Role of NF-kappaB signaling in hepatocyte growth factor/scatter factor-mediated cell protection.
The cytokine scatter factor/hepatocyte growth factor (HGF/SF) protects epithelial, carcinoma, and other cell types against cytotoxicity and apoptosis induced by DNA-damaging agents such as ionizing radiation and adriamycin (ADR, a topoisomerase IIalpha inhibitor). We investigated the role of nuclear factor kappa B (NF-kappaB) signaling in HGF/SF-mediated protection of human prostate cancer (DU-145) and Madin-Darby canine kidney (MDCK) epithelial cells against ADR. HGF/SF caused the rapid nuclear translocation of the p65 (RelA) subunit of NF-kappaB associated with the transient loss of the inhibitory subunit IkappaB-alpha. Exposure to HGF/SF caused the activation of an NF-kappaB luciferase reporter that was blocked or attenuated by the expression of a mutant 'super-repressor' IkappaB-alpha. Electrophoretic mobility shift assay supershift assays revealed that HGF/SF treatment induced the transient binding of various NF-kappaB family proteins (p65, p50, c-Rel, and RelB) with radiolabeled NF-kappaB-binding oligonucleotides. The HGF/SF-mediated protection of DU-145 and MDCK cells against ADR (demonstrated using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assays) was abrogated by the IkappaB-alpha super-repressor. The ability of HGF/SF to activate NF-kappaB signaling was dependent on c-Akt --> Pak1 (p21-associated kinase-1) signaling (with Pak1 downstream of c-Akt) and was inhibited by the tumor suppressor PTEN (phosphatase and tensin homolog). Inhibitors of phosphatidylinositol-3'-kinase and Src family kinases significantly inhibited HGF/SF-mediated activation of NF-kappaB, while inhibitors of MEK, protein kinase C, and p70 S6 kinase had a modest effect or no effect on NF-kappaB activity. HGF/SF induced the expression of several known NF-kappaB target genes (cIAP-1 (cellular inhibitor of apoptosis-1), cIAP-2, and TRAF-2 (TNF receptor-associated factor-2)) in an NF-kappaB-dependent manner; HGF/SF blocked the inhibition of expression of these genes by ADR. Experimental manipulation of expression of these genes suggests that they (particularly TRAF-2 and cIAP-2) contribute to the protection against ADR by HGF/SF. These findings suggest that HGF/SF activates NF-kappaB through a c-Akt --> Pak1 signaling pathway that is also dependent on Src, and that NF-kappaB contributes to HGF/SF-mediated protection against ADR.
Fan
Saijun
S
Department of Oncology, Lombardi Cancer Center/Georgetown University, 3970 Reservoir Road, NW, Box 571469, Washington, DC 20057-1469, USA.
Gao
Min
M
Meng
Qinghui
Q
Laterra
John J
JJ
Symons
Marc H
MH
Coniglio
Sal
S
Pestell
Richard G
RG
Goldberg
Itzhak D
ID
Rosen
Eliot M
EM
D021581:Active Transport, Cell Nucleus; D000818:Animals; D002478:Cells, Cultured; D019610:Cytoprotection; D004247:DNA / Q000378:metabolism; D004285:Dogs; D004317:Doxorubicin / Q000494:pharmacology; D017228:Hepatocyte Growth Factor / Q000494:pharmacology*; D006801:Humans; D016328:NF-kappa B / Q000502:physiology*; D051059:PTEN Phosphohydrolase; D010744:Phosphoric Monoester Hydrolases / Q000502:physiology; D017346:Protein Serine-Threonine Kinases / Q000502:physiology; D011518:Proto-Oncogene Proteins / Q000502:physiology; D051057:Proto-Oncogene Proteins c-akt; D015398:Signal Transduction; D047992:TNF Receptor-Associated Factor 2 / Q000494:pharmacology; D014158:Transcription, Genetic; D025521:Tumor Suppressor Proteins / Q000502:physiology; D054462:p21-Activated Kinases
D016328:NF-kappa B; D011518:Proto-Oncogene Proteins; D047992:TNF Receptor-Associated Factor 2; D025521:Tumor Suppressor Proteins; D017228:Hepatocyte Growth Factor; D004317:Doxorubicin; D004247:DNA; C494918:AKT1 protein, human; C515903:PAK1 protein, human; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D054462:p21-Activated Kinases; D010744:Phosphoric Monoester Hydrolases; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human