Roy S, McPherson R A, Apolloni A, Yan J, Lane A, Clyde-Smith J, Hancock J F
Queensland Cancer Fund Laboratory of Experimental Oncology, Department of Pathology, University of Queensland Medical School, Brisbane 4006, Australia.
Mol Cell Biol. 1998 Jul;18(7):3947-55. doi: 10.1128/MCB.18.7.3947.
14-3-3 proteins complex with many signaling molecules, including the Raf-1 kinase. However, the role of 14-3-3 in regulating Raf-1 activity is unclear. We show here that 14-3-3 is bound to Raf-1 in the cytosol but is totally displaced when Raf-1 is recruited to the plasma membrane by oncogenic mutant Ras, in vitro and in vivo. 14-3-3 is also displaced when Raf-1 is targeted to the plasma membrane. When serum-starved cells are stimulated with epidermal growth factor, some recruitment of 14-3-3 to the plasma membrane is evident, but 14-3-3 recruitment correlates with Raf-1 dissociation and inactivation, not with Raf-1 recruitment. In vivo, overexpression of 14-3-3 potentiates the specific activity of membrane-recruited Raf-1 without stably associating with the plasma membrane. In vitro, Raf-1 must be complexed with 14-3-3 for efficient recruitment and activation by oncogenic Ras. Recombinant 14-3-3 facilitates Raf-1 activation by membranes containing oncogenic Ras but reduces the amount of Raf-1 that associates with the membranes. These data demonstrate that the interaction of 14-3-3 with Raf-1 is permissive for recruitment and activation by Ras, that 14-3-3 is displaced upon membrane recruitment, and that 14-3-3 may recycle Raf-1 to the cytosol. A model that rationalizes many of the apparently discrepant observations on the role of 14-3-3 in Raf-1 activation is proposed.
14-3-3蛋白可与许多信号分子形成复合物，包括Raf-1激酶。然而，14-3-3在调节Raf-1活性中的作用尚不清楚。我们在此表明，在体外和体内，14-3-3在细胞质中与Raf-1结合，但当致癌突变型Ras将Raf-1招募到质膜时，14-3-3会完全解离。当Raf-1靶向质膜时，14-3-3也会解离。当用表皮生长因子刺激血清饥饿的细胞时，可明显观察到一些14-3-3被招募到质膜，但14-3-3的招募与Raf-1的解离和失活相关，而与Raf-1的招募无关。在体内，14-3-3的过表达增强了膜招募的Raf-1的比活性，但并不与质膜稳定结合。在体外，Raf-1必须与14-3-3形成复合物才能被致癌性Ras有效招募和激活。重组14-3-3可促进含致癌性Ras的膜对Raf-1的激活，但会减少与膜结合的Raf-1的量。这些数据表明，14-3-3与Raf-1的相互作用有利于Ras的招募和激活，14-3-3在膜招募时会被取代，并且14-3-3可能会将Raf-1循环回细胞质。本文提出了一个模型，该模型解释了许多关于14-3-3在Raf-1激活中作用的明显矛盾的观察结果。
