MEK/ERK‐dependent uPAR expression is required for motility via phosphorylation of P70S6K in human hepatocarcinoma cells

Abstract

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.

MEK drives cyclin D1 hyperelevation during geroconversion

“…16 In fact, the MAPK pathway is known to affect phosphorylation of Raptor, p70S6K and RSK, thereby inducing S6 phosphorylation. [47][48][49] Incidentally, we observed that inhibition of MEK did not inhibit the MTOR pathway in some cell lines. This may provide the opportunity to elucidate MTOR-independent effects of MEK inhibition.…”

“…Inhibitors of MEK can indirectly inhibit the MTOR pathway in a cell type-specific manner, because MEK/ERK phosphorylates and activates raptor, p70 S6K and RSK, thereby leading to phosphorylation of S6. [47][48][49] Importantly, inhibitors of MEK completely eliminated cyclin D1, regardless whether they suppressed MTOR or not (compare U0126 effects in HT-p21 versus MEL10 cells). In contrast to HT-p21 cells, U0126 treatment did not result in inhibition of MTOR in MEL10 cells, but inhibited cell proliferation, thus causing senescence in these cells, instead of suppressing it.…”

RNAi-mediated ERK2 knockdown inhibits growth of tumor cells in vitro and in vivo

“…Other data strongly support the role of the MEK/MAPK pathway in the resistance of breast cancer cells to gefitinib and provide the rationale for novel therapeutic approaches based on combinations of signal transduction inhibitors (Normanno et al, 2006). Some recent studies have been conducted on MEK/ERK pathway by RNA interference (RNAi) in tumor cell lines suggesting a specific role of each isoform (ERK1/2) to contribute to a regulated cell cycle, motility and apoptosis progression (Vantaggiato et al, 2006;Bessard et al, 2007). Dominant-negative forms of ERK, and also ERK antisense nucleotides, inhibited proliferation of NIH 3T3 fibroblasts (Pages et al, 1993).…”

Epidermal Growth Factor Receptor Pathway Analysis Identifies Amphiregulin as a Key Factor for Cisplatin Resistance of Human Breast Cancer Cells

“…Several reports from the literature show that ERK1 and ERK2 have different functions (30). Although ERK1 is linked to cell proliferation and the survival of tumor cells (30), ERK2 has been linked to the regulation of cell motility (31). Thus, the activation of ERK1 in MCF-7 CisR cells can contribute to increased cell proliferation and cell survival.…”