Trzeciak W H, Simpson E R, Scallen T J, Vahouny G V, Waterman M R
J Biol Chem. 1987 Mar 15;262(8):3713-7.
The effects of ACTH or dibutyryl cyclic AMP (Bt2cAMP) on the synthesis of sterol carrier protein-2 (SCP2) have been studied in rat adrenocortical cells in monolayer culture. Radiolabeling of total cellular proteins with [35S]methionine and immunoprecipitation with antibodies directed against rat liver SCP2, followed by polyacrylamide gel electrophoresis and fluorography, showed a 3-4-fold increase in the rate of synthesis of SCP2 in cells treated for 48 h with ACTH (1 microM) or Bt2cAMP (0.1 mM). The induction of SCP2 synthesis depended upon the concentrations of ACTH or Bt2cAMP with an ED50 of 8 and 100 nM, respectively, and increased linearly with time between 12 and 48 h of treatment. Immunoprecipitation of SCP2 synthesized in a rabbit reticulocyte in vitro translation system programmed with RNA isolated from cells treated with ACTH or Bt2cAMP revealed increased synthesis of SCP2 compared to RNA from control cells. The immunoprecipitable rat adrenal SCP2, synthesized in a cell-free translation system, showed mobility corresponding to Mr of 14,400 upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis and was clearly larger than immunodetectable SCP2 synthesized in cultured adrenal cells (Mr = 11,300). The electrophoretic mobilities of rat liver SCP2 synthesized in cultured cells and in a cell-free translation system were the same as the respective forms from rat adrenal. It is concluded that the synthesis of SCP2 in rat adrenocortical cells is induced by ACTH and that the induction is mediated by cAMP and may involve increased levels of translatable mRNA encoding a higher molecular weight precursor form of SCP2, which presumably undergoes post-translational processing yielding the mature form.
在单层培养的大鼠肾上腺皮质细胞中，研究了促肾上腺皮质激素（ACTH）或二丁酰环磷腺苷（Bt2cAMP）对固醇载体蛋白-2（SCP2）合成的影响。用[35S]甲硫氨酸对总细胞蛋白进行放射性标记，并用针对大鼠肝脏SCP2的抗体进行免疫沉淀，随后进行聚丙烯酰胺凝胶电泳和荧光自显影，结果显示，用ACTH（1 microM）或Bt2cAMP（0.1 mM）处理48小时的细胞中，SCP2的合成速率增加了3至4倍。SCP2合成的诱导取决于ACTH或Bt2cAMP的浓度，其半数有效浓度（ED50）分别为8和100 nM，并且在处理12至48小时之间随时间呈线性增加。用从经ACTH或Bt2cAMP处理的细胞中分离的RNA编程的兔网织红细胞体外翻译系统中合成的SCP2进行免疫沉淀，与对照细胞的RNA相比，显示SCP2的合成增加。在无细胞翻译系统中合成的可免疫沉淀的大鼠肾上腺SCP2，在十二烷基硫酸钠-聚丙烯酰胺凝胶电泳上显示出与分子量14,400相对应的迁移率，明显大于在培养的肾上腺细胞中合成的可免疫检测的SCP2（分子量 = 11,300）。在培养细胞和无细胞翻译系统中合成的大鼠肝脏SCP2的电泳迁移率与来自大鼠肾上腺的相应形式相同。结论是，ACTH诱导大鼠肾上腺皮质细胞中SCP2的合成，并且该诱导由cAMP介导，可能涉及编码SCP2较高分子量前体形式的可翻译mRNA水平的增加，推测该前体形式经过翻译后加工产生成熟形式。
