PKR-dependent and -independent mechanisms are involved in translational shutoff during Sindbis virus infection.
The replication of Sindbis virus (SIN) profoundly affects the metabolism of infected vertebrate cells. One of the main events during SIN infection is the strong inhibition of translation of cellular mRNAs. In this study, we used a combination of approaches, including the study of SIN replication in PKR(-/-) mouse embryo fibroblasts or in the presence of an excess of catalytically inactive PKR. We show that the PKR-dependent inhibition of translation is not the only and most likely not the major pathway mediating translational shutoff during SIN infection. The PKR-independent mechanism strongly affects the translation of cellular templates, whereas translation of SIN subgenomic RNA is resistant to inhibition, and this leads to a benefit for viral replication. Our findings suggest that both PKR-dependent and non-PKR-dependent mechanisms of SIN-induced translational shutoff can be manipulated by using SIN replicons expressing mutated SIN nsP2 or kinase-defective PKR. Specifically, we show that expression of heterologous genes from SIN-based and most likely other alphavirus-based replicons can be increased by downregulating both the PKR-dependent and PKR-independent translational shutoffs.
Department of Microbiology and Immunology, University of Texas Medical Branch, 301 University Blvd., Galveston, TX 77555-1019, USA.
Rodion
R
Gorchakov
Elena
E
Frolova
Bryan R G
BR
Williams
Charles M
CM
Rice
Ilya
I
Frolov
D000818:Animals; D002460:Cell Line; D002478:Cells, Cultured; D006224:Cricetinae; D003546:Cysteine Endopeptidases / Q000235:genetics / Q000378:metabolism; D005347:Fibroblasts / Q000821:virology*; D005786:Gene Expression Regulation; D051379:Mice; D008810:Mice, Inbred C57BL; D041681:NIH 3T3 Cells; D014176:Protein Biosynthesis; D012845:Sindbis Virus / Q000472:pathogenicity*; D014779:Virus Replication; D019892:eIF-2 Kinase / Q000235:genetics / Q000378:metabolism*
D019892:eIF-2 Kinase; D003546:Cysteine Endopeptidases; C078306:nsP2 proteinase