Mixed lineage kinase 3 negatively regulates IKK activity and enhances etoposide-induced cell death.
Mixed lineage kinase 3 (MLK3) is a mitogen activated protein kinase kinase kinase (MAP3K) that activates multiple MAPK signaling pathways. Nuclear factor kappa B (NF-kappaB) is a transcription factor that has important functions in inflammation, immunity and cell survival. We found that silencing mlk3 expression with RNA interference (RNAi) in SKOV3 human ovarian cancer epithelial cells and NIH-3T3 murine fibroblasts led to a reduction in the level of the inhibitor of kappa B alpha (IkappaBalpha) protein. In addition, we observed enhanced basal IkappaB kinase (IKK) activity in HEK293 cells transiently transfected with MLK3 siRNA and in NIH3T3 cells stably expressing MLK3 shRNA (shMLK3). Furthermore, the basal level of NF-kappaB-dependent gene transcription was elevated in shMLK3 cells. Silencing mlk3 expression conferred resistance of cells to etoposide-induced apoptotic cell death and overexpression of wild type MLK3 (MLK3-WT) or kinase-dead MLK3 (MLK3-KD) promoted apoptotic cell death and cleavage of poly (ADP-ribose) polymerase (PARP). Overexpression of MLK3-WT or MLK3-KD enhanced etoposide-induced apoptotic cell death and cleavage of PARP. These data suggest that MLK3 functions to limit IKK activity, and depleting MLK3 helps protect cells from etoposide-induced cell death through activation of IKK-dependent signaling.
Department of Biological Sciences, University of Toledo, 2801 West Bancroft Street, Toledo, OH 43606, USA.
Eric T
ET
Cole
Yu
Y
Zhan
Widian F
WF
Abi Saab
Amanda C
AC
Korchnak
Brian P
BP
Ashburner
Deborah N
DN
Chadee
D000818:Animals; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects* / Q000235:genetics; D045744:Cell Line, Tumor; D005047:Etoposide / Q000494:pharmacology*; D006801:Humans; D051550:I-kappa B Kinase / Q000235:genetics / Q000378:metabolism*; D020672:I-kappa B Proteins / Q000235:genetics / Q000378:metabolism; D020930:MAP Kinase Kinase Kinases / Q000235:genetics / Q000378:metabolism*; D020935:MAP Kinase Signaling System / Q000187:drug effects* / Q000235:genetics; D051379:Mice; D041681:NIH 3T3 Cells; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases / Q000235:genetics / Q000378:metabolism; D000097767:Mitogen-Activated Protein Kinase Kinase Kinase 11
D000972:Antineoplastic Agents, Phytogenic; D005047:Etoposide; D051550:I-kappa B Kinase; D020672:I-kappa B Proteins; D020930:MAP Kinase Kinase Kinases; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; D000097767:Mitogen-Activated Protein Kinase Kinase Kinase 11; C499320:PARP1 protein, human; C499321:Parp1 protein, mouse