THE protein Raf-1, a key mediator of mitogenesis and differentiation, associates with p21(ras) (refs 1-3). However, the regulation of the serine/threonine kinase activity of Raf-1 is still not understood(4-13). Using the yeast two-hybrid systems(8,14-16), we identified two structurally related proteins that interact with the aminoterminal region of Raf-1. These proteins, 14-3-3 zeta (PLA(2)) and 14-3-3 beta (HS1), are members of the 14-3-3 family of proteins(17-23). Expression of 14-3-3 proteins in Xenopus oocytes enhanced Raf-1 activity and promoted Raf-1-dependent oocyte maturation. A dominant negative mutant of Raf-1 blocked tile effects of 14-3-3 protein.