Downregulation of the c-Fes protein-tyrosine kinase inhibits the proliferation of human renal carcinoma cells.
The c-Fes protein-tyrosine kinase is associated with growth and differentiation of hematopoietic, neuronal, vascular endothelial and epithelial cell types. In this study, we investigated whether small interfering RNA (siRNA)-mediated knockdown of c-Fes expression affected proliferation of the human renal carcinoma cell lines, ACHN and VMRC-RCW. Immunofluorescence microscopy showed that c-Fes was expressed in both the cytosol and nuclei of these cells, and siRNA treatment preferentially downregulated c-Fes expression in the cytosol. Knock-down of c-Fes inhibited cellular proliferation in a dose-dependent manner with minimal increase in cell death. c-Fes siRNA treatment also downregulated the phosphorylation of Akt1 on S473 and IKKalpha on T23, and cyclin D1 expression, enhanced the expression of IkappaBalpha, and prevented the nuclear localization of NFkappaB. Treatment with an NFkappaB inhibitory peptide (SN50) also blocked the proliferation and nuclear localization of NFkappaB in these cells. The effect of SN50 treatment was not enhanced by c-Fes siRNA, suggesting that downregulation of c-Fes expression inhibited cell cycle progression through the Akt1/NFkappaB pathway. In contrast to siRNA-mediated knockdown, ectopic expression of either wild-type or kinase-inactive c-Fes in renal carcinoma cells failed to alter their proliferation in vitro and in vivo. Thus, suppression of proliferation resulting from siRNA-mediated knockdown may depend upon an expression of c-Fes protein rather than its kinase activity. Taken together, our results indicate that downregulation of c-Fes expression may be a potential therapeutic strategy for advanced human renal cell carcinoma and inhibition of its kinase activity as an antiangiogenic therapy does not seem to induce the growth of human renal carcinoma cells.
Kanda
Shigeru
S
Department of Molecular Microbiology and Immunology, Division of Endothelial Cell Biology, Nagasaki University Graduate School of Biomedical Sciences, 1-7-1, Sakamoto, Nagasaki 851-0251, Japan. skanda-jua@umin.net
Miyata
Yasuyoshi
Y
Kanetake
Hiroshi
H
Smithgall
Thomas E
TE
D000818:Animals; D002292:Carcinoma, Renal Cell / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D002467:Cell Nucleus; D049109:Cell Proliferation; D015536:Down-Regulation; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D020672:I-kappa B Proteins / Q000378:metabolism; D015151:Immunoblotting; D007124:Immunoenzyme Techniques; D047468:Immunoprecipitation; D007680:Kidney Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D051578:Proto-Oncogene Proteins c-fes / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000494:pharmacology; D015398:Signal Transduction; D014407:Tumor Cells, Cultured
D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; C000606205:Nfkbia protein, mouse; D034741:RNA, Small Interfering; D000072000:NF-KappaB Inhibitor alpha; D011494:Protein Kinases; C496645:FES protein, human; D051578:Proto-Oncogene Proteins c-fes; D051057:Proto-Oncogene Proteins c-akt