Synergistic Roles of Eukaryotic Translation Elongation Factors 1Bγ and 1A in Stimulation of Tombusvirus Minus-Strand Synthesis

Zsuzsanna Sasvari,Lara Izotova,Terri Goss Kinzy,Peter D Nagy,Dan Gallie

doi:10.1371/journal.ppat.1002438

Zsuzsanna Sasvari, Lara Izotova + Show 3 more

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https://doi.org/10.1371/journal.ppat.1002438

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Journal: PLoS Pathogens	Publication Date: Dec 15, 2011
Citations: 123	License type: CC BY 4.0

Affiliation: University of Kentucky

Abstract

Host factors are recruited into viral replicase complexes to aid replication of plus-strand RNA viruses. In this paper, we show that deletion of eukaryotic translation elongation factor 1Bgamma (eEF1Bγ) reduces Tomato bushy stunt virus (TBSV) replication in yeast host. Also, knock down of eEF1Bγ level in plant host decreases TBSV accumulation. eEF1Bγ binds to the viral RNA and is one of the resident host proteins in the tombusvirus replicase complex. Additional in vitro assays with whole cell extracts prepared from yeast strains lacking eEF1Bγ demonstrated its role in minus-strand synthesis by opening of the structured 3′ end of the viral RNA and reducing the possibility of re-utilization of (+)-strand templates for repeated (-)-strand synthesis within the replicase. We also show that eEF1Bγ plays a synergistic role with eukaryotic translation elongation factor 1A in tombusvirus replication, possibly via stimulation of the proper positioning of the viral RNA-dependent RNA polymerase over the promoter region in the viral RNA template.These roles for translation factors during TBSV replication are separate from their canonical roles in host and viral protein translation.

Highlights

Plus-stranded (+)RNA viruses recruit numerous host proteins to facilitate their replication and spread [1,2]
Expression of eEF1Bc (Tef4p) in TEF4-encoded form of eEF1Bc (tef4D) yeast increased Tomato bushy stunt virus (TBSV) replication to,80%, demonstrating that the defect in TBSV repRNA replication in tef4D yeast can be complemented.Altogether, these data established that eEF1Bc plays an important stimulatory role in TBSV replication
The cell-free extracts (CFE) extracts were programmed with the TBSV (+)repRNA and purified recombinant p33 and p92pol obtained from E. coli

Summary

Introduction

Plus-stranded (+)RNA viruses recruit numerous host proteins to facilitate their replication and spread [1,2]. Among the identified host proteins are RNA-binding proteins (RBPs), such as ribosomal proteins, translation factors and RNA-modifying enzymes [1,2,3,4,5]. The subverted host proteins likely affect several steps in viral RNA replication, including the assembly of the replicase complex and initiation of RNA synthesis. The detailed functions of recruited host RBPs in (+)RNA virus replication are known only for a small number of host factors [2,6,7,8]. Recent genome-wide screens and global proteomics approaches with TBSV and a yeast host revealed a large number of host factors interacting with viral components or affecting TBSV replication. The identified host proteins are involved in various cellular processes, such as translation, RNA metabolism, protein modifications and intracellular transport or membrane modifications [14,15,16,17]

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