The interferon-induced double-stranded RNA-activated protein kinase PKR will phosphorylate serine, threonine, or tyrosine at residue 51 in eukaryotic initiation factor 2alpha.
The family of eukaryotic initiation factor 2alpha (eIF2alpha) protein kinases plays an important role in regulating cellular protein synthesis under stress conditions. The mammalian kinases PKR and HRI and the yeast kinase GCN2 specifically phosphorylate Ser-51 on the alpha subunit of the translation initiation factor eIF2. By using an in vivo assay in yeast, the substrate specificity of these three eIF2alpha kinases was examined by substituting Ser-51 in eIF2alpha with Thr or Tyr. In yeast, phosphorylation of eIF2 inhibits general translation but derepresses translation of the GCN4 mRNA. All three kinases phosphorylated Thr in place of Ser-51 and were able to regulate general and GCN4-specific translation. In addition, both PKR and HRI were found to phosphorylate eIF2alpha-S51Y and stimulate GCN4 expression. Isoelectric focusing analysis of eIF2alpha followed by detection using anti-eIF2alpha and anti-phosphotyrosine-specific antibodies demonstrated that PKR and HRI phosphorylated eIF2alpha-S51Y on Tyr in vivo. These results provide new insights into the substrate recognition properties of the eIF2alpha kinases, and they are intriguing considering the potential for alternate substrates for PKR in cellular signaling and growth control pathways.
Lu
J
J
Laboratory of Eukaryotic Gene Regulation, NICHD, National Institutes of Health, Bethesda, Maryland 20892-2716, USA.
O'Hara
E B
EB
Trieselmann
B A
BA
Romano
P R
PR
Dever
T E
TE
D000483:Alleles; D002460:Cell Line; D015852:Eukaryotic Initiation Factor-2 / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D007525:Isoelectric Focusing; D010766:Phosphorylation; D014176:Protein Biosynthesis; D012694:Serine / Q000378:metabolism*; D013379:Substrate Specificity; D013912:Threonine / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism*; D019892:eIF-2 Kinase / Q000378:metabolism*
D015852:Eukaryotic Initiation Factor-2; D013912:Threonine; D014443:Tyrosine; D012694:Serine; D019892:eIF-2 Kinase