Herpes simplex virus 1 alkaline nuclease is required for efficient egress of capsids from the nucleus.
We previously described the isolation of AN-1, a null mutant of the HSV-1 alkaline nuclease gene, which is able to synthesize near wild-type levels of viral DNA and late viral proteins under nonpermissive growth conditions; these results lead us to conclude that the alkaline nuclease is not essential for viral DNA synthesis (Weller, S. K., Seghatoleslami, R. M., Shao, L., Rowse, D., and Carmichael, E. P., 1990, J. Gen. Virol. 71, 2941-2952). AN-1 was found to be deficient in the production of infectious virions suggesting that the nuclease may play a role in processing or packaging of viral DNA into infectious virions. In this report we demonstrate that the defect is distinct from that observed in other late HSV-1 mutants which make but fail to process viral DNA under nonpermissive growth conditions. Following restriction enzyme digestion, specific terminal fragments are observed in DNA from AN-1-infected Vero cells, indicating that specific cleavage has occurred; moreover, the efficiency of cleavage is at near wild-type levels. Also in contrast to cells infected with previously described late mutants, DNase I or staphylococcal nuclease resistant DNA is observed in these cells further indicating that encapsidation has occurred. Three lines of evidence suggest, however, that maturation of DNA-containing AN-1 capsids is defective in some ways. First, in contrast to wild-type, very small amounts of protected DNA is detected in cytoplasmic extracts from AN-1-infected cells. Second, very few if any mature, DNA-containing C capsids are observed in the cytoplasm when analyzed by electron microscopy or sucrose gradient sedimentation. Finally, analysis of nuclei by sucrose gradient sedimentation indicates an elevated ratio of A to B capsids. These data indicate that AN-1 may be defective for the production of capsids competent to mature into the cytoplasm. Possible models for the nature of the defect in AN-1 will be discussed.
Department of Microbiology, University of Connecticut Health Center, Farmington 06030.
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D000818:Animals; D015139:Blotting, Southern; D002213:Capsid / Q000378:metabolism*; D002467:Cell Nucleus / Q000382:microbiology*; D004279:DNA, Viral / Q000378:metabolism; D005784:Gene Amplification; D005816:Genetic Complementation Test; D006863:Hydrogen-Ion Concentration; D015151:Immunoblotting; D009154:Mutation; D015183:Restriction Mapping; D012260:Ribonucleases / Q000235:genetics / Q000378:metabolism*; D018139:Simplexvirus / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000648:ultrastructure; D014709:Vero Cells
D004279:DNA, Viral; D012260:Ribonucleases; C066964:viral alkaline nuclease