Gene–environment interactions are believed to play a role in multifactorial phenotypes, although poorly described mechanistically. Cleft lip/palate (CLP), the most common craniofacial malformation, has been associated with both genetic and environmental factors, with little gene–environment interaction experimentally demonstrated. Here, we study CLP families harbouring CDH1/E-Cadherin variants with incomplete penetrance and we explore the association of pro-inflammatory conditions to CLP. By studying neural crest (NC) from mouse, Xenopus and humans, we show that CLP can be explained by a 2-hit model, where NC migration is impaired by a combination of genetic (CDH1 loss-of-function) and environmental (pro-inflammatory activation) factors, leading to CLP. Finally, using in vivo targeted methylation assays, we demonstrate that CDH1 hypermethylation is the major target of the pro-inflammatory response, and a direct regulator of E-cadherin levels and NC migration. These results unveil a gene–environment interaction during craniofacial development and provide a 2-hit mechanism to explain cleft lip/palate aetiology.

基因-环境相互作用被认为在多因素表型中发挥作用，尽管在机制上描述不佳。唇裂/腭裂 （CLP） 是最常见的颅面畸形，与遗传和环境因素有关，实验证明基因-环境相互作用很少。在这里，我们研究了携带不完全外显率的 CDH1/E-钙粘蛋白变体的 CLP 家族，并探讨了促炎性疾病与 CLP 的关系。通过研究小鼠、非洲爪蟾和人类的神经嵴 （NC），我们表明 CLP 可以用 2-hit 模型来解释，其中 NC 迁移受到遗传 （CDH1 功能丧失） 和环境 （促炎激活） 因素的组合损害，导致 CLP。最后，使用体内靶向甲基化测定，我们证明 CDH1 高甲基化是促炎反应的主要靶标，也是 E-钙粘蛋白水平和 NC 迁移的直接调节因子。这些结果揭示了颅面发育过程中的基因-环境相互作用，并提供了解释唇裂/腭裂病因的 2-hit 机制。