Fate of parental ∅X174-DNA upon infection of starved thymine-requiring host cells

Abstract

Escherichia coli HF 4704, a thymine-requiring host for ∅X174, and a derivative, defective in the replication of ∅X replicative form, were starved in buffer and infected under various conditions with ∅Xam3 containing labeled DNA. If the infection was carried out in buffer without an energy source the parental DNA was recovered as intact single-stranded circles. If starved cells were infected in growth-medium a small percentage of the parental label was found in replicative form DNA whereas the majority was degraded. The noninfectious degradation-product was shown to contain parental DNA of varying sizes shorter than unit-length linear viral strands. The proportion of parental DNA being degraded was independent of the multiplicity of infection and the presence or absence of thymine in the medium. Washing of starved, infected cells with borate-EDTA liberated most of the parental label in particles that sedimented slower than intact phages but faster than intact DNA. They were resistant to sarcosyl but the DNA became freely sedimenting after treatment with sodium dodecyl sulfate and pronase. This DNA showed a similar distribution of degraded material as that recovered from the total lysate of infected cells, except that it contained no replicative form. All replicative form DNA synthesized under these conditions was found remaining with the cells after elution of the viral particles. These observations show that both penetration of the viral strand and synthesis of the complementary strand are impaired in starved cells and suggest, that active DNA-synthesis is required for productive infection—possibly replicating the infecting DNA out of the viral coat into the host cell.