Herpes simplex virus mutant KG111 contains a
nonsense mutation at
codon 44 of the viral
thymidine kinase (tk) gene and produces low amounts of a truncated tk
polypeptide. We tested mutant KG111 and related viruses that specify varying amounts of similar truncated tk
polypeptides for their sensitivities to
antiviral nucleoside analogs at different temperatures using plaque reduction assays. The results of these assays showed that the
nonsense mutation confers high resistance to bromovinyldeoxyuridine (BVdU) at any temperature and temperature-dependent resistance to
acyclovir (ACV),
buciclovir (BCV),
ganciclovir (
DHPG), and fluoroiodoarabinouracil (
FIAU). Above relatively low threshold levels of tk that varied depending on the
drug tested, viruses exhibited full sensitivity to ACV, BCV,
DHPG, and
FIAU at 34 degrees. Below these threshold levels, however, decreases in
drug sensitivity were linear with decreases in tk levels, forming the basis of a pharmacological assay for tk gene expression. Studies of
thymidine (TdR) anabolism in infected 143 tk-cells showed that when high TdR concentrations were added to the medium, KG111 directed
thymidine monophosphate (
TMP) formation at rates consonant with the amount of tk
polypeptide produced by the mutant. When low concentrations to TdR were added to the medium, however, KG111 directed
TMP formation at a rate similar to that directed by wild-type virus, indicating that the truncation of the tk
polypeptide had little or no effect on tk activity at 34 degrees. Subsequent anabolism to
thymidine diphosphate and
thymidine triphosphate was reduced in KG111-infected cells, indicating a defect in
TMP kinase activity that explains this mutant's resistance to BVdU. Despite the low levels of tk and
TMP kinase activity expressed by KG111, this mutant established reactivatable
latent infections as efficiently as wild-type virus in a mouse model.