Synthesis of Seoul virus RNA and structural proteins in cultured cells

Summary. Seoul virus is a hantavirus that causes hemorrhagic fever with renal syndrome (HFRS). The virion has a tripartite (S, M, and L) negative-stranded RNA genome, which is characteristic of the family Bunyaviridae. However, the molecular basis of virus replication is not well known. We established a Northern blot hybridization (NB) procedure using digoxygenin-labeled RNA probes, to quantitate the hantaviral plus- and minus-strand RNAs separately. Virus RNA replication was analyzed in infected Vero E6 cells. When the Vero E6 cells were infected with Seoul virus strain KI-83-262 (KI) at m.o.i.=0.25, the plus-strand RNA was detected within 1h post-infection (hpi), and the minus-strand RNA was detected subsequently. Using laser confocal microscopy, the nucleocapsid protein (NP) was detected within 2hpi, and accumulated as scattered granules in the cytoplasm until 24hpi. In contrast, the G2 protein first appeared at 8hpi, was immediately transported to the Golgi, and accumulated in the Golgi until 24hpi. Infectious virus particles were released into the medium at 24hhpi. These findings indicate that hantavirus RNA replication starts with the appearance of NP at 2hpi, glycoproteins then accumulate gradually in the Golgi, and virion formation is initiated once the viral RNAs and proteins have accumulated.Received October 10, 2002; accepted April 25, 2003 Published online July 17, 2003