Disruption of cellular translational control by a viral truncated eukaryotic translation initiation factor 2α kinase homolog

Phosphorylation of eukaryotic translation initiation factor 2α (eIF2α) is a common cellular mechanism to limit protein synthesis in stress conditions. Baculovirus PK2, which resembles the C-terminal half of a protein kinase domain, was found to inhibit both human and yeast eIF2α kinases. Insect cells infected with wild-type, but not pk2-deleted, baculovirus exhibited reduced eIF2α phosphorylation and increased translational activity. The negative regulatory effect of human protein kinase RNA-regulated (PKR), an eIF2α kinase, on virus production was counteracted by PK2, indicating that baculoviruses have evolved a unique strategy for disrupting a host stress response. PK2 was found in complex with PKR and blocked kinase autophosphorylation in vivo, suggesting a mechanism of kinase inhibition mediated by interaction between truncated and intact kinase domains.

Recognition of Yeast mRNAs as “Nonsense Containing” Leads to Both Inhibition of mRNA Translation and mRNA Degradation: Implications for the Control of mRNA Decapping

A critical step in the degradation of many eukaryotic mRNAs is a decapping reaction that exposes the transcript to 5′ to 3′ exonucleolytic degradation. The dual role of the cap structure as a target of mRNA degradation and as the site of assembly of translation initiation factors has led to the hypothesis that the rate of decapping would be specified by the status of the cap binding complex. This model makes the prediction that signals that promote mRNA decapping should also alter translation. To test this hypothesis, we examined the decapping triggered by premature termination codons to determine whether there is a down-regulation of translation when mRNAs were recognized as “nonsense containing.” We constructed an mRNA containing a premature stop codon in which we could measure the levels of both the mRNA and the polypeptide encoded upstream of the premature stop codon. Using this system, we analyzed the effects of premature stop codons on the levels of protein being produced per mRNA. In addition, by using alterations either in cis or in trans that inactivate different steps in the recognition and degradation of nonsense-containing mRNAs, we demonstrated that the recognition of a nonsense codon led to a decrease in the translational efficiency of the mRNA. These observations argue that the signal from a premature termination codon impinges on the translation machinery and suggest that decapping is a consequence of the change in translational status of the mRNA.

Actin Depolymerization Affects Stress-Induced Translational Activity of Potato Tuber Tissue1

Changes in polymerized actin during stress conditions were correlated with potato (Solanum tuberosum L.) tuber protein synthesis. Fluorescence microscopy and immunoblot analyses indicated that filamentous actin was nearly undetectable in mature, quiescent aerobic tubers. Mechanical wounding of postharvest tubers resulted in a localized increase of polymerized actin, and microfilament bundles were visible in cells of the wounded periderm within 12 h after wounding. During this same period translational activity increased 8-fold. By contrast, low-oxygen stress caused rapid reduction of polymerized actin coincident with acute inhibition of protein synthesis. Treatment of aerobic tubers with cytochalasin D, an agent that disrupts actin filaments, reduced wound-induced protein synthesis in vivo. This effect was not observed when colchicine, an agent that depolymerizes microtubules, was used. Neither of these drugs had a significant effect in vitro on run-off translation of isolated polysomes. However, cytochalasin D did reduce translational competence in vitro of a crude cellular fraction containing both polysomes and cytoskeletal elements. These results demonstrate the dependence of wound-induced protein synthesis on the integrity of microfilaments and suggest that the dynamics of the actin cytoskeleton may affect translational activity during stress conditions.