Biogenesis of vaccinia: Specific inhibition of rapidly labeled host DNA in vaccinia inoculated cells

Abstract

Because separation of the bulk and nascent (i.e., rapidly labeled) DNA fractions of HeLa cells could be effected satisfactorily it became possible to ascertain the effects of inoculating uv-irradiated vaccinia virus on in vivo host DNA synthesis. In both control and infected cells, labeled 4s fragments, formed during 1-min pulses with [ 3H]Tdr, existed as single- or double-stranded molecules covalently linked to short stretches of RNA. During the chase period, most of the rapidly labeled DNA within uninfected cells became associated with the bulk DNA component but failed to do so within inoculated cells. Host DNA ligase activity was unaffected by the infection, and there was a rapid breakdown of the 4s DNA fragments, leading to the conclusion that arrest of HeLa nuclear DNA synthesis by vaccinia is, most probably, due to the in vivo hydrolysis of nascent DNA. The present data further substantiate previous studies from this laboratory demonstrating the hydrolysis of in vitro synthesized nascent DNA by a DNAse activity originating from the virus core.