们证明Golgb1突变胚胎在腭突抬高方面存在内在缺陷。在野生型同窝小鼠腭突抬高的发育阶段之前,Golgb1突变胚胎在腭间充质中表现出细胞密度增加、透明质酸积累减少和蛋白质糖基化受损。总之,这些结果表明,尽管Golgb1是一种广泛表达的高尔基体相关蛋白,但它在蛋白质糖基化和组织形态发生中具有特定功能。