Panganiban G E, Rashka K E, Neitzel M D, Hoffmann F M
McArdle Laboratory for Cancer Research, University of Wisconsin, Madison 53706.
Mol Cell Biol. 1990 Jun;10(6):2669-77. doi: 10.1128/mcb.10.6.2669-2677.1990.
The decapentaplegic (dpp) gene of Drosophila melanogaster is required for pattern formation in the embryo and for viability of the epithelial cells in the imaginal disks. The dpp protein product predicted from the DNA sequence is similar to members of a family of growth factors that includes transforming growth factor beta (TGF-beta). We have produced polyclonal antibodies to a recombinant dpp protein made in bacteria and used a metallothionein promoter to express a dpp cDNA in Drosophila S2 cells. Similar to other proteins in the TGF-beta family, the dpp protein produced by the Drosophila cells was proteolytically cleaved, and both portions of the protein were secreted from the cells. The amino-terminal 47-kilodalton (kDa) peptide was found in the medium and in the proteins adhering to the plastic petri dish. The carboxy-terminal peptide, the region with sequence similarity to the active ligand portion of TGF-beta, was found extracellularly as a 30-kDa homodimer. Most of the 30-kDa homodimer was in the S2 cell protein adsorbed onto the surface of the plastic dish. The dpp protein could be released into solution by increased salt concentration and nonionic detergent. Under these conditions, the amino-terminal and carboxy-terminal portions of dpp were not associated in a stable complex.
果蝇的截瘫（dpp）基因对于胚胎中的模式形成以及成虫盘上皮细胞的存活是必需的。从DNA序列预测的dpp蛋白产物类似于包括转化生长因子β（TGF-β）在内的生长因子家族成员。我们已经制备了针对细菌中产生的重组dpp蛋白的多克隆抗体，并使用金属硫蛋白启动子在果蝇S2细胞中表达dpp cDNA。与TGF-β家族中的其他蛋白质类似，果蝇细胞产生的dpp蛋白被蛋白水解切割，并且该蛋白的两个部分都从细胞中分泌出来。在培养基和附着在塑料培养皿上的蛋白质中发现了氨基末端47千道尔顿（kDa）的肽。羧基末端肽，即与TGF-β活性配体部分具有序列相似性的区域，在细胞外作为30-kDa同二聚体被发现。大部分30-kDa同二聚体存在于吸附在塑料培养皿表面的S2细胞蛋白中。通过增加盐浓度和非离子去污剂，dpp蛋白可以释放到溶液中。在这些条件下，dpp的氨基末端和羧基末端部分不会形成稳定的复合物。
