Abstract
Mammalian arenavirus (mammarenavirus) mRNAs are characterized by 5′-capped and 3′-nonpolyadenylated untranslated regions (UTRs). We previously reported that the nonpolyadenylated 3′-UTR of viral mRNA (vmRNA), which is derived from the noncoding intergenic region (IGR), regulates viral protein levels at the posttranscriptional level. This finding provided the basis for the development of novel live-attenuated vaccines (LAVs) against human pathogenic mammarenaviruses. Detailed information about the roles of specific vmRNA 3′-UTR sequences in controlling translation efficiency will help in understanding the mechanism underlying attenuation by IGR manipulations. Here, we characterize the roles of cis-acting mRNA regulatory sequences of a prototypic mammarenavirus, lymphocytic choriomeningitis virus (LCMV), in modulating translational efficiency. Using in vitro transcribed RNA mimics encoding a reporter gene, we demonstrate that the 3′-UTR of nucleoprotein (NP) mRNA without a poly(A) tail promotes translation in a poly(A)-binding protein-independent manner. Comparison with the 3′-UTR of glycoprotein precursor mRNA, which is translated less efficiently, revealed that a 10-nucleotide sequence proximal to the NP open reading frame is essential for promoting translation. Modification of this 10-nucleotide sequence also impacted reporter gene expression in recombinant LCMV. Our findings will enable rational design of the 10-nucleotide sequence to further improve our mammarenavirus LAV candidates and to develop a novel LCMV vector capable of controlling foreign gene expression.
Highlights
Emergence of novel mammarenaviruses that cause hemorrhagic fever [2, 3]
We previously demonstrated that intergenic region (IGR) sequences play a critical role in viral mRNA (vmRNA) translation efficiency [17]
To examine whether the length of the 30-untranslated region (UTR) of lymphocytic choriomeningitis virus (LCMV) NP mRNA plays a critical role in translation efficiency, we assessed ZsG levels in cells transfected with a series of 30-end-truncated vmRNA-like mRNA (vlmRNA)
Summary
Mounting evidence has indicated that a globally distributed prototypic mammarenavirus, lymphocytic choriomeningitis virus (LCMV), is an important human pathogen of clinical significance, especially in immunocompromised or pregnant individuals [4,5,6,7] Despite their significant impact on human health, medical countermeasures against human pathogenic mammarenaviruses are limited. We previously used the finding that IGR sequences regulate vmRNA translation efficiency to develop a novel genetic approach for rational attenuation of Translation regulation by the 30-UTR of LCMV mRNAs mammarenaviruses. Partial deletions in the L segment IGRs (L-IGRs) of other mammarenaviruses, Lujo virus and Machupo virus (the causative agent of Bolivian hemorrhagic fever), were associated with an attenuated phenotype in cultured cells [20] and in vivo [21]. Our findings will enable rational design of the 10-nt sequence to further improve our mammarenavirus LAV candidates and to develop a novel LCMV vector capable of controlling foreign gene expression
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