Kunihiko Asakura

Theiler's Murine Encephalomyelitis Virus Leader Protein Amino Acid Residue 57 Regulates Subgroup-Specific Virus Growth on BHK-21 Cells

ABSTRACT Strains of Theilers murine encephalomyelitis virus (TMEV) are divided into two subgroups, TO and GDVII. TMEV strains show subgroup-specific virus growth and cell tropism and induce subgroup-specific diseases. Using site-directed mutagenesis, we demonstrated that the amino acid at position 57 of the leader protein (L57), which is located at the most N-terminal part of the polyprotein, regulates subgroup-specific virus growth on BHK-21 cells. Further study suggested that L57 may regulate viral RNA encapsidation, although it does not affect the synthesis of viral proteins or the assembly of viral intermediates.

Distinct cell death mechanisms by Theiler's murine encephalomyelitis virus (TMEV) infection in microglia and macrophage

DA strain of Theilers murine encephalomyelitis virus (TMEV) persists in the mouse central nervous system (CNS) and induces demyelination while GDVII strain fails to persist or demyelinate. L* protein, which is synthesized only in DA but not GDVII, is believed important in virus persistence and demyelination. Because a major reservoir for DA persistence is infiltrated macrophages or microglia, a resident macrophage of the CNS, we investigated TMEV infection of Ra2 cells, a murine microglial cell line. We found that DA strain grew well in Ra2 cells, but not GDVII strain or DAL*-1 virus (which fails to synthesize L* protein), suggesting that L* protein plays an important role in virus growth in microglia. Interestingly, in contrast to virus growth, most Ra2 cells infected with DA strain survived with no evidence of virus-induced apoptosis. These results may be important in clarifying the pathogenesis of DA-induced demyelinating disease.

Epitope-Tagged L* Protein of Theiler's Murine Encephalomyelitis Virus Is Expressed in the Central Nervous System in the Acute Phase of Infection

ABSTRACT TO subgroup strains of Theilers murine encephalomyelitis virus (TMEV) synthesize L* protein from an alternative initiation codon. We first demonstrated L* expression in the central nervous system (CNS) of TMEV-infected mice during the acute phase of infection by immunoprecipitation and immunoblotting with anti-L* antibody. In addition, we generated mutant viruses which synthesize FLAG or 3xFLAG epitope-tagged L* protein. With a mutant virus expressing 3xFLAG epitope-tagged L*, designated DA/3xFLAGL*, we investigated L* in the CNS in the acute phase of infection. DA/3xFLAGL* did not change the virus tropism in comparison with wild-type virus, and L* was clearly identified in the CNS in both susceptible and resistant strains of mice. Double immunolabeling studies showed that L* is colocalized with TMEV polyprotein and exclusively expressed in neurons.