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. 1995 Jul;69(7):4548–4551. doi: 10.1128/jvi.69.7.4548-4551.1995

The baculovirus transactivator IE1 binds to viral enhancer elements in the absence of insect cell factors.

J Choi 1, L A Guarino 1
PMCID: PMC189203  PMID: 7769721

Abstract

The transregulatory IE1 protein of Autographa californica nuclear polyhedrosis virus binds to the viral enhancer element hr5. To test whether IE1 binds independently of host cell factors, IE1 was translated in rabbit reticulocyte extracts and tested for DNA binding activity by an electrophoretic mobility shift assay. Complexes with the hr5 probe were detected with translation reaction mixtures primed with ie1 RNA but not with control translation reaction mixtures. However, the DNA-protein complexes formed with IE1 translated in vitro migrated more slowly than complexes formed with IE1 that was transiently expressed in insect cells. Phosphatase treatment of the translation reactions resulted in an increase in the mobility of the DNA-protein complexes, suggesting that hyperphosphorylation was responsible for the altered migration. To further verify that IE1 was capable of binding DNA in the absence of host cell factors, an N-terminal truncation of IE1 was synthesized in vitro, and shown to interact with hr5. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of IE1 translated in vitro revealed that the mobility of the protein was heterogeneous. This pattern was altered by translation in the presence of an oligonucleotide corresponding to the IE1 specific binding site but was not affected by translation in the presence of a nonspecific DNA. These results suggest that binding of IE1 to DNA causes a conformational change in the protein that alters the accessibility of IE1 to protein kinases.

