Characterization of the untranslated region of lymphocytic choriomeningitis virus S segment

Abstract

ABSTRACTLymphocytic choriomeningitis virus (LCMV) is a prototypic arenavirus. The viral genome consists of two RNA segments, L and S. The 5’- and 3’-termini of both L and S segments are highly conserved among arenaviruses. These regions consist of 19 complementary base pairs and are essential for viral genome replication and transcription. In addition to these 19 nucleotides in the 5’- and 3’-termini, there are untranslated regions (UTRs) composed of 58 and 41 nucleotide residues in the 5’ and 3’ UTRs, respectively, in the LCMV S segment. Their functional roles, however, have yet to be elucidated. In this study, a reverse genetics and a minigenome system for the LCMV strain WE were established and used to analyze the function of these regions. The results obtained from these analyses, plus RNA secondary structure prediction, revealed that not only these 19 nucleotides but also the 20th–40th and 20th–38th nucleotides located downstream of the 19 nucleotides in the 5’- and 3’-termini, respectively, are heavily involved in viral genome replication and transcription. Furthermore, the introduction of mutations in these regions depressed viral propagation in vitro and enhanced attenuation in vivo. Conversely, recombinant LCMVs (rLCMVs), which had various deletions in the other UTRs, propagated as well as wild-type LCMV in vitro but were attenuated in vivo. Most mice previously infected with rLCMVs with mutated UTRs, when further infected with a lethal dose of wild-type LCMV, survived. These results suggest that rLCMVs with mutated UTRs could be candidates for an LCMV vaccine.IMPORTANCEThe function of untranslated regions (UTRs) of the arenavirus genome has not well been studied except for the 19 nucleotides of the 5’- and 3’-termini. In this study the function of the UTRs of the LCMV S segment was analyzed. It was found that not only the 19 nucleotides of the 5’- and 3’-termini but also the 20th–40th and 20th–38th nucleotides located downstream of the 19 nucleotides in the 5’- and 3’-termini, respectively, were involved in viral genome replication and transcription. Furthermore, other UTRs in the S segment were involved in virulence in vivo. The introduction of mutations to these regions makes it possible to establish attenuated LCMV and potentially develop LCMV vaccine candidates.