Dumaz Nicolas, Marais Richard
Cancer Research UK Centre for Cell and Molecular Biology, Institute of Cancer Research, 237 Fulham Road, London SW3 6JB, United Kingdom.
J Biol Chem. 2003 Aug 8;278(32):29819-23. doi: 10.1074/jbc.C300182200. Epub 2003 Jun 11.
Cyclic AMP (cAMP) blocks Raf-1 activation by stimulating its phosphorylation on serine 43 (Ser43), serine 233 (Ser233), and serine 259 (Ser259). We show here that phosphorylation of all three sites blocks Raf-1 binding to Ras.GTP in vivo and that cAMP stimulates binding of 14-3-3 proteins to Ser233 and Ser259. We also show that Raf-1 and protein kinase A (PKA) form a complex in vivo that is disrupted by cAMP and that ablation of PKA by use of small interfering RNA blocks phosphorylation by cAMP. The ability of PKA to block Raf-1 activation is ablated by the PKA inhibitor H89. These studies suggest that Raf-1 and cAMP form a signaling complex in cells. Upon activation of PKA, Raf-1 is phosphorylated and 14-3-3 binds, blocking Raf-1 recruitment to the plasma membrane and preventing its activation.
环磷酸腺苷（cAMP）通过刺激丝氨酸43（Ser43）、丝氨酸233（Ser233）和丝氨酸259（Ser259）的磷酸化来阻断Raf-1的激活。我们在此表明，这三个位点的磷酸化在体内均会阻断Raf-1与Ras.GTP的结合，且cAMP会刺激14-3-3蛋白与Ser233和Ser259的结合。我们还表明，Raf-1与蛋白激酶A（PKA）在体内形成复合物，该复合物会被cAMP破坏，并且使用小干扰RNA消除PKA会阻断cAMP的磷酸化作用。PKA抑制剂H89消除了PKA阻断Raf-1激活的能力。这些研究表明，Raf-1与cAMP在细胞中形成信号复合物。PKA激活后，Raf-1发生磷酸化，14-3-3与之结合，从而阻断Raf-1募集至质膜并阻止其激活。
