Gcn2 eIF2a kinase mediates combinatorial translational regulation through nucleotide motifs and uORFs in the mRNA 5’ leader regions

Cellular signaling controls translation through cis regulatory elements found in mRNAs. Gcn2 is the master regulator of translation during nutrient limitation in normal and cancer cells. Activated Gcn2 phosphorylates eIF2a, thereby repressing general translation while activating translation of specific mRNAs with upstream ORFs (uORFs) in its leader regions. Here we performed genome-wide measurement of mRNA translation during histidine starvation in fission yeast Schizosaccharomyces pombe. Polysomal microarray hybridization experiments identified a group of 1779 genes whose translation is up-regulated in Gcn2-dependent manner. We find that translation is reprogrammed to enhance organellar synthesis and transcription and repress ribosome synthesis. The 1779 genes included gcn5 and hri2 shown to promote growth under histidine starvation. They encode histone acetyl transferase and heme-binding eIF2a kinase activated by oxidative stress, and their 5’-leader contains 3 or 4 uORFs, respectively. Our reporter studies show that uORFs in gcn5 and hri2 operate similarly to those found in S. cerevisiae GCN4, the founding case for uORF-dependent regulation. Moreover, motif analysis identified 5’-UGA(C/G)GG-3’ as a motif promoting translation during starvation. Using hrd1 5’UTR with such a motif, we demonstrated its requirement in Gcn2-dependent translational control. To our knowledge, this is the first nucleotide motif found to be responsible for translational control by Gcn2. We propose that Gcn2 mediates translational control of more specific mRNAs than previously anticipated Polysome profile at exponentially-growing phase in the fission yeast with and without 3AT.