Human cytomegalovirus up-regulates the phosphatidylinositol 3-kinase (PI3-K) pathway: Inhibition of PI3-K activity inhibits viral replication and virus-induced signaling

Infection of quiescent fibroblasts with human cytomegalovirus (HCMV) was fo und to cause a rapid activation of cellular phosphatidylinositol 3-kinase ( PI3-K), Maximum PIS-K activation occurred from 15 to 30 min postinfection. This activation was transient, and by 2 h postinfection (hpi), PI3-K activi ty had declined to preinfection levels. However, at 4 hpi, a second tier of PI3-K activation was detected, and PI3-K activity remained elevated relati ve to that of mock-infected cells for the remainder of infection. The cellu lar kinases Akt and p70S6K and the transcription factor NF-KB were activate d in a PI3-K-dependent manner at similar times following HCMV infection. An alysis using UV-irradiated virus indicated that no viral protein synthesis was necessary for the first phase of P13-K activation, but viral protein ex pression was required for the second tier of PI3-K activation, Treatment of infected fibroblasts with LY294002, a potent and specific inhibitor of PI3 -K kinase activity, caused a 4-log decrease in viral titers, LY294002 did n ot inhibit viral entry, but it did decrease viral immediate-early gene expr ession, In addition, the protein levels of two viral early genes required f or DNA replication, UL84 and UL44, were significantly lower in the presence of LY294002, Furthermore, viral DNA replication was strongly inhibited by LY294002 treatment. This inhibition of viral DNA replication could be rever sed by adding back the products of PD-K activity (PI-3,4-P-2 and PI-3,4,5-P -3), demonstrating that the effect of LY294002 on the viral life cycle was specifically due to the inhibition of PI3-R activity. These results are the first to suggest that PI3-K mediates HCMV-induced activation of host cell mitogenic pathways, They also provide strong evidence that PI3-K activation is important for initiation of viral DNA replication and completion of the viral lytic life cycle.