Wülbeck C, Fromental-Ramain C, Campos-Ortega J A
Institut für Entwicklungsbiologie, Universität zu Köln, Germany.
Mech Dev. 1994 May;46(2):73-85. doi: 10.1016/0925-4773(94)90077-9.
We have identified a zebrafish homologue of the human E12 protein, which we have called ZFE12. It shows a high degree of identity to HE12 throughout its entire sequence, particularly in the bHLH domain (93%); amino acid sequence identity of the bHLH domain of ZFE12 to Drosophila DAUGHTERLESS is also very high (75%). During early embryogenesis, expression of ZfE12 is widespread. Following gastrulation, ZfE12 transcripts can be detected in all embryonic tissues with the exception of the notochord, although zones with relatively higher densities of transcripts are present in the developing brain and in the somites. To assay biological activities associated with the ZFE12 protein, two P-element constructs were made, each carrying a Drosophila daughterless gene that had been modified by replacing either the HLH domain or the entire C-terminus including the bHLH domain, by the equivalent domains of ZfE12. These constructs were used to transform flies and tested for their ability to rescue the daughterless mutant phenotype. Complete rescue of the neural phenotype and of the embryonic lethality was obtained. However, the daughterless function could not be completely restored. Although fertile flies transheterozygous for a hypomorphic and an amorphic mutation and carrying the construct encoding the zebrafish HLH domain emerged, they lacked various types of sensory organs on head and thorax and showed slight wing defects. Mutants carrying the second construct occasionally reached pupal stages but died subsequently. These data demonstrate that the HLH domain of ZFE12 can carry out most, but not all, functions performed by the corresponding region of the DAUGHTERLESS protein in Drosophila.
我们鉴定出了人类E12蛋白的斑马鱼同源物，我们将其命名为ZFE12。它在整个序列中与HE12具有高度的同一性，尤其是在bHLH结构域（93%）；ZFE12的bHLH结构域与果蝇的无女儿基因（DAUGHTERLESS）的氨基酸序列同一性也非常高（75%）。在胚胎早期发育过程中，ZfE12的表达很广泛。原肠胚形成后，除了脊索外，在所有胚胎组织中都能检测到ZfE12转录本，不过在发育中的大脑和体节中存在转录本密度相对较高的区域。为了检测与ZFE12蛋白相关的生物学活性，构建了两个P因子构建体，每个构建体都携带一个果蝇无女儿基因，该基因已通过用ZfE12的等效结构域替换HLH结构域或包括bHLH结构域在内的整个C末端进行了修饰。这些构建体用于转化果蝇，并测试它们拯救无女儿基因突变表型的能力。获得了神经表型和胚胎致死性的完全拯救。然而，无女儿基因的功能不能完全恢复。虽然携带编码斑马鱼HLH结构域的构建体、对于一个亚效和无效突变是反式杂合的可育果蝇出现了，但它们的头部和胸部缺少各种类型的感觉器官，并且翅膀有轻微缺陷。携带第二个构建体的突变体偶尔能发育到蛹期，但随后死亡。这些数据表明，ZFE12的HLH结构域可以执行果蝇中无女儿蛋白相应区域所执行的大部分但不是全部功能。
