MLK-3 ACTIVATES THE SAPK JNK AND P38/RK PATHWAYS VIA SEK1 AND MKK3/6/

Mixed lineage kinase-3 (MLK-3) is a 97 kDa serine/threonine kinase wit h multiple interaction domains, including a Cdc42 binding motif, but u nknown function. Cdc42 and the related small GTP binding protein Rac1 can activate the SAPK/JNK and p38/RK stress-responsive kinase cascades , suggesting that MLK-3 may have a role in upstream regulation of thes e pathways . In support of this role, we demonstrate that MLK-3 can sp ecifically activate the SAPK/JNK and p38/RK pathways, but has no effec t on the activation of ERKs. Immunoprecipitated MLK-3 catalyzed the ph osphorylation of SEK1 in vitro, and co-transfected MLK-3 induced phosp horylation of SEK1 and MKK3 at sites required for activation, suggesti ng direct regulation of these protein kinases. Furthermore, interactio ns between MLK-3 and SEK and MLK-3 and MKK6 were observed in co-precip itation experiments. Finally, kinase-dead mutants of MLK-3 blocked act ivation of the SAPK pathway by a newly identified mammalian analog of Ste20, germinal center kinase, but not by MEKK, suggesting that MLK-3 functions to activate the SAPK/JNK and p38/RK cascades in response to stimuli transduced by Ste20-like kinases.