Stress and Translation lab

PROJECT TITLE : Role of GCN2 and eukaryotic translation initiation factor 2 (eIF2) phosphorylation in cell survival/transformation under different stress conditions

SUMMARY: Stress response in eukaryotes is characterized by changes in gene expression, and one of the first steps of this response is the activation of kinases which phosphorylate the translation initiation factor eIF2 (eIF2alpha kinases). Mammalian cells express four eIF2alpha kinases which respond especificaly to different forms of stress: GCN2, PKR, PERK and HRI. The phosphorylation of eIF2alpha leads to a global inhibition of protein synthesis, but also to the specific activation of the translation of some mRNAs involved on recovery, adaptation and survival. It is still unknown the relevance of this response in cell survival under certain stress conditions. Thus, the main goals of this project are to study:

1. The implication of GCN2 activation in response to nutrient deprivation and ultraviolet (UV) radiation in short and long term cell survival

2. The role of GCN2 activation and eIF2 phosphorylation in the oncogenic transformation of UV-irradiated cells

3. Regulation of eIF2 phosphorylation and stress specific protein expression by the GCN2 modulator GCN1

For this purpose we will use cell cultures of immortalized mouse embryonic fibroblasts from wild type and GCN2 knock out mice. We will also produce GCN1 knockdown cells. Activation of GCN2 (auto-phosphorylation) and the subsequent eIF2alpha phosphorylation, together with the expression of the transcription factor ATF4 (whose translation is activated under stress conditions) will be analyzed. Short term survival, long term survival and proliferation will also be assayed,. Finally, we will test oncogenic transformation by soft agar colony-forming assay.

With this project we aim to explore the possible use of GCN2 and functionally associated proteins as therapeutic targets to prevent oncogenic transformation of cells in response to UV radiation.