A dimeric 14-3-3 protein is an essential cofactor for Raf kinase activity

被引:392
作者
Tzivion, G
Luo, ZJ
Avruch, J [1 ]
机构
[1] Massachusetts Gen Hosp, Diabet Unit, Boston, MA 02114 USA
[2] Massachusetts Gen Hosp, Med Serv, Boston, MA 02114 USA
[3] Massachusetts Gen Hosp, Dept Biol Mol, Boston, MA 02114 USA
[4] Harvard Univ, Sch Med, Dept Med, Boston, MA 02114 USA
关键词
D O I
10.1038/27938
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
cRaf-1 is a mitogen-activated protein kinase that is the main effector recruited by GTP-bound Ras in order to activate the MAP kinase pathway(1). Inactive Raf is found in the cytosol in a complex with Hsp90, Hsp50 (Cdc37)(2,3) and the 14-3-3 proteins(4). GTP-bound Ras binds Raf and is necessary but not sufficient for the stable activation of Raf that occurs in response to serum, epidermal growth factor, platelet-derived growth factor or insulin(5-8). These agents cause a two- to threefold increase in overall phosphorylation of Raf on serine/threonine residues(8,9), and treatment of cRaf-1 with protein (serine/threonine) phosphatases can deactivate it, at least partially(10). The role of 14-3-3 proteins in the regulation of Raf's kinase activity is uncertain(4,11) and is investigated here. Active Raf can be almost completely deactivated in vitro by displacement of 14-3-3 using synthetic phosphopeptides. Deactivation can be substantially reversed by addition of purified recombinant bacterial 14-3-3; however, Raf must have been previously activated in vivo to be reactivated by 14-3-3 in vitro. The ability of 14-3-3 to support Raf activity is dependent on phosphorylation of serine residues on Raf and on the integrity of the 14-3-3 dimer; mutant monomeric forms of 14-3-3, although able to bind Raf in vivo, do not enable Raf to be activated in vivo or restore Raf activity after displacement of 14-3-3 in vitro. The 14-3-3 protein is not required to induce dimerization of Raf, We propose that dimeric 14-3-3 is needed both to maintain Raf in an inactive state in the absence of GTP-bound Ras and to stabilize an active conformation of Raf produced during activation in vivo.
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页码:88 / 92
页数:5
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