Cytokine-induced activation of mixed lineage kinase 3 requires TRAF2 and TRAF6.
Mixed lineage kinase 3 (MLK3) is a mitogen-activated protein kinase kinase kinase (MAP3K) that activates multiple mitogen-activated protein kinase (MAPK) pathways in response to growth factors, stresses and the pro-inflammatory cytokine, tumor necrosis factor (TNF). MLK3 is required for optimal activation of stress activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK) signaling by TNF, however, the mechanism by which MLK3 is recruited and activated by the TNF receptor remains poorly understood. Here we report that both TNF and interleukin-1 beta (IL-1 beta) stimulation rapidly activate MLK3 kinase activity. We observed that TNF stimulates an interaction between MLK3 and TNF receptor associated factor (TRAF) 2 and IL-1 beta stimulates an interaction between MLK3 and TRAF6. RNA interference (RNAi) of traf2 or traf6 dramatically impairs MLK3 activation by TNF indicating that TRAF2 and TRAF6 are critically required for MLK3 activation. We show that TNF also stimulates ubiquitination of MLK3 and MLK3 can be conjugated with lysine 48 (K48)- and lysine 63 (K63)-linked polyubiquitin chains. Our results suggest that K48-linked ubiquitination directs MLK3 for proteosomal degradation while K63-linked ubiquitination is important for MLK3 kinase activity. These results reveal a novel mechanism for MLK3 activation by the pro-inflammatory cytokines TNF and IL-1 beta.
Korchnak
Amanda C
AC
Department of Biological Sciences, University of Toledo, 2801 West Bancroft Street, MS601, Toledo, OH 43606, USA.
Zhan
Yu
Y
Aguilar
Michael T
MT
Chadee
Deborah N
DN
D002460:Cell Line; D006801:Humans; D053583:Interleukin-1beta / Q000494:pharmacology*; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D020930:MAP Kinase Kinase Kinases / Q000378:metabolism*; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D015398:Signal Transduction; D047992:TNF Receptor-Associated Factor 2 / Q000235:genetics / Q000378:metabolism*; D048029:TNF Receptor-Associated Factor 6 / Q000235:genetics / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D054875:Ubiquitination; D000097767:Mitogen-Activated Protein Kinase Kinase Kinase 11
D053583:Interleukin-1beta; D048031:JNK Mitogen-Activated Protein Kinases; D020930:MAP Kinase Kinase Kinases; D034741:RNA, Small Interfering; D047992:TNF Receptor-Associated Factor 2; D048029:TNF Receptor-Associated Factor 6; D014409:Tumor Necrosis Factor-alpha; D000097767:Mitogen-Activated Protein Kinase Kinase Kinase 11