Gerwin N, La Rosée A, Sauer F, Halbritter H P, Neumann M, Jäckle H, Nauber U
Max-Planck-Institut für biophysikalische Chemie, Abteilung Molekulare Entwicklungsbiologie, Göttingen, Germany.
Mol Cell Biol. 1994 Dec;14(12):7899-908. doi: 10.1128/mcb.14.12.7899-7908.1994.
The Drosophila gap gene knirps (kni) is required for abdominal segmentation. It encodes a steroid/thyroid orphan receptor-type transcription factor which is distributed in a broad band of nuclei in the posterior region of the blastoderm. To identify essential domains of the kni protein (KNI), we cloned and sequenced the DNA encompassing the coding region of nine kni mutant alleles of different strength and kni-homologous genes of related insect species. We also examined in vitro-modified versions of KNI in various assay systems both in vitro and in tissue culture. The results show that KNI contains several functional domains which are arranged in a modular fashion. The N-terminal 185-amino-acid region which includes the DNA-binding domain and a functional nuclear location signal fails to provide kni activity to the embryo. However, a truncated KNI protein that contains additional 47 amino acids exerts rather strong kni activity which is functionally defined by a weak kni mutant phenotype of the embryo. The additional 47-amino-acid stretch includes a transcriptional repressor domain which acts in the context of a heterologous DNA-binding domain of the yeast transcriptional activator GAL4. The different domains of KNI as defined by functional studies are conserved during insect evolution.
果蝇缺口基因驼背(kni)对于腹部体节的形成是必需的。它编码一种类固醇/甲状腺孤儿受体型转录因子,该因子分布于囊胚后部区域的一条宽核带中。为了鉴定驼背蛋白(KNI)的关键结构域,我们克隆并测序了包含九个不同强度的kni突变等位基因的编码区以及相关昆虫物种的kni同源基因的DNA。我们还在各种体外和组织培养检测系统中检测了KNI的体外修饰版本。结果表明,KNI包含几个以模块化方式排列的功能结构域。包含DNA结合结构域和功能性核定位信号的N端185个氨基酸区域不能为胚胎提供kni活性。然而,包含额外47个氨基酸的截短KNI蛋白具有相当强的kni活性,其功能由胚胎的弱kni突变表型定义。额外的47个氨基酸片段包括一个转录抑制结构域,该结构域在酵母转录激活因子GAL4的异源DNA结合结构域的背景下起作用。通过功能研究定义的KNI的不同结构域在昆虫进化过程中是保守的。