EXPRESSION OF VACCINIA VIRUS K3L PROTEIN IN YEAST INHIBITS EUKARYOTICINITIATION FACTOR-II KINASE GCN2 AND THE GENERAL AMINO-ACID CONTROL PATHWAY

Phosphorylation of the alpha subunit of eukaryotic initiation factor-2 (eIF-2) is a well characterized mechanism regulating protein synthesi s. Viral and cellular proteins have been identified that regulate the activity of the eIF-2 alpha kinases. The regulatory protein, K3L, from vaccinia virus is homologous to the amino terminus of eIF-2 alpha and is thought to inhibit the activity of the double-stranded RNA-depende nt kinase suppressing the antiviral mechanism mediated by this kinase. We investigated whether K3L can inhibit the activity of the yeast eIF -2 alpha kinase GCN2. Expression of K3L protein in yeast reduced the l evel of eIF-2 alpha phosphorylation by GCN2 and blocked the stimulatio n of the general amino acid control pathway in response to starvation conditions. Accompanying in vitro studies showed that recombinant K3L protein reduced GCN2 autophosphorylation and phosphorylation elF-2 alp ha. In agreement with the hypothesis that K3L inhibits eIF-2 alpha kin ases by functioning as a pseudosubstrate, we observed that K3L directl y interacted with the kinase catalytic domain of GCN2. Together, these results indicate that K3L is a specific inhibitor of eIF-2 alpha kina ses from mammals and yeast and suggest that the kinases contain common structural features important for recognition of their substrate eIF- 2 alpha.