Human cytomegalovirus UL97 kinase confers ganciclovir susceptibility to recombinant vaccinia virus.
We analyzed whether the phosphotransferase encoded by the UL97 open reading frame of human cytomegalovirus (HCMV) alone is sufficient to confer ganciclovir (GCV) susceptibility to a foreign virus. Two vaccinia virus recombinants (T1 and A5) containing the UL97 open reading frames from a GCV-sensitive HCMV and from a GCV-resistant strain were constructed. T1 exhibited a GCV-sensitive phenotype in plaque reduction assays, whereas A5 did not. Moreover, T1-infected cell cultures showed a strongly increased incorporation of [14C]GCV triphosphate into macromolecular DNA, compared with recombinant A5 or vaccinia virus controls, which could be inhibited by the addition of guanosine. This shows that UL97 kinase is the only additional gene product required to make vaccinia virus susceptible to GCV, and guanosine seems to be one natural substrate for the enzyme. The system described here should be very helpful for fast and detailed functional analyses of UL97 mutations found in GCV-resistant HCMV isolates.
Metzger
C
C
Department of Virology, University of Ulm, Germany.
Michel
D
D
Schneider
K
K
Lüske
A
A
Schlicht
H J
HJ
Mertens
T
T
D001483:Base Sequence; D001859:Bone Neoplasms; D003001:Cloning, Molecular; D003587:Cytomegalovirus / Q000201:enzymology*; D017931:DNA Primers; D004279:DNA, Viral / Q000096:biosynthesis*; D015774:Ganciclovir / Q000378:metabolism / Q000494:pharmacology*; D006151:Guanosine / Q000494:pharmacology; D006801:Humans; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D012516:Osteosarcoma; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000096:biosynthesis / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000378:metabolism; D011995:Recombination, Genetic; D012044:Regression Analysis; D015183:Restriction Mapping; D017384:Sequence Deletion; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured; D014616:Vaccinia virus / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D010948:Viral Plaque Assay
D017931:DNA Primers; D004279:DNA, Viral; D011994:Recombinant Proteins; D006151:Guanosine; D017853:Phosphotransferases (Alcohol Group Acceptor); C075365:ganciclovir kinase; D015774:Ganciclovir