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. 1995 Jan;69(1):182–188. doi: 10.1128/jvi.69.1.182-188.1995

CCAAT box-dependent activation of the TATA-less human DNA polymerase alpha promoter by the human cytomegalovirus 72-kilodalton major immediate-early protein.

G P Hayhurst 1, L A Bryant 1, R C Caswell 1, S M Walker 1, J H Sinclair 1
PMCID: PMC188562  PMID: 7983709

Abstract

Human cytomegalovirus (HCMV) immediate-early (IE) proteins are known potent transregulators of viral and cellular gene expression upon HCMV infection. HCMV is known to activate a number of cellular genes intimately associated with the cell cycle and DNA replication by mechanisms involving the viral major IE 86-kDa protein (IE2). We have recently shown that IE2 mediates this activation in a TATA-dependent manner and interacts directly with the TATA-binding protein. However, a number of TATA-less cellular promoters, e.g., DNA polymerase alpha and dihydrofolate reductase, are also activated by HCMV infection. Consequently, we have asked how HCMV mediates this activation. We show that, consistent with its known TATA dependency, IE2 does not activate the DNA polymerase alpha promoter. In contrast, this promoter is strongly activated by the major IE 72-kDa protein (IE1). Whilst deletion of ATF or E2F sites within the DNA polymerase alpha promoter had little effect on IE1-mediated activation, removal of the CCAAT box appeared to abolish high levels of activation by IE1. Consistent with this observation, we also find that IE1 interacts directly with the CCAAT box binding factor CTF1 in vitro and massively augments CTF1-mediated activation of the DNA polymerase alpha promoter in transient transfection assays.

