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Eur J Biochem. 1999 Oct;265(2):754-62.

Characterization of a mammalian homolog of the GCN2 eukaryotic initiation factor 2alpha kinase.

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1
Centro de Biología Molecular 'Severo Ochoa', Consejo Superior de Investigaciones Científicas, Universidad Autónoma de Madrid, Spain.

Abstract

In eukaryotic cells, protein synthesis is regulated in response to various environmental stresses by phosphorylating the alpha subunit of the eukaryotic initiation factor 2 (eIF2alpha). Three different eIF2alpha kinases have been identified in mammalian cells, the heme-regulated inhibitor (HRI), the interferon-inducible RNA-dependent kinase (PKR) and the endoplasmic reticulum-resident kinase (PERK). A fourth eIF2alpha kinase, termed GCN2, was previously characterized from Saccharomyces cerevisiae, Drosophila melanogaster and Neurospora crassa. Here we describe the cloning of a mouse GCN2 cDNA (MGCN2), which represents the first mammalian GCN2 homolog. MGCN2 has a conserved motif, N-terminal to the kinase subdomain V, and a large insert of 139 amino acids located between subdomains IV and V that are characteristic of the known eIF2alpha kinases. Furthermore, MGCN2 contains a class II aminoacyl-tRNA synthetase domain and a degenerate kinase segment, downstream and upstream of the eIF2alpha kinase domain, respectively, and both are singular features of GCN2 protein kinases. MGCN2 mRNA is expressed as a single message of approximately 5.5 kb in a wide range of different tissues, with the highest levels in the liver and the brain. Specific polyclonal anti-(MGCN2) immunoprecipitated an eIF2alpha kinase activity and recognized a 190 kDa phosphoprotein in Western blots from either mouse liver or MGCN2-transfected 293 cell extracts. Interestingly, serum starvation increased eIF2alpha phosphorylation in MGCN2-transfected human 293T cells. This finding provides evidence that GCN2 is the unique eIF2alpha kinase present in all eukaryotes from yeast to mammals and underscores the role of MGCN2 kinase in translational control and its potential physiological significance.

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