Untranslated regions of diverse plant viral RNAs vary greatly in translation enhancement efficiency

Qiuling Fan,W Allen Miller,Krzysztof Treder

doi:10.1186/1472-6750-12-22

Qiuling Fan, W Allen Miller + Show 1 more

Open Access

https://doi.org/10.1186/1472-6750-12-22

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Abstract

BackgroundWhole plants or plant cell cultures can serve as low cost bioreactors to produce massive amounts of a specific protein for pharmacological or industrial use. To maximize protein expression, translation of mRNA must be optimized. Many plant viral RNAs harbor extremely efficient translation enhancers. However, few of these different translation elements have been compared side-by-side. Thus, it is unclear which are the most efficient translation enhancers. Here, we compare the effects of untranslated regions (UTRs) containing translation elements from six plant viruses on translation in wheat germ extract and in monocotyledenous and dicotyledenous plant cells.ResultsThe highest expressing uncapped mRNAs contained viral UTRs harboring Barley yellow dwarf virus (BYDV)-like cap-independent translation elements (BTEs). The BYDV BTE conferred the most efficient translation of a luciferase reporter in wheat germ extract and oat protoplasts, while uncapped mRNA containing the BTE from Tobacco necrosis virus-D translated most efficiently in tobacco cells. Capped mRNA containing the Tobacco mosaic virus omega sequence was the most efficient mRNA in tobacco cells. UTRs from Satellite tobacco necrosis virus, Tomato bushy stunt virus, and Crucifer-infecting tobamovirus (crTMV) did not stimulate translation efficiently. mRNA with the crTMV 5′ UTR was unstable in tobacco protoplasts.ConclusionsBTEs confer the highest levels of translation of uncapped mRNAs in vitro and in vivo, while the capped omega sequence is most efficient in tobacco cells. These results provide a basis for understanding mechanisms of translation enhancement, and for maximizing protein synthesis in cell-free systems, transgenic plants, or in viral expression vectors.

Highlights

Whole plants or plant cell cultures can serve as low cost bioreactors to produce massive amounts of a specific protein for pharmacological or industrial use
All constructs consisted of the firefly luciferase Open reading frame (ORF) flanked by the complete untranslated region (UTR) of each virus (Figure 1)
The omega sequence of Tobacco mosaic virus (TMV) and the Crucifer-infecting tobamovirus (crTMV) internal ribosome entry site (IRES) are both located in 5′ UTRs

Summary

Introduction

Whole plants or plant cell cultures can serve as low cost bioreactors to produce massive amounts of a specific protein for pharmacological or industrial use. Many plant viral RNAs harbor extremely efficient translation enhancers. Few of these different translation elements have been compared side-by-side. We compare the effects of untranslated regions (UTRs) containing translation elements from six plant viruses on translation in wheat germ extract and in monocotyledenous and dicotyledenous plant cells. All host cellular mRNAs and some viral RNAs have a 5′ cap to recruit the translational machinery via binding of the eIF4E subunit of translation initiation factor complex eIF4F [1,2]. Many plant viral RNAs lack a 5′ cap and have either an internal ribosome entry site (IRES) in the 5′ untranslated region (UTR) or a cap-independent translation element (CITE) in the 3′ UTR to facilitate translation (reviewed in [3,4,5,6]). STNV-1 RNA contains a completely different type of 3′ CITE, called the translation enhancer domain (TED), CC UU

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