Skip to main content
NCBI home page
Journal of Virology logoLink to Journal of Virology
. 1980 Dec;36(3):816–828. doi: 10.1128/jvi.36.3.816-828.1980

Analysis of herpes simplex virus nucleoprotein complexes extracted from infected cells.

P F Pignatti, E Cassai
PMCID: PMC353709  PMID: 6257929

Abstract

HEp-2 cells were infected with herpes simplex virus type 1 and labeled with [3H]thymidine and 14C-amino acids. Infected cells or nuclei prepared from them were extracted with Triton X-100 and NaCl, utilizing a method recently described, and the low-speed supernatant (extract) was partially purified by sedimentation on sucrose gradients. A nucleoprotein complex which sedimented as a wide peak around 200S was identified. The nucleoprotein complex contained viral DNA, which banded at the expected density in CsCl isopycnic gradients and was intact after measurements taken on electron microscopic photographic enlargements. The autoradiographic pattern of 14C-labeled proteins after electrophoresis showed that only a few of the virus-specific polypeptides were present in the nucleoprotein complexes, in particular, VP5, VP12, VP15.2, VP19, and VP24. Cellular histones were absent. The extracts and the nucleoprotein complexes were centrifuged to equilibrium in metrizamide density gradients without prefixation. Electron microscopic direct visualization of the nucleoprotein complexes after sucrose or metrizamide purification revealed that the proteins were preferentially associated with one end of the DNA molecule and formed large irregular terminal thickenings or capsid-like transparent shells enclosing polyglobular cores. No nucleosomes were observed on herpes simplex virus nucleoprotein complexes. The same type of complex was detected after phosphonoacetic acid addition, and grossly altered nucleocapsids were formed.

Full text

PDF
816

Images in this article

820Image
on p.820
821Image
on p.821
822Image
on p.822
823Image
on p.823
825Image
on p.825
827Image
on p.827

