Cloning and characterization of a cDNA encoding a protein synthesis initiation factor-2alpha (eIF-2alpha) kinase from Drosophila melanogaster. Homology To yeast GCN2 protein kinase.
Phosphorylation of the alpha subunit of the eukaryotic initiation factor 2 (eIF-2alpha) is one of the best-characterized mechanisms for downregulating protein synthesis in mammalian cells in response to various stress conditions. In Drosophila, such a regulatory mechanism has not been elucidated. We report the molecular cloning and characterization of DGCN2, a Drosophila eIF-2alpha kinase related to yeast GCN2 protein kinase. DGCN2 contains all of the 12 catalytic subdomains characteristic of eukaryotic Ser/Thr protein kinases and the conserved sequence of eIF-2alpha kinases in subdomain V. A large insert of 94 amino acids, which is characteristic of eIF-2alpha kinases, is also present between subdomains IV and V. It is particularly notable that DGCN2 possesses an amino acid sequence related to class II aminoacyl-tRNA synthetases, a unique feature of yeast GCN2 protein kinase. DGCN2 expression is developmentally regulated. During embryogenesis, DGCN2 mRNA is dynamically expressed in several tissues. Interestingly, at later stages this expression becomes restricted to a few cells of the central nervous system. Affinity-purified antibodies, raised against a synthetic peptide based on the predicted DGCN2 sequence, specifically immunoprecipitated an eIF-2alpha kinase activity and recognized an approximately 175 kDa phosphoprotein in Western blots of Drosophila embryo extracts.
Santoyo
J
J
Centro de Biología Molecular "Severo Ochoa", Consejo Superior de Investigaciones Científicas, Universidad Autónoma de Madrid, Canto Blanco, 28049 Madrid, Spain.
Alcalde
J
J
Méndez
R
R
Pulido
D
D
de Haro
C
C
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D003001:Cloning, Molecular; D017124:Conserved Sequence; D018076:DNA, Complementary / Q000737:chemistry; D029721:Drosophila Proteins / Q000235:genetics*; D004331:Drosophila melanogaster; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011494:Protein Kinases / Q000235:genetics*; D017346:Protein Serine-Threonine Kinases / Q000235:genetics*; D012333:RNA, Messenger / Q000378:metabolism; D011817:Rabbits; D019892:eIF-2 Kinase
D018076:DNA, Complementary; D029721:Drosophila Proteins; D012333:RNA, Messenger; C585051:GCN2 protein, Drosophila; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D019892:eIF-2 Kinase