Abstract
In cells that allow replication of vesicular stomatitis virus (VSV), there are two phases of translation inhibition: an early block of host translation and a later inhibition of viral translation. We investigated the phosphorylation of the alpha subunit of the eIF2 complex during these two phases of viral infection. In VSV-infected cells, the accumulation of phosphorylated (inactivated) eIF2alpha did not begin until well after host protein synthesis was inhibited, suggesting that it only plays a role in blocking viral translation later after infection. Consistent with this, cells expressing an unphosphorylatable eIF2alpha showed prolonged viral protein synthesis without an effect on host protein synthesis inhibition. Induction of eIF2alpha phosphorylation at early times of viral infection by treatment with thapsigargin showed that virus and host translation are similarly inhibited, demonstrating that viral and host messages are similarly sensitive to eIF2alpha phosphorylation. A recombinant virus that expresses a mutant matrix protein and is defective in the inhibition of host and virus protein synthesis showed an altered phosphorylation of eIF2alpha, demonstrating an involvement of viral protein function in inducing this antiviral response. This analysis of eIF2alpha phosphorylation, coupled with earlier findings that the eIF4F complex is modified earlier during VSV infection, supports a temporal/kinetic model of translation control, where at times soon after infection, changes in the eIF4F complex result in the inhibition of host protein synthesis; at later times, inactivation of the eIF2 complex blocks VSV protein synthesis.
Highlights
Tion of both host and viral translation are apparent during infection with the prototype rhabdovirus vesicular stomatitis virus (VSV).1 During VSV infection, there is a rapid inhibition of host mRNA translation early after infection
A substantial body of evidence implicates the phosphorylation of the initiation factor eIF2␣ as a major player in inhibiting viral translation [5]. eIF2␣ is a subunit of eIF2, the multiprotein complex that is responsible for recruiting the initiator tRNA to the 40 S subunit of the ribosome in a GTP-dependent manner
We showed that after VSV infection, there is a rapid alteration of the eIF4F complex, indicated by the dephosphorylation of the eIF4E subunit [16], which was correlated with the inhibition of host protein synthesis
Summary
Tion of both host and viral translation are apparent during infection with the prototype rhabdovirus vesicular stomatitis virus (VSV).1 During VSV infection, there is a rapid inhibition of host mRNA translation early after infection. The rates of protein synthesis were determined from images similar to A as detailed under “Experimental Procedures.” D, extracts from mock- and VSV-infected cells were analyzed by Western blotting with antibodies against eIF2␣, phosphorylated at serine 51 (eIF2␣-P), and total eIF2␣.
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