Mutations in the 2C region of poliovirus responsible for altered sensitivity to benzimidazole derivatives.
MRL-1237, [1-(4-fluorophenyl)-2-(4-imino-1,4-dihydropyridin-1-yl) methylbenzimidazole hydrochloride], is a potent and selective inhibitor of the replication of enteroviruses. To reveal the target molecule of MRL-1237 in viral replication, we selected spontaneous MRL-1237-resistant poliovirus mutants. Of 15 MRL-1237-resistant mutants obtained, 14 were cross-resistant to guanidine hydrochloride (mrgr), while 1 was susceptible (mrgs). Sequence analysis of the 2C region revealed that the 14 mrgr mutants contained a single nucleotide substitution that altered an amino acid residue from Phe-164 to Tyr. The mrgs mutant, on the other hand, contained a substitution of Ile-120 to Val. Through the construction of a cDNA-derived mutant, we confirmed that the single mutation at Phe-164 was really responsible for the reduced susceptibility to MRL-1237. MRL-1237 inhibited poliovirus-specific RNA synthesis in HeLa cells infected with a wild strain but not with an F164Y mutant. We furthermore examined the effect of mutations of the 2C region on the drug sensitivity of cDNA-derived guanidine-resistant and -dependent mutants. Two guanidine-resistant mutants were cross-resistant to MRL-1237 but remained susceptible to another benzimidazole, enviroxime. Either MRL-1237 or guanidine stimulated the viral replication of two guanidine-dependent mutants, but enviroxime did not. These results indicate that MRL-1237, like guanidine, targets the 2C protein of poliovirus for its antiviral effect.
Shimizu
H
H
Department of Virology II, National Institute of Infectious Diseases, Musashimurayama-shi, Tokyo 208-0011, Japan.
Agoh
M
M
Agoh
Y
Y
Yoshida
H
H
Yoshii
K
K
Yoneyama
T
T
Hagiwara
A
A
Miyamura
T
T
D000595:Amino Acid Sequence; D000998:Antiviral Agents / Q000737:chemistry / Q000494:pharmacology*; D001562:Benzimidazoles / Q000737:chemistry / Q000494:pharmacology*; D002352:Carrier Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D004352:Drug Resistance, Microbial; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D015394:Molecular Structure; D016297:Mutagenesis, Site-Directed; D017955:Poliovirus / Q000187:drug effects* / Q000235:genetics; D011725:Pyridines / Q000494:pharmacology*; D012367:RNA, Viral / Q000096:biosynthesis; D017386:Sequence Homology, Amino Acid; D017361:Viral Nonstructural Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*
D000998:Antiviral Agents; D001562:Benzimidazoles; D002352:Carrier Proteins; C407906:MRL 1237; D011725:Pyridines; D012367:RNA, Viral; D017361:Viral Nonstructural Proteins; C076023:2C protein, viral