MEK/ERK-dependent uPAR expression is required for motility via phosphorylation of P70S6K in human hepatocarcinoma cells.
Motility and invasiveness events require specific intracellular signaling cascade activations. In cancer liver cells, one of these mechanisms could involve the MAPK MEK/ERK cascade activation which has been shown over expressed and activated in hepatocellular carcinoma. To study whether the MEK/ERK cascade is involved in the motility of HCC, we examined the effect of MEK inhibitor and ERK2 silencing using monolayer wound-healing assays and fluoroblock invasion systems. Evidence was provided that the MAPK cascade is a key transduction pathway which controls HCC cells motility and invasiveness. We could disconnect proliferation to motility using mitomycin C and we established that RNAi-mediated inhibition of ERK2 led to strongly reduced cell motility. To improve our understanding, we analysed the regulation and the role of urokinase receptor (uPAR) in this process. We provided evidence that uPAR was under a MEK/ERK dependent mechanism and blocking uPAR activity using specific antagonist or inhibiting its expression by RNA interference which resulted in complete inhibition of motility. Moreover, we found in MAPK inhibited cultures and in uPAR silencing cells that p70S6K phosphorylation on residue Thr-389 was significantly reduced, whereas Ser-421/Thr-424 phosphorylation did not change. We highlighted that the FRAP/mTOR pathway did not affect motility and Thr-389 phosphorylation. Furthermore, we demonstrated that p70S6K inhibition by RNA interference completely inhibited hepatocarcinoma cell motility. Therefore, targeting uPAR and/or MEK/ERK/S6K by RNA interference could be a major therapeutic strategy for the future treatment of invasive hepatocarcinoma cells.
Bessard
Anne
A
INSERM U522, IFR 140, Université de Rennes1, Rennes, France.
Frémin
Christophe
C
Ezan
Frédéric
F
Coutant
Alexandre
A
Baffet
Georges
G
D002070:Butadienes / Q000494:pharmacology; D006528:Carcinoma, Hepatocellular / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation; D006801:Humans; D008113:Liver Neoplasms / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology; D020935:MAP Kinase Signaling System; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D016685:Mitomycin / Q000494:pharmacology; D009361:Neoplasm Invasiveness; D009570:Nitriles / Q000494:pharmacology; D010456:Peptides, Cyclic / Q000494:pharmacology; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D055293:Receptors, Urokinase Plasminogen Activator; D038762:Ribosomal Protein S6 Kinases, 70-kDa / Q000235:genetics / Q000378:metabolism*; D013997:Time Factors; D014945:Wound Healing
D002070:Butadienes; D009570:Nitriles; C524898:PLAUR protein, human; D010456:Peptides, Cyclic; D047428:Protein Kinase Inhibitors; D034741:RNA, Small Interfering; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; C113580:U 0126; D016685:Mitomycin; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3