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Selected References

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  1. Ayres M. D., Howard S. C., Kuzio J., Lopez-Ferber M., Possee R. D. The complete DNA sequence of Autographa californica nuclear polyhedrosis virus. Virology. 1994 Aug 1;202(2):586–605. doi: 10.1006/viro.1994.1380. [DOI] [PubMed] [Google Scholar]
  2. Carson D. D., Summers M. D., Guarino L. A. Transient expression of the Autographa californica nuclear polyhedrosis virus immediate-early gene, IE-N, is regulated by three viral elements. J Virol. 1991 Feb;65(2):945–951. doi: 10.1128/jvi.65.2.945-951.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Chisholm G. E., Henner D. J. Multiple early transcripts and splicing of the Autographa californica nuclear polyhedrosis virus IE-1 gene. J Virol. 1988 Sep;62(9):3193–3200. doi: 10.1128/jvi.62.9.3193-3200.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Grässer F. A., Mann K., Walter G. Removal of serine phosphates from simian virus 40 large T antigen increases its ability to stimulate DNA replication in vitro but has no effect on ATPase and DNA binding. J Virol. 1987 Nov;61(11):3373–3380. doi: 10.1128/jvi.61.11.3373-3380.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Guarino L. A., Dong W. Expression of an enhancer-binding protein in insect cells transfected with the Autographa californica nuclear polyhedrosis virus IE1 gene. J Virol. 1991 Jul;65(7):3676–3680. doi: 10.1128/jvi.65.7.3676-3680.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Guarino L. A., Dong W. Functional dissection of the Autographa california nuclear polyhedrosis virus enhancer element hr5. Virology. 1994 May 1;200(2):328–335. doi: 10.1006/viro.1994.1197. [DOI] [PubMed] [Google Scholar]
  7. Guarino L. A., Gonzalez M. A., Summers M. D. Complete Sequence and Enhancer Function of the Homologous DNA Regions of Autographa californica Nuclear Polyhedrosis Virus. J Virol. 1986 Oct;60(1):224–229. doi: 10.1128/jvi.60.1.224-229.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Guarino L. A., Summers M. D. Functional mapping of a trans-activating gene required for expression of a baculovirus delayed-early gene. J Virol. 1986 Feb;57(2):563–571. doi: 10.1128/jvi.57.2.563-571.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Guarino L. A., Summers M. D. Interspersed Homologous DNA of Autographa californica Nuclear Polyhedrosis Virus Enhances Delayed-Early Gene Expression. J Virol. 1986 Oct;60(1):215–223. doi: 10.1128/jvi.60.1.215-223.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Guarino L. A., Summers M. D. Nucleotide sequence and temporal expression of a baculovirus regulatory gene. J Virol. 1987 Jul;61(7):2091–2099. doi: 10.1128/jvi.61.7.2091-2099.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kool M., van den Berg P. M., Tramper J., Goldbach R. W., Vlak J. M. Location of two putative origins of DNA replication of Autographa californica nuclear polyhedrosis virus. Virology. 1993 Jan;192(1):94–101. doi: 10.1006/viro.1993.1011. [DOI] [PubMed] [Google Scholar]
  12. Kovacs G. R., Choi J., Guarino L. A., Summers M. D. Functional dissection of the Autographa californica nuclear polyhedrosis virus immediate-early 1 transcriptional regulatory protein. J Virol. 1992 Dec;66(12):7429–7437. doi: 10.1128/jvi.66.12.7429-7437.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kovacs G. R., Guarino L. A., Graham B. L., Summers M. D. Identification of spliced baculovirus RNAs expressed late in infection. Virology. 1991 Dec;185(2):633–643. doi: 10.1016/0042-6822(91)90534-i. [DOI] [PubMed] [Google Scholar]
  14. Kovacs G. R., Guarino L. A., Summers M. D. Novel regulatory properties of the IE1 and IE0 transactivators encoded by the baculovirus Autographa californica multicapsid nuclear polyhedrosis virus. J Virol. 1991 Oct;65(10):5281–5288. doi: 10.1128/jvi.65.10.5281-5288.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Passarelli A. L., Miller L. K. Three baculovirus genes involved in late and very late gene expression: ie-1, ie-n, and lef-2. J Virol. 1993 Apr;67(4):2149–2158. doi: 10.1128/jvi.67.4.2149-2158.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Pearson M., Bjornson R., Pearson G., Rohrmann G. The Autographa californica baculovirus genome: evidence for multiple replication origins. Science. 1992 Sep 4;257(5075):1382–1384. doi: 10.1126/science.1529337. [DOI] [PubMed] [Google Scholar]
  17. Prives C. The replication functions of SV40 T antigen are regulated by phosphorylation. Cell. 1990 Jun 1;61(5):735–738. doi: 10.1016/0092-8674(90)90179-i. [DOI] [PubMed] [Google Scholar]
  18. Ribeiro B. M., Hutchinson K., Miller L. K. A mutant baculovirus with a temperature-sensitive IE-1 transregulatory protein. J Virol. 1994 Feb;68(2):1075–1084. doi: 10.1128/jvi.68.2.1075-1084.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Rodems S. M., Friesen P. D. The hr5 transcriptional enhancer stimulates early expression from the Autographa californica nuclear polyhedrosis virus genome but is not required for virus replication. J Virol. 1993 Oct;67(10):5776–5785. doi: 10.1128/jvi.67.10.5776-5785.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Theilmann D. A., Stewart S. Identification and characterization of the IE-1 gene of Orgyia pseudotsugata multicapsid nuclear polyhedrosis virus. Virology. 1991 Feb;180(2):492–508. doi: 10.1016/0042-6822(91)90063-h. [DOI] [PubMed] [Google Scholar]
  21. Wang E. H., Bhattacharyya S., Prives C. The replication functions of polyomavirus large tumor antigen are regulated by phosphorylation. J Virol. 1993 Nov;67(11):6788–6796. doi: 10.1128/jvi.67.11.6788-6796.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Yoo S., Guarino L. A. The Autographa californica nuclear polyhedrosis virus ie2 gene encodes a transcriptional regulator. Virology. 1994 Aug 1;202(2):746–753. doi: 10.1006/viro.1994.1396. [DOI] [PubMed] [Google Scholar]

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