Involvement of RhoA and RalB in geranylgeranyltransferase I inhibitor-mediated inhibition of proliferation and migration of human oral squamous cell carcinoma cells.
OBJECTIVE
Geranylgeranyltransferase I is required for the prenylation of the small GTPases. The effect of GGTase I inhibitors (GGTIs) on oral squamous cell carcinoma (SCC) cells was examined.


METHODS
The GGTI-treated cells were examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay, flow cytometric analysis, transwell chamber assays, and immunofluorescent staining. Small GTPases were detected by immunoblot analysis, and siRNA were used for silencing RalA and RalB.


RESULTS
GGTI suppressed the proliferation of oral SCC cells and induced cell cycle arrest at G(1), but the sub-G(1) fraction was small. The expression of the cyclin-dependent kinase (CDK) inhibitor p21(Waf1/Cip1), but not p27(Kip1), was markedly increased by GGTI. There was an apparent increase in the expression and reduction in the membrane localization of RhoA and RalB, but not Ras and RalA. Assays with transwell chambers and wound healing and invasion revealed the migrative and invasive capabilities of SAS cells to be inhibited by GGTI. Actin filaments were rearranged and stress fibers and peripheral cell processes were lost, accompanying cell rounding. siRNA for RalB, but not RalA, significantly suppressed the migration of SAS cells.


CONCLUSIONS
These results suggest that GGTI inhibits the geranylgeranylation of RhoA and increases the p21(Waf1/Cip1) level, resulting in cell cycle arrest at G(1) to decrease cell proliferation, and that of RalB to suppress the migration and invasion by oral SCC cells. GGTIs may be useful as inhibitors of invasion and metastasis in cases of oral SCC.
Department of Oral and Maxillofacial Surgery, Osaka University Graduate School of Dentistry, Osaka, Japan. hmdmskz@dent.osaka-u.ac.jp
Masakazu
M
Hamada
Tetsuei
T
Miki
Soichi
S
Iwai
Hidetaka
H
Shimizu
Yoshiaki
Y
Yura
D019883:Alkyl and Aryl Transferases / Q000037:antagonists & inhibitors*; D015153:Blotting, Western; D002294:Carcinoma, Squamous Cell / Q000188:drug therapy* / Q000201:enzymology / Q000473:pathology; D002458:Cell Fractionation; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D050756:Cyclin-Dependent Kinase Inhibitor Proteins / Q000494:pharmacology; D003599:Cytoskeleton / Q000378:metabolism / Q000648:ultrastructure; D020558:GTP Phosphohydrolases / Q000378:metabolism; D006801:Humans; D007202:Indicators and Reagents; D009062:Mouth Neoplasms / Q000188:drug therapy* / Q000201:enzymology / Q000473:pathology; D009361:Neoplasm Invasiveness / Q000473:pathology; D017368:Protein Prenylation / Q000187:drug effects; D034741:RNA, Small Interfering; D014945:Wound Healing; D020725:ral GTP-Binding Proteins / Q000235:genetics*; D018631:ras Proteins / Q000096:biosynthesis; D020742:rhoA GTP-Binding Protein / Q000235:genetics*
D050756:Cyclin-Dependent Kinase Inhibitor Proteins; D007202:Indicators and Reagents; D034741:RNA, Small Interfering; C483163:Ralb protein, human; C066737:RHOA protein, human; D019883:Alkyl and Aryl Transferases; C083291:geranylgeranyltransferase type-I; D020558:GTP Phosphohydrolases; D020725:ral GTP-Binding Proteins; D018631:ras Proteins; D020742:rhoA GTP-Binding Protein