We previously identified the protein Lbh as necessary for cranial neural crest (CNC) cell migration in Xenopus through the use of morpholinos. However, Lbh is a maternally deposited protein and morpholinos achieve knockdowns through prevention of translation. In order to investigate the role of Lbh in earlier embryonic events, we employed the new technique “Trim-Away” to degrade this maternally deposited protein. Trim-Away utilizes the E3 ubiquitin ligase trim21 to degrade proteins targeted with an antibody and was developed in mammalian systems. Our results show that Xenopus is amenable to the Trim-Away technique. We also show that early knockdown of Lbh in Xenopus results in defects in gastrulation that present with a decrease in fibronectin matrix assembly, an increased in mesodermal cell migration and decrease in endodermal cell cohesion. We further show that the technique is also effective on a second abundant maternal protein PACSIN2. We discuss potential advantages and limit of the technique in Xenopus embryos as well as the mechanism of gastrulation inhibition.

我们之前通过使用morpholinos 将蛋白质Lbh 确定为非洲爪蟾中颅神经嵴（CNC）细胞迁移所必需的。然而，Lbh 是一种母体沉积的蛋白质，而吗啉通过阻止翻译实现击倒。为了研究 Lbh 在早期胚胎事件中的作用，我们采用了新技术“ Trim-Away ”来降解这种母体沉积的蛋白质。Trim-Away 利用 E3 泛素连接酶 trim21 降解抗体靶向的蛋白质，并在哺乳动物系统中开发。我们的结果表明，非洲爪蟾适合 Trim-Away 技术。我们还表明非洲爪蟾中 Lbh 的早期敲除导致原肠形成缺陷，表现为纤连蛋白基质组装减少，中胚层细胞迁移增加和内胚层细胞凝聚力降低。我们进一步表明该技术对第二种丰富的母体蛋白 PACSIN2 也有效。我们讨论了该技术在非洲爪蟾胚胎中的潜在优势和局限性以及抑制原肠胚形成的机制。