PHOSPHORYLATION OF PLANT EUKARYOTIC INITIATION FACTOR-II BY THE PLANT-ENCODED DOUBLE-STRANDED RNA-DEPENDENT PROTEIN-KINASE, PPKR, AND INHIBITION OF PROTEIN-SYNTHESIS IN-VITRO

Regulation of protein synthesis by eukaryotic initiation factor-2 alph a (eIF-2 alpha) phosphorylation is a highly conserved phenomenon in eu karyotes that occurs in response to various stress conditions. Protein kinases capable of phosphorylating eIF-2 alpha have been characterize d from mammals and yeast. However, the phenomenon of eIF2-alpha-mediat ed regulation of protein synthesis and the presence of an elF-2 alpha kinase has not been demonstrated in higher plants. We show that plant eIF-2 alpha (peIF-2 alpha) and mammalian eIF-2 alpha (meIF-2 alpha) ar e phosphorylated similarly by both the double-stranded RNA-binding kin ase, pPKR, present in plant ribosome salt wash fractions and the meIF- 2 alpha kinase, PHR. By several criteria, phosphorylation of peIF-2 al pha is directly correlated with pPKR. protein and autophosphorylation levels. Significantly, pPKR is capable of specifically phosphorylating Ser(51) in a synthetic eIF-2 alpha peptide, a key characteristic of t he eIF-2 alpha kinase family. Taken together, these data support the c oncept that pPKR is a member of the eIF-2 alpha kinase family. In addi tion, the inhibition of brome mosaic virus RNA in vitro translation in wheat germ lysates by the addition of double-stranded RNA, phosphoryl ated peIF-2 alpha; meIF-2 alpha, or activated human PKR suggests that plant protein synthesis may be regulated via phosphorylation of eIF-2 alpha.