Concatemeric forms of intracellular herpesvirus DNA

Abstract

Newly synthesized pseudorabies viral DNA sediments in neutral sucrose gradients as molecules with high S values. The fast-sedimenting structures are precursors of mature viral DNA and represent replicative intermediates of this DNA. The sedimentation characteristics of the replicative intermediates are not affected by treatments which should eliminate protein, membrane fragments, or RNA. The buoyant density of the replicative intermediates in cesium chloride indicates that they probably consist of DNA only. Electron microscopic examination of intracellular viral DNA purified by equilibrium sedimentation in CsCl revealed DNA “tangles,” as well as longer than unit-size linear molecules. Mature viral-DNA preparations treated similarly did not contain such structures. Thus, concatemeric forms of viral DNA are present in the infected cells. Maturation of newly synthesized viral DNA to unit-size length occurs in the absence of further DNA synthesis but is dependent on protein synthesis. The “Hirt” procedure separates mature viral DNA from cellular DNA and also separates viral replicative intermediates from mature viral DNA.