A mammalian homologue of GCN2 protein kinase important for translational control by phosphorylation of eukaryotic initiation factor-2 alpha

A family of protein kinases regulates translation in response to different cellular stresses by phosphorylation of the alpha subunit of eukaryotic ini tiation factor-2 (eIF-2 alpha). In yeast, an eIF-2 alpha kinase, GCN2, func tions in translational control in response to amino acid starvation. It is thought that uncharged tRNA that accumulates during amino acid limitation b inds to sequences in GCN2 homologous to histidyl-tRNA synthetase (HisRS) en zymes, leading to enhanced kinase catalytic activity. Given that starvation for amino acids also stimulates phosphorylation of eIF-2 alpha in mammalia n cells, we searched for and identified a GCN2 homologue in mice. We cloned three different cDNAs encoding mouse GCN2 isoforms, derived from a single gene, that vary in their amino-terminal sequences. Like their yeast counter part, the mouse GCN2 isoforms contain HisRS-related sequences juxtaposed to the kinase catalytic domain. While GCN2 mRNA was found in all mouse tissue s examined, the isoforms appear to be differentially expressed. Mouse GCN2 expressed in yeast was found to inhibit growth by hyperphosphorylation of e IF-2 alpha, requiring both the kinase catalytic domain and the HisRS-relate d sequences. Additionally, lysates prepared from yeast expressing mGCN2 wer e found to phosphorylate recombinant eIF-2 alpha substrate, Mouse GCN2 acti vity in both the in vivo and in vitro assays required the presence of serin e-51, the known regulator phosphorylation site in eIF-2 alpha. Together, ou r studies identify a new mammalian eIF-2 alpha kinase, GCN2, that can media te translational control.