Abstract
BackgroundUnderstanding the biochemical mechanisms contributing to melanoma development and progression is critical for therapeutical intervention. LKB1 is a multi-task Ser/Thr kinase that phosphorylates AMPK controlling cell growth and apoptosis under metabolic stress conditions. Additionally, LKB1Ser428 becomes phosphorylated in a RAS-Erk1/2-p90RSK pathway dependent manner. However, the connection between the RAS pathway and LKB1 is mostly unknown.Methodology/Principal FindingsUsing the UV induced HGF transgenic mouse melanoma model to investigate the interplay among HGF signaling, RAS pathway and PI3K pathway in melanoma, we identified LKB1 as a protein directly modified by HGF induced signaling. A variety of molecular techniques and tissue culture revealed that LKB1Ser428 (Ser431 in the mouse) is constitutively phosphorylated in BRAFV600E mutant melanoma cell lines and spontaneous mouse tumors with high RAS pathway activity. Interestingly, BRAFV600E mutant melanoma cells showed a very limited response to metabolic stress mediated by the LKB1-AMPK-mTOR pathway. Here we show for the first time that RAS pathway activation including BRAFV600E mutation promotes the uncoupling of AMPK from LKB1 by a mechanism that appears to be independent of LKB1Ser428 phosphorylation. Notably, the inhibition of the RAS pathway in BRAFV600E mutant melanoma cells recovered the complex formation and rescued the LKB1-AMPKα metabolic stress-induced response, increasing apoptosis in cooperation with the pro-apoptotic proteins Bad and Bim, and the down-regulation of Mcl-1.Conclusions/SignificanceThese data demonstrate that growth factor treatment and in particular oncogenic BRAFV600E induces the uncoupling of LKB1-AMPKα complexes providing at the same time a possible mechanism in cell proliferation that engages cell growth and cell division in response to mitogenic stimuli and resistance to low energy conditions in tumor cells. Importantly, this mechanism reveals a new level for therapeutical intervention particularly relevant in tumors harboring a deregulated RAS-Erk1/2 pathway.
Highlights
Melanoma is the most lethal human skin cancer and its incidence is rapidly rising world-wide [1]
We proposed to identify novel molecules involved in melanoma development and progression analyzing the hepatocyte growth factor (HGF) specific signaling in the UV induced HGF transgenic melanoma mouse model
Analysis of B16F1 melanoma cells showed that the phosphorylation of the LKB1Ser431 was p-Erk1/2 dependent (Fig. 1C) and, inhibition of Mek1/2 after HGF treatment in 37-31E cells totally abolished the phosphorylation of Erk1/2, p90RSK and LKB1Ser431 (Fig. 1C)
Summary
Melanoma is the most lethal human skin cancer and its incidence is rapidly rising world-wide [1]. Dissecting the aberrant signaling pathways that are critical to melanomagenesis and understanding the mechanisms by which these pathways interact with each other has become the recent focus of research directed at melanoma therapeutic intervention. HGF signaling activates Ras-Erk1/2 and PI3K-AKT pathways, and Ras pathway activation has been shown to play a role in melanoma development and maintenance [2]. Support for PI3K-AKT pathway signaling dysfunction in melanomagenesis has been demonstrated by the documented loss of the tumor suppressor PTEN-containing chromosomal region in 5–20% of melanomas as well as the over expression of AKT3 in the advanced stages of this disease [4,5]. Understanding the biochemical mechanisms contributing to melanoma development and progression is critical for therapeutical intervention. The connection between the RAS pathway and LKB1 is mostly unknown
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