Abstract #4414: ETV6-NTRK3 and K-Ras oncoproteins block eEF2 kinase activity under nutrient starvation by modulating both AMPK and p90RSK signaling pathways.
Authors: Gabriel Leprivier, Barak Rotblat, Eric Jan, Adi Barokas, Keith Laderoute, Christopher Proud, Poul Sorensen
Journal: Cancer Research (2009)

AACR Annual Meeting-- Apr 18-22, 2009; Denver, CO

Nutrient deprivation is a common form of cell stress during tumor development, but little is known regarding the biological response of tumor cells to this stress. To better understand this process, we sought to characterize the impact of the fusion oncoprotein ETV6-NTRK3, a constitutively activated tyrosine kinase receptor that was first discovered in pediatric sarcomas in our laboratory, and K-Ras on the response of tumor cells to nutrient starvation. Both of these kinases are known to activate the classical PI3K-Akt and Ras-Erk pathways. Stable expression of ETV6-NTRK3 and K-Ras in mouse NIH3T3 fibroblasts both lead to massive death under short-term nutrient starvation (glucose and amino acids) compared to control cells, which remained largely viable. This was not due to differences in mTOR activity, as p70S6K, S6 ribosomal protein, and 4EBP1 phosphorylation was equivalently lost in transformed and control cells in the absence of nutrients, strongly suggesting that translation initiation is blocked. Although transformed cells do not show higher proliferation compared to control cells under these conditions, both PI3K-Akt and Ras-Erk pathways remained activated and cyclin D1 expression is maintained in transformed cells. Furthermore, and in contrast to parental NIH3T3 cells, the energy sensor AMP-activated protein kinase (AMPK) fails to become fully activated in transformed cells. This kinase is crucial in control cells for maintaining survival under nutrient starvation, as demonstrated using an AMPK inhibitor (compound C) or AMPK-/- mouse embryonic fibroblasts. Interestingly, the effect on AMPK in transformed cells strongly correlates with diminished phosphorylation of one of its downstream effectors, the translation elongation factor eEF2. Reactivation of AMPK by AICAR or SMER28 compounds in these cells led to an increase in eEF2 phosphorylation, a mark of translation elongation inhibition. Therefore, eEF2 activity appears to be released from AMPK negative control in transformed cells. Moreover, we found that in transformed cells eEF2 phosphorylation is also blocked by a Ras-Erk-p90RSK signaling cascade. Indeed, small molecule inhibition of either MEK or p90RSK activities induced an increase of eEF2 phosphorylation through loss of eEF2 kinase (eEK2K) phosphorylation on serine 366 (an inhibitory phosphorylation site known to be targeted by p70S6K and p90RSK), independently of AMPK activity. These results indicate that under nutrient starvation, in contrast to parental non-transformed cells, ETV6-NTRK3 and K-Ras oncoproteins reduce phosphorylation of the major elongation factor eEF2 by inhibiting eEF2K. This can be explained by reduced AMPK activation and increased Ras-Erk-p90RSK activity. Ongoing studies are focused on determining whether transformed cells thus maintain translation elongation activity in the absence of nutrients.

Citation Information: In: Proc Am Assoc Cancer Res; 2009 Apr 18-22; Denver, CO. Philadelphia (PA): AACR; 2009. Abstract nr 4414.