Replication of herpesvirus DNA. V. Maturation of concatemeric DNA of pseudorabies virus to genome length is related to capsid formation

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Abstract
The maturation of pseudorabies virus DNA from the replicative concatemeric form to molecules fo genome length was examined using 9 DNA+temperature-sensitive (ts) mutants of pseudorabies virus, each belonging to a different complementation group. At the nonpermissive temperature, cells [rabbit kidney] infected with each of the mutants synthesized concatemeric DNA. Cleavage of the concatemeric DNA to genome-length viral DNA was defective in all the DNA+ ts mutants tested, indicating that several viral gene products are involved in the DNA maturation process. In none of the ts mutant-infected cells were capsids with electron-dense cores (containing DNA) formed. Empty capdids with electron-translucent cores were formed in cells infected with 6 of the 9 ts mutants; in cells infected with 3 of the mutants, no capsid assembly occurred. Because these 3 mutants are deficient in maturation of DNA and in the assembly of viral capsids, maturation of viral DNA is apparently dependent on the assembly of capsids. In cells infected with 2 of the mutants (tsN and tsIE13), normal maturation of viral DNA occurred after shiftdown of the cells to the permissive temperature in the presence of cycloheximide, indicating that the ts proteins involved in DNA maturation became functional after shiftdown. Because cycloheximide reduces maturation of DNA in wild-type-infected cells but not in cells infected with these 2 mutants, a protein(s) necessary for the maturation of concatemeric DNA, which is present in limiting amounts during the normal course of infection, probably accumulated in the mutant-infected cells at the nonpermissive temperature. Concomitant with cleavage of concatemeric DNA, full capsids with electron-dense cores appeared after shiftdown of tsN-infected cells to the permissive temperature, indicating that there may be a correlation between maturation of DNA and formation of full capsids. The number of empty and full capsids (containing electron-dense cores) present in tsN-infected cells incubated at the nonpermissive temperature, as well as after shiftdown to the permissive temperature in the presence of cycloheximide, was determined by EM and by sedimentation analysis in sucrose gradients. After shiftdown to the permissive temperature in the presence of cycloheximide, the number of empty capsids present in tsN-infected cells decreased with a concomitant accumulation of full capsids, indicating that empty capsids are precursors to full capsids.