Effect of inositol hexakisphosphate kinase 2 on transforming growth factor beta-activated kinase 1 and NF-kappaB activation.
We previously showed that inositol hexakisphosphate kinase 2 (IHPK2) functions as a growth-suppressive and apoptosis-enhancing kinase during cell stress. Overexpression of IHPK2 sensitized ovarian carcinoma cell lines to the growth-suppressive and apoptotic effects of interferon beta (IFN-beta), IFN-alpha2, and gamma-irradiation. Expression of a kinase-dead mutant abrogated 50% of the apoptosis induced by IFN-beta. Because the kinase-dead mutant retained significant response to cell stressors, we hypothesized that a portion of the death-promoting function of IHPK2 was independent of its kinase activity. We now demonstrate that IHPK2 binds to tumor necrosis factor (TNF) receptor-associated factor (TRAF) 2 and interferes with phosphorylation of transforming growth factor beta-activated kinase 1 (TAK1), thereby inhibiting NF-kappaB signaling. IHPK2 contains two sites required for TRAF2 binding, Ser-347 and Ser-359. Compared with wild type IHPK2-transfected cells, cells expressing S347A and S359A mutations displayed 3.5-fold greater TAK1 activation following TNF-alpha. This mutant demonstrated a 6-10-fold increase in NF-kappaB DNA binding following TNF-alpha compared with wild type IHPK2-expressing cells in which NF-kappaB DNA binding was inhibited. Cells transfected with wild type IHPK2 or IHPK2 mutants that lacked S347A and S359A mutations displayed enhanced terminal deoxynucleotidyltransferase-mediated dUTP nick end-labeling staining following TNF-alpha. We believe that IHPK2-TRAF2 binding leads to attenuation of TAK1- and NF-kappaB-mediated signaling and is partially responsible for the apoptotic activity of IHPK2.
Morrison
Bei H
BH
Center for Hematology and Oncology Molecular Therapeutics, Taussig Cancer Center, and Department of Cancer Biology, Lerner Research Institute, Cleveland Clinic, Cleveland, Ohio 44195, USA.
Bauer
Joseph A
JA
Lupica
Joseph A
JA
Tang
Zhuo
Z
Szugye
Heidi
H
DiDonato
Joseph A
JA
Lindner
Daniel J
DJ
D019943:Amino Acid Substitution; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics / Q000528:radiation effects; D045744:Cell Line, Tumor; D005720:Gamma Rays; D015870:Gene Expression; D006801:Humans; D016898:Interferon-alpha / Q000494:pharmacology; D016899:Interferon-beta / Q000494:pharmacology; D020930:MAP Kinase Kinase Kinases / Q000235:genetics / Q000378:metabolism*; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000235:genetics / Q000528:radiation effects; D051379:Mice; D020125:Mutation, Missense; D016328:NF-kappa B / Q000378:metabolism; D017856:Phosphotransferases (Phosphate Group Acceptor) / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding / Q000235:genetics; D047992:TNF Receptor-Associated Factor 2 / Q000235:genetics / Q000378:metabolism*; D000099151:MAP Kinase Kinase Kinase 7
D016898:Interferon-alpha; D016899:Interferon-beta; D020930:MAP Kinase Kinase Kinases; D016328:NF-kappa B; D017856:Phosphotransferases (Phosphate Group Acceptor); D047992:TNF Receptor-Associated Factor 2; D000099151:MAP Kinase Kinase Kinase 7; C099438:inositol hexakisphosphate kinase