3PK, A NOVEL MITOGEN-ACTIVATED PROTEIN (MAP) KINASE-ACTIVATED PROTEIN-KINASE, IS TARGETED BY 3 MAP KINASE PATHWAYS

Recently we have identified a mitogen-activated protein kinase (MAPK)- activated protein kinase, named 3pK (G. Sithanandam, F. Latif, U. Smol a, R. A. Bernal, F.-M. Duh, H. Li, I. Kuzmin, V. Wirier, L. Geil, S. S hresta, P. A. Lloyd, S. Bader, Y. Sekido, K. D. Tartof, V. I. Kashuba, E. R. Zabarovsky, M. Dean, G. Klein, B. Zbar, M. I. Lerman, J. D. Min na, U. R. Rapp, and A. Allikmets, Mel. Cell. Biol. 16:868-876, 1996). In vitro characterization of the kinase revealed that 3pK is activated by ERK. It was further shown that 3pK is phosphorylated in vivo after stimulation of cells with serum. However, the in vivo relevance of th is observation in terms of involvement of the Raf/MEK/ERK cascade has not been established. Here we show that 3pK is activated in vivo by th e growth inducers serum and tetradecanoyl phorbol acetate in promyeloc ytic HL60 cells and transiently transfected embryonic kidney 293 cells . Activation of 3pK was Raf dependent and was mediated by the Raf/MEK/ ERK kinase cascade. 3pK was also shown to be activated after stress st imulation of cells. In vitro studies with recombinant proteins demonst rate that in addition to ERK, members of other subgroups of the MARK f amily, namely, p38RK and Jun-N-terminal kinases/stress-activated prote in kinases, were also able to phosphorylate and activate 3pK. Cotransf ection experiments as well as the use of a specific inhibitor of p38RK showed that these in vitro upstream activators also function In vivo, identifying 3pK as the first kinase to be activated through all three MAPK cascades. Thus, 3pK is a novel convergence point of different MA PK pathways and could function as an integrative element of signaling in both mitogen and stress responses.