Adaptation of Theiler's Virus to L929 Cells: Mutations in the Putative Receptor Binding Site on the Capsid Map to Neutralization Sites and Modulate Viral Persistence

Abstract

Persistent strains of Theiler's virus, a murine picornavirus, produce a life-long infection of the central nervous system of the mouse and induce a chronic demyelinating disease. Strain DA1, a molecular clone of such a persistent strain, produces a prominent cytopathic effect in BHK-21 cells but is less efficient at infecting L929 cells. We cloned the cDNA of a derivative of virus DA1, adapted to promote a rapid cytopathic effect in L929 cells. Adaptation of the new variant (named KJ6) to L929 cells correlated with an enhanced viral entry rather than with an increased replication rate of the genome. Mutations responsible for L929 cells adaptation occurred in amino acids exposed at the surface of the capsid, in the CD loop of VP1 (100-102) and in the EF loop of VP2 (162-171-173), suggesting that these residues could be involved in receptor recognition. These two clusters of amino acids are precisely known to be part of neutralization epitopes. They also differentiate persistent from neurovirulent strains of Theiler's virus. Adaptation of the virus to L929 cells was accompanied by attenuation of its virulence for the mouse. Taken together, these data suggest a close relationship between receptor binding, virus neutralization, and virus phenotype.

The Leader Protein of Theiler's Virus Inhibits Immediate-Early Alpha/Beta Interferon Production

“…The capsid coding region of pTM598, contained in a BsiWI-BamHI fragment (nt 1265 to 3925 of DA1), was then replaced by the corresponding region of pKJ6 (12), a variant of pTMDA1 with mutations in the capsid coding region that enhance infection of L929 cells. The recombinant carrying the L cys mutations and the capsid adapted to L929 cells was called pTM659 (Table 1).…”

“…2fTGH, U3A (23), BALB/3T3, and L929 cells were cultured in Dulbecco's modified Eagle medium (Gibco-BRL) supplemented with 10% fetal bovine serum (Gibco-BRL), 100 IU of penicillin/ml, 100 g of streptomycin/ml, and 1 mM sodium pyruvate. TMEV derivatives were produced by electroporation of BHK-21 cells (24) with the genomic RNA transcribed in vitro from plasmids carrying the corresponding cDNAs: pTMDA1 (21,24), pKJ6 (12), pTM564 (24), pTM598, and pTM659 (this work).…”

Characterization of Interferon-α 13, a Novel Constitutive Murine Interferon-α Subtype

“…The KJ6 and TM659 are the corresponding wild-type and L cys -mutant viruses carrying a capsid adapted to infect L929 cells (29,31). These viruses were produced as described previously by transfection of BHK-21 cells with viral RNAs transcribed in vitro from the corresponding infectious cDNA clones, pTMDA1, pTM598, pKJ6, and pTM659 (29).…”

TGF-β1 suppresses T cell infiltration and VP2 puff B mutation enhances apoptosis in acute polioencephalitis induced by Theiler's virus

“…These results support the hypothesis that VP2 puff B is important for cell binding. In addition, studies of recombinant viruses and monoclonal antibody escape mutant viruses have provided support for the involvement of this region, formed by VP1 loop II (Jnaoui and Michiels, 1998;Lin et al, 1998;Wada et al, 1994;Zurbriggen et al, 1989) and VP2 puff B (Jnaoui and Michiels, 1998;Sato et al, 1996), in viral persistence and differences in disease phenotype.…”