MECHANISMS OF RAF ACTIVATION

Raf-1蛋白丝氨酸/苏氨酸激酶(PSK)是一种重要的 连接生长因子受体刺激和 膜蛋白激酶C(PKC)与早期生长的激活 细胞核中的反应基因。最近我们已经证明了另一位成员 Raf家族中的B-Raf在神经生长后的PC12细胞中被激活 因素刺激。Raf-1和B-Raf激活的受体偶联 受p21Ras基因调控。细胞的生长因子刺激导致 Raf-1和Raf-1的磷酸化和伴随的激酶活性增加 B-Raf。除了调节磷酸化外，磷酸化- Raf-1激活过程中的脂类产物已经被认为是 在Raf的N-末端半部分中存在半胱氨酸指基序。 1蛋白，在其他Raf PSK家族成员和相关成员中保守 与PKC中的类似结构相同。我们现在已经扩展了对Raf-1的研究 和B-Raf到A-Raf，并能够证明(I)A-Raf的激酶激活。 RAF在相同的条件下发生，并具有类似的动力学 以前在Raf-1中看到的，(Ii)PKC-α磷酸化并激活 Raf-1，(Iii)磷脂酰胆碱的水解物 激活Raf-1，以及(Iv)我们已经确定了PSK癌基因mos和cot 作为潜在的Raf激酶。

Raf-1 protein serine/threonine kinase (PSK) functions as a critical shuttle enzyme that connects stimulation of growth factor receptors and protein kinase C (PKC) at the membrane with activation of early growth response genes in the nucleus. Recently we have shown that another member of the Raf family, B-Raf, is activated in PC12 cells after nerve growth factor stimulation. Receptor coupling of Raf-1 and B-Raf activation is controlled by p21Ras. Growth factor stimulation of cells results in the increased phosphorylation and concomitant kinase activation of Raf-1 and B-Raf. In addition to regulatory phosphorylations, a role for phospho- lipid-derived products in the activation of Raf-1 has been suggested by the presence of a cysteine finger motif in the N-terminal half of the Raf- 1 protein, which is conserved in other Raf PSK family members and related to a similar structure in PKC. We have now extended our studies on Raf-1 and B-Raf to A-Raf, and were able to show that (i) kinase activation of A- Raf occurs under identical conditions and with similar kinetics as previously seen with Raf-1, (ii) PKC-alpha phosphorylates and activates Raf-1, (iii) products derived from the hydrolysis of phosphatidylcholine activate Raf-1, and (iv) we have identified the PSK oncogenes mos and cot as potential Raf kinase kinases.