Rincón-Limas D E, Lu C H, Canal I, Botas J
Department of Molecular and Human Genetics, Baylor College of Medicine, One Baylor Plaza, Houston, TX 77030, USA.
EMBO J. 2000 Jun 1;19(11):2602-14. doi: 10.1093/emboj/19.11.2602.
The LIM homeodomain (LIM-HD) protein Apterous (Ap) and its cofactor DLDB/CHIP control dorso- ventral (D/V) patterning and growth of Drosophila wing. To investigate the molecular mechanisms of Ap/CHIP function we altered their relative levels of expression and generated mutants in the LIM1, LIM2 and HD domains of Ap, as well as in the LIM-interacting and self-association domains of CHIP. Using in vitro and in vivo assays we found that: (i) the levels of CHIP relative to Ap control D/V patterning; (ii) the LIM1 and LIM2 domains differ in their contributions to Ap function; (iii) Ap HD mutations cause weak dominant negative effects; (iv) overexpression of ChipDeltaSAD mutants mimics Ap lack-of-function, and this dominant negative phenotype is caused by titration of Ap because it can be rescued by adding extra Ap; and (v) overexpression of ChipDeltaLID mutants also causes an Ap lack-of-function phenotype, but it cannot be rescued by extra Ap. These results support the model that the Ap-CHIP active complex in vivo is a tetramer.
LIM 同源结构域（LIM-HD）蛋白无翅（Ap）及其辅因子 DLDB/CHIP 控制果蝇翅膀的背腹（D/V）模式形成和生长。为了研究 Ap/CHIP 功能的分子机制，我们改变了它们的相对表达水平，并在 Ap 的 LIM1、LIM2 和 HD 结构域以及 CHIP 的 LIM 相互作用和自缔合结构域中产生了突变体。通过体外和体内试验，我们发现：（i）CHIP 相对于 Ap 的水平控制 D/V 模式形成；（ii）LIM1 和 LIM2 结构域对 Ap 功能的贡献不同；（iii）Ap HD 突变产生微弱的显性负效应；（iv）ChipDeltaSAD 突变体的过表达模拟了 Ap 功能缺失，这种显性负表型是由 Ap 的滴定引起的，因为添加额外的 Ap 可以挽救它；（v）ChipDeltaLID 突变体的过表达也导致 Ap 功能缺失表型，但额外的 Ap 不能挽救它。这些结果支持了体内 Ap-CHIP 活性复合物是四聚体的模型。
