Overexpression of cellular dihydrofolate reductase abolishes the anticytomegaloviral activity of methotrexate.

Cytomegalovirus (CMV) stimulates numerous cellular pathways upon infection. One of these pathways involves activation of dihydrofolate reductase (DHFR), an essential enzyme in the biosynthesis of purines and thymidylate. Here we report that methotrexate (MTX), an inhibitor of DHFR, suppresses murine CMV replication at the level of DNA synthesis in quiescent NIH 3T3 cells. However, MTX has no antiviral activity in NIH 3T3 sublines resistant to MTX due to DHFR overexpression. These results directly link MTX antiviral activity to DHFR and demonstrate that DHFR plays an essential role for CMV replication in quiescent cells.