Formation of virus-like particles using a replicon system for Tacaribe Arenavirus

N. LÓPEZ, J. C. CASABONA AND M. T. FRANZE-FERNÁNDEZ

Salamanca, España

Conferencia; XIII International Conference on Negative Strand Viruses. Salamanca, Spain, June 17th-22nd, 2006; 2006

Negative Strand Viruses (B. Mahy, D. Kolakofsky, E. Villar)

FORMATION OF VIRUS-LIKE PARTICLES USING A REPLICON SYSTEM FOR TACARIBE ARENAVIRUS. N. López, J. C. Casabona and M. T. Franze-Fernández. CEVAN, Consejo Nacional de Investigaciones Científicas, Buenos Aires, Argentina. Tacaribe virus (TacV) is the prototype of New World arenaviruses. This group includes Junin virus (JunV) among other South American pathogens causing hemorrhagic fever. TacV is an enveloped virus whose bisegmented genome encodes four proteins: the nucleoprotein (N), the precursor for the envelope glycoproteins (GPC), the RNA polymerase (L) and a RING finger protein (Z). N and L proteins have been identified as the minimal trans-acting factors required for supporting viral RNA transcription and replication, Z protein being a powerful inhibitor of these processes. For Lassa and lymphocytic choriomeningitis (Old World) arenaviruses, a key role of Z in virus budding has also been reported. Here, we use a plasmid-driven reverse genetic approach to recover a TacV minigenome (MG) into infectious virus-like particles (VLPs). The expression of individual arenavirus proteins from transfected plasmids was directed by the T7 RNA polymerase, stably expressed in BSR-T7/5 cells. A TacV MG containing the chloramphenicol acetyltransferase (CAT) reporter gene was supplied by co-transfection with a polymerase I-driven plasmid. VLPs carrying the MG were detected by a CAT assay after transferring the supernatant to fresh BSR-T7/5 cells which were subsequently transfected with TacV N and L-expressing plasmids. Co-expression of TacV L, N, Z and GPC, resulted in undetectable levels of CAT activity passage. The replacement of TacV GPC and Z proteins with their JunV counterparts allowed the transfer of CAT activity, which was inhibited by anti-JunV neutralizing antibodies. This suggested that JunV GPC and Z proteins were competent in rescuing synthetic TacV ribonucleoproteins into infectious VLPs.