NUB1, an interferon-inducible protein, mediates anti-proliferative actions and apoptosis in renal cell carcinoma cells through cell-cycle regulation.
BACKGROUND
NEDD8 ultimate buster 1 (NUB1) is an interferon (IFN)-inducible protein that downregulates NEDD8 expression and its conjugation system. Although overexpression of NUB1 induces a growth-inhibitory effect in cells, the mechanisms underlying the anti-mitogenic actions of NUB1 in cancer cells remain uncertain. We investigated the anti-cancer effects of NUB1 in human renal cell carcinoma (RCC) cells.


METHODS
Nine human RCC cells were used for this study. The proliferation of RCC cells exposed to IFN-alpha was measured by water-soluble tetrazolium salt assay. The expression level of NUB1 in cells was measured by quantitative reverse transcriptase PCR or western blot analysis. Apoptosis and cell-cycle analysis were performed by flow cytometry. Silencing of NUB1 was performed using a small interfering RNA.


RESULTS
Both NUB1 messenger RNA and protein were significantly induced by IFN-alpha in seven out of nine selected RCC cell lines, and the NUB1 expressions induced by IFN-alpha correlated positively with cell growth inhibition. Overexpression of NUB1 remarkably induced S-phase transition during cell cycle and apoptosis in IFN-alpha-resistant A498 cells, in which NUB1 is not induced by IFN-alpha. The expression levels of two cell-cycle regulator proteins, cyclin E and p27, were increased under the aforementioned conditions. The knockdown of NUB1 enhanced cell proliferation of IFN-alpha-resistant A498 cells and suppressed IFN-alpha-induced growth inhibition in IFN-alpha-sensitive 4TUHR cells.


CONCLUSIONS
NUB1 may be a key factor involved not only in cell growth inhibition by IFN-alpha in RCC cells but also in the anti-cancer effect against IFN-alpha-resistant RCC cells.
Hosono
T
T
Department of Urology, Osaka City University Graduate School of Medicine, 1-4-3, Asahimachi, Abeno-ku, Osaka, Japan.
Tanaka
T
T
Tanji
K
K
Nakatani
T
T
Kamitani
T
T
D048868:Adaptor Proteins, Signal Transducing; D017209:Apoptosis; D015153:Blotting, Western; D002292:Carcinoma, Renal Cell / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D019927:Cyclin E / Q000235:genetics / Q000378:metabolism; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000235:genetics / Q000378:metabolism; D006801:Humans; D016898:Interferon-alpha / Q000494:pharmacology*; D007680:Kidney Neoplasms / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology*; D015513:Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016196:S Phase; D014157:Transcription Factors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; C475820:CCNE1 protein, human; D019927:Cyclin E; D016898:Interferon-alpha; C430242:NUB1 protein, human; D015513:Oncogene Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D014157:Transcription Factors; D050760:Cyclin-Dependent Kinase Inhibitor p27