The multiplication of parvovirus Lu III in a synchronized culture system

Summary

The multiplication of parvovirus Lu III in asynchronously growing and synchronized HeLa cell cultures was followed by means of immunofluorescence staining, infectivity titration, hemagglutination assay, and pulse chase labeling for replication of viral DNA. Metabolic inhibitors were used to characterize the dependence of early intracytoplasmic and late intranuclear viral antigen formation on DNA as well as on RNA synthesis.

Formation of early intracytoplasmic antigen was not controlled by the actual status of cellular physiology and occurred at a maximum rate between 4 and 8 hours p.i. Only constant low levels of infective, hemagglutinating virus could be detected during that space of time. Synthesis of early antigen did not depend onde novo synthesis of DNA. proved to be unaffected byα-amanitin and rifampicin, but was reduced in the presence of 4 μg/ml actinomycin D.

After infection in early S phase, the amount of intranuclear viral antigen increased rapidly in synchronously growing cells from 10 to 12 hours p.i. Its appearance was well correlated with replication of viral DNA as well as with maturation of progeny virus which could be demonstrated as early as 8 hours following infection. Both inhibitors specific for DNA synthesis (FUdR, ara-C, mitomycin C) and those interfering with RNA synthesis (actinomycin D,α-amanitin, rifampicin) were able to prevent the accumulation of intranuclear antigen. The only specific action, however, could be ascribed to rifampicin, since all the other inhibitors used are known to block cellular metabolic activity as well and, hence, might have stopped the cell cycle already before the cellular helper function necessary in parvovirus replication was displayed. The results suggest that the helper function is available late in S phase of the cell cycle and obviously controls replication of viral DNA.