Department of Medicine, New York University School of Medicine, New York, NY 10016, USA.
Mol Cell Biol. 2010 Dec;30(24):5649-57. doi: 10.1128/MCB.00635-10. Epub 2010 Oct 11.
Ras proteins associate with cellular membranes as a consequence of a series of posttranslational modifications of a C-terminal CAAX sequence that include prenylation and are thought to be required for biological activity. In Drosophila melanogaster, Ras1 is required for eye development. We found that Drosophila Ras1 is inefficiently prenylated as a consequence of a lysine in the A(1) position of its CAAX sequence such that a significant pool remains soluble in the cytosol. We used mosaic analysis with a repressible cell marker (MARCM) to assess if various Ras1 transgenes could restore photoreceptor fate to eye disc cells that are null for Ras1. Surprisingly, we found that whereas Ras1 with an enhanced efficiency of membrane targeting could not rescue the Ras1 null phenotype, Ras1 that was not at all membrane targeted by virtue of a mutation of the CAAX cysteine was able to fully rescue eye development. In addition, constitutively active Ras1(12V,C186S) not targeted to membranes produced a hypermorphic phenotype and stimulated mitogen-activated protein kinase (MAPK) signaling in S2 cells. We conclude that the membrane association of Drosophila Ras1 is not required for eye development.
Ras 蛋白通过一系列 C 末端 CAAX 序列的翻译后修饰（包括 prenylation）与细胞膜结合，这些修饰被认为是生物活性所必需的。在果蝇中，Ras1 对于眼睛发育是必需的。我们发现，由于其 CAAX 序列的 A(1)位的赖氨酸，果蝇 Ras1 的 prenylation 效率很低，以至于很大一部分仍然存在于细胞质中可溶。我们使用可抑制细胞标记物的镶嵌分析（MARCM）来评估各种 Ras1 转基因是否可以将感光器命运恢复到 Ras1 缺失的眼盘细胞中。令人惊讶的是，我们发现，虽然靶向膜效率更高的 Ras1 不能挽救 Ras1 缺失表型，但由于 CAAX 半胱氨酸突变而根本不靶向膜的 Ras1 能够完全挽救眼睛发育。此外，不靶向膜的组成性激活 Ras1(12V,C186S)产生了超形表型，并刺激 S2 细胞中的丝裂原活化蛋白激酶（MAPK）信号转导。我们得出结论，果蝇 Ras1 的膜结合对于眼睛发育不是必需的。