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Birnie G. D., Rickwood D., Hell A. Buoyant densities and hydration of nucleic acids, proteins and nucleoprotein complexes in metrizamide. Biochim Biophys Acta. 1973 Dec 7;331(2):283–294. doi: 10.1016/0005-2787(73)90441-3. [DOI] [PubMed] [Google Scholar]
  2. Burgoyne L. A., Wagar M. A., Atkinson M. R. Calcium-dependent priming of DNA synthesis in isolated rat liver nuclei. Biochem Biophys Res Commun. 1970 Apr 24;39(2):254–259. doi: 10.1016/0006-291x(70)90786-2. [DOI] [PubMed] [Google Scholar]
  3. Cremisi C., Pignatti P. F., Croissant O., Yaniv M. Chromatin-like structures in polyoma virus and simian virus 10 lytic cycle. J Virol. 1975 Jan;17(1):204–211. doi: 10.1128/jvi.17.1.204-211.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Dubochet J., Ducommun M., Zollinger M., Kellenberger E. A new preparation method for dark-field electron microscopy of biomacromolecules. J Ultrastruct Res. 1971 Apr;35(1):147–167. doi: 10.1016/s0022-5320(71)80148-x. [DOI] [PubMed] [Google Scholar]
  5. Ejercito P. M., Kieff E. D., Roizman B. Characterization of herpes simplex virus strains differing in their effects on social behaviour of infected cells. J Gen Virol. 1968 May;2(3):357–364. doi: 10.1099/0022-1317-2-3-357. [DOI] [PubMed] [Google Scholar]
  6. Favre M., Breitburd F., Croissant O., Orth G. Chromatin-like structures obtained after alkaline disruption of bovine and human papillomaviruses. J Virol. 1977 Mar;21(3):1205–1209. doi: 10.1128/jvi.21.3.1205-1209.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Furlong D., Swift H., Roizman B. Arrangement of herpesvirus deoxyribonucleic acid in the core. J Virol. 1972 Nov;10(5):1071–1074. doi: 10.1128/jvi.10.5.1071-1074.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Gibson W., Roizman B. Proteins specified by herpes simplex virus. 8. Characterization and composition of multiple capsid forms of subtypes 1 and 2. J Virol. 1972 Nov;10(5):1044–1052. doi: 10.1128/jvi.10.5.1044-1052.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Griffith J. D. Chromatin structure: deduced from a minichromosome. Science. 1975 Mar 28;187(4182):1202–1203. doi: 10.1126/science.187.4182.1202. [DOI] [PubMed] [Google Scholar]
  10. Heine J. W., Honess R. W., Cassai E., Roizman B. Proteins specified by herpes simplex virus. XII. The virion polypeptides of type 1 strains. J Virol. 1974 Sep;14(3):640–651. doi: 10.1128/jvi.14.3.640-651.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Henry C. J. The core particles of herpes simplex virus. Can J Microbiol. 1974 May;20(5):731–734. doi: 10.1139/m74-110. [DOI] [PubMed] [Google Scholar]
  12. Honess R. W., Roizman B. Proteins specified by herpes simplex virus. XI. Identification and relative molar rates of synthesis of structural and nonstructural herpes virus polypeptides in the infected cell. J Virol. 1973 Dec;12(6):1347–1365. doi: 10.1128/jvi.12.6.1347-1365.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Houssais J. F. Séparation des particules ribonucléoprotéiques (Hn-RNP) des noyaux de cellules L, en deux classes, par centrifugation isopycnique en gradients de métrizamide. FEBS Lett. 1975 Aug 15;56(2):341–347. doi: 10.1016/0014-5793(75)81123-9. [DOI] [PubMed] [Google Scholar]
  14. Kedinger C., Brison O., Perrin F., Wilhelm J. Structural analysis of viral replicative intermediates isolated from adenovirus type 2-infected HeLa cell nuclei. J Virol. 1978 May;26(2):364–379. doi: 10.1128/jvi.26.2.364-379.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kieff E. D., Bachenheimer S. L., Roizman B. Size, composition, and structure of the deoxyribonucleic acid of herpes simplex virus subtypes 1 and 2. J Virol. 1971 Aug;8(2):125–132. doi: 10.1128/jvi.8.2.125-132.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Meneguzzi G., Pignatti P. F., Barbanti-Brodano G., Milanesi G. Minichromosome from BK virus as a template for transcription in vitro. Proc Natl Acad Sci U S A. 1978 Mar;75(3):1126–1130. doi: 10.1073/pnas.75.3.1126. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Nii S., Yasuda I. Detection of viral cores having toroid structures in eight herpes viruses. Biken J. 1975 Mar;18(1):41–46. [PubMed] [Google Scholar]
  18. Pignatti P. F., Cassai E., Bertazzoni U. Herpes simplex virus DNA synthesis in a partially purified soluble extract from infected cells. J Virol. 1979 Dec;32(3):1033–1036. doi: 10.1128/jvi.32.3.1033-1036.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Pignatti P. F., Cassai E., Meneguzzi G., Chenciner N., Milanesi G. Herpes simplex virus DNA isolation from infected cells with a novel procedure. Virology. 1979 Feb;93(1):260–264. doi: 10.1016/0042-6822(79)90295-2. [DOI] [PubMed] [Google Scholar]
  20. Pignatti P. F., Cremisi C., Croissant O., Yaniv M. SV 40 nucleoprotein complexes: structural modifications after isopycnic centrifugation in metrizamide gradients. FEBS Lett. 1975 Dec 15;60(2):369–373. doi: 10.1016/0014-5793(75)80751-4. [DOI] [PubMed] [Google Scholar]
  21. Wadsworth S., Jacob R. J., Roizman B. Anatomy of herpes simplex virus DNA. II. Size, composition, and arrangement of inverted terminal repetitions. J Virol. 1975 Jun;15(6):1487–1497. doi: 10.1128/jvi.15.6.1487-1497.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Wilhelm J., Brison O., Kedinger C., Chambon P. Characterization of adenovirus type 2 transcriptional complexes isolated from infected HeLa cell nuclei. J Virol. 1976 Jul;19(1):61–81. doi: 10.1128/jvi.19.1.61-81.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Virology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES