Although phospho rpS6 was maintained in RSK1, RSK3, and RSK4 over

Although phospho rpS6 was maintained in RSK1, RSK3, and RSK4 overex pressing cells, phospho eIF4B was only detectable in RSK3 and RSK4 overexpressing cells following PI3K inhibition. These benefits are in line with our proliferation studies suggesting that, while RSK1, RSK3 and RSK4 lower the sensitivity of cells to PI3K inhibitors, only RSK3 and RSK4 overexpressing cells exhibit a strong resistance phenotype. Two classes of protein kinases are recognized to phosphorylate rpS6 directly. The kinases primarily accountable for rpS6 phosphoryla tion will be the mTOR regulated S6 kinases, which are extremely sensitive to PI3K mTOR inhibition. The second class could be the RSK family of kinases, that are regulated by ERK signaling and are activated following mitogenic stimulation.
Based on our observation that retention of rpS6 and eIF4B phosphorylation correlates with resistance to PI3K pathway inhibitors, we hypothesized that cell lines with larger levels of activated ERK and or RSK signaling may preserve higher levels of phosphorylated S6235 236 upon PI3K block ade and as a result be somewhat insensitive to PI3K inhibition. To investi gate this possibility, we surveyed 27 breast selleck chemicals Bosutinib cancer cell lines by immu noblotting and queried Oncomine to identify breast cancer cell lines with high levels of RSK4. Notably, the two breast cancer cell lines exhibiting high levels of RSK4 in Oncomine, HCC1143 and HCC38, also demonstrated resistance towards the PI3K inhibitor GSK 1059615. As anticipated, when subjected to therapy with PI3K inhibitors, cell lines with higher levels of RSK4 activity exhibited a decrease in sensi tivity compared with the sensitive cell line MCF7. Fur thermore, each AU565 and MDA MB 231, but not MCF7, retained rpS6 and eIF4B phosphorylation when treated with diverse PI3K pathway inhibitors.
These final results suggest that, though inhibitor Sorafenib rpS6 and eIF4B phosphorylation is principally regulated by the PI3K AKT mTOR axis, within the context of RSK overexpression or activation by upstream elements, RSKs can sustain rpS6 and eIF4B phosphorylation throughout PI3K pathway downregulation. In eukaryotic cells, initiation of protein translation may be the big price limiting step in protein synthesis. Current studies have suggested that phosphorylation of Ser235 236 in rpS6 and eIF4B Ser422 is needed for cap dependent translation of mRNA. To figure out the effects of RSK4 overexpression on translation, we monitored new protein synthesis rates in vivo by labeling cells with S35 methionine. Indeed, we observed that RSK4 overexpressing cells had larger levels of total protein synthesis in both typical and PI3K inhibitor treated conditions compared with handle cells. Collectively, our data suggest that RSK overexpression prevents response to PI3K inhibition via maintenance of pro tein translation and by means of the inhibition of apoptosis.

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