Temesvari L A, Seastone D J, Cardelli J A
Department of Microbiology and Immunology, Louisiana State University Medical Center, Shreveport 71130, USA.
Gene. 1996 Dec 12;183(1-2):47-51. doi: 10.1016/s0378-1119(96)00444-1.
We describe the cloning and characterization of a cDNA, DdApm1, encoding a putative medium chain subunit of a clathrin-associated protein (adaptor or assembly protein [AP]) complex in Dictyostelium discoideum. The DdApm1 clone is predicted to encode a polypeptide of 439 amino acids (aa) with a molecular mass of 49.9 kDa. The predicted translation product (DdApm1p) shares at least 51.7% identity and 76.3% similarity with the medium chain subunits of plasma membrane (mb)-associated clathrin AP complexes from rat and Caenorhabditis elegans. The deduced aa sequence also demonstrates significant but lesser homology to a number of medium chain subunits of Golgi-associated clathrin AP complexes. Since DdApm1p demonstrates significantly greater homology to plasma mb-associated clathrin AP complex medium chains than to their Golgi-associated counterparts, we suggest that DdApm1p may be a medium chain subunit of an AP complex involved in clathrin function at the plasma mb of D. discoideum. Southern blot analysis indicated that DdApm1 gene defines a single copy gene in the D. discoideum genome. Northern blot analysis of RNA purified at different times during growth and development demonstrated that the DdApm1 gene is expressed at relatively constant levels throughout the life cycle of the organism. DdApm1 is the first reported full-length cDNA encoding a subunit of an AP complex in D. discoideum, and thus provides the first evidence for the existence of AP complexes in this organism.
我们描述了一种cDNA（DdApm1）的克隆和特性分析，该cDNA编码盘基网柄菌中一种假定的网格蛋白相关蛋白（衔接蛋白或组装蛋白[AP]）复合物的中链亚基。DdApm1克隆预计编码一个由439个氨基酸（aa）组成的多肽，分子量为49.9 kDa。预测的翻译产物（DdApm1p）与大鼠和秀丽隐杆线虫质膜（mb）相关的网格蛋白AP复合物的中链亚基具有至少51.7%的同一性和76.3%的相似性。推导的氨基酸序列与一些高尔基体相关的网格蛋白AP复合物的中链亚基也显示出显著但较低的同源性。由于DdApm1p与质膜相关的网格蛋白AP复合物中链的同源性明显高于其与高尔基体相关对应物的同源性，我们认为DdApm1p可能是参与盘基网柄菌质膜网格蛋白功能的AP复合物的中链亚基。Southern印迹分析表明，DdApm1基因在盘基网柄菌基因组中定义了一个单拷贝基因。对生长和发育过程中不同时间纯化的RNA进行Northern印迹分析表明，DdApm1基因在生物体的整个生命周期中以相对恒定的水平表达。DdApm1是首次报道的编码盘基网柄菌中AP复合物亚基的全长cDNA，因此为该生物体中AP复合物的存在提供了首个证据。