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Selected References

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  1. Arlt H., Lang D., Gebert S., Stamminger T. Identification of binding sites for the 86-kilodalton IE2 protein of human cytomegalovirus within an IE2-responsive viral early promoter. J Virol. 1994 Jul;68(7):4117–4125. doi: 10.1128/jvi.68.7.4117-4125.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Biegalke B. J., Geballe A. P. Sequence requirements for activation of the HIV-1 LTR by human cytomegalovirus. Virology. 1991 Jul;183(1):381–385. doi: 10.1016/0042-6822(91)90151-z. [DOI] [PubMed] [Google Scholar]
  3. Boldogh I., Fons M. P., Albrecht T. Increased levels of sequence-specific DNA-binding proteins in human cytomegalovirus-infected cells. Biochem Biophys Res Commun. 1993 Dec 30;197(3):1505–1510. doi: 10.1006/bbrc.1993.2647. [DOI] [PubMed] [Google Scholar]
  4. Boshart M., Weber F., Jahn G., Dorsch-Häsler K., Fleckenstein B., Schaffner W. A very strong enhancer is located upstream of an immediate early gene of human cytomegalovirus. Cell. 1985 Jun;41(2):521–530. doi: 10.1016/s0092-8674(85)80025-8. [DOI] [PubMed] [Google Scholar]
  5. Burns L. J., Waring J. F., Reuter J. J., Stinski M. F., Ginder G. D. Only the HLA class I gene minimal promoter elements are required for transactivation by human cytomegalovirus immediate early genes. Blood. 1993 Mar 15;81(6):1558–1566. [PubMed] [Google Scholar]
  6. Caswell R., Hagemeier C., Chiou C. J., Hayward G., Kouzarides T., Sinclair J. The human cytomegalovirus 86K immediate early (IE) 2 protein requires the basic region of the TATA-box binding protein (TBP) for binding, and interacts with TBP and transcription factor TFIIB via regions of IE2 required for transcriptional regulation. J Gen Virol. 1993 Dec;74(Pt 12):2691–2698. doi: 10.1099/0022-1317-74-12-2691. [DOI] [PubMed] [Google Scholar]
  7. Cherrington J. M., Mocarski E. S. Human cytomegalovirus ie1 transactivates the alpha promoter-enhancer via an 18-base-pair repeat element. J Virol. 1989 Mar;63(3):1435–1440. doi: 10.1128/jvi.63.3.1435-1440.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Colberg-Poley A. M., Santomenna L. D., Harlow P. P., Benfield P. A., Tenney D. J. Human cytomegalovirus US3 and UL36-38 immediate-early proteins regulate gene expression. J Virol. 1992 Jan;66(1):95–105. doi: 10.1128/jvi.66.1.95-105.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. DeMarchi J. M., Schmidt C. A., Kaplan A. S. Patterns of transcription of human cytomegalovirus in permissively infected cells. J Virol. 1980 Aug;35(2):277–286. doi: 10.1128/jvi.35.2.277-286.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Dudding L., Haskill S., Clark B. D., Auron P. E., Sporn S., Huang E. S. Cytomegalovirus infection stimulates expression of monocyte-associated mediator genes. J Immunol. 1989 Nov 15;143(10):3343–3352. [PubMed] [Google Scholar]
  11. Furnari B. A., Poma E., Kowalik T. F., Huong S. M., Huang E. S. Human cytomegalovirus immediate-early gene 2 protein interacts with itself and with several novel cellular proteins. J Virol. 1993 Aug;67(8):4981–4991. doi: 10.1128/jvi.67.8.4981-4991.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Geist L. J., Monick M. M., Stinski M. F., Hunninghake G. W. The immediate early genes of human cytomegalovirus upregulate tumor necrosis factor-alpha gene expression. J Clin Invest. 1994 Feb;93(2):474–478. doi: 10.1172/JCI116995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Ghazal P., Young J., Giulietti E., DeMattei C., Garcia J., Gaynor R., Stenberg R. M., Nelson J. A. A discrete cis element in the human immunodeficiency virus long terminal repeat mediates synergistic trans activation by cytomegalovirus immediate-early proteins. J Virol. 1991 Dec;65(12):6735–6742. doi: 10.1128/jvi.65.12.6735-6742.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hagemeier C., Walker S. M., Sissons P. J., Sinclair J. H. The 72K IE1 and 80K IE2 proteins of human cytomegalovirus independently trans-activate the c-fos, c-myc and hsp70 promoters via basal promoter elements. J Gen Virol. 1992 Sep;73(Pt 9):2385–2393. doi: 10.1099/0022-1317-73-9-2385. [DOI] [PubMed] [Google Scholar]
  15. Hagemeier C., Walker S., Caswell R., Kouzarides T., Sinclair J. The human cytomegalovirus 80-kilodalton but not the 72-kilodalton immediate-early protein transactivates heterologous promoters in a TATA box-dependent mechanism and interacts directly with TFIID. J Virol. 1992 Jul;66(7):4452–4456. doi: 10.1128/jvi.66.7.4452-4456.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hermiston T. W., Malone C. L., Witte P. R., Stinski M. F. Identification and characterization of the human cytomegalovirus immediate-early region 2 gene that stimulates gene expression from an inducible promoter. J Virol. 1987 Oct;61(10):3214–3221. doi: 10.1128/jvi.61.10.3214-3221.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hirai K., Watanabe Y. Induction of alpha type DNA polymerases in human cytomegalovirus-infected WI-38 cells. Biochim Biophys Acta. 1976 Oct 18;447(3):328–339. doi: 10.1016/0005-2787(76)90056-3. [DOI] [PubMed] [Google Scholar]
  18. Huang E. S. Human cytomegalovirus. III. Virus-induced DNA polymerase. J Virol. 1975 Aug;16(2):298–310. doi: 10.1128/jvi.16.2.298-310.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Huang L., Malone C. L., Stinski M. F. A human cytomegalovirus early promoter with upstream negative and positive cis-acting elements: IE2 negates the effect of the negative element, and NF-Y binds to the positive element. J Virol. 1994 Apr;68(4):2108–2117. doi: 10.1128/jvi.68.4.2108-2117.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Hunninghake G. W., Monks B. G., Geist L. J., Monick M. M., Monroy M. A., Stinski M. F., Webb A. C., Dayer J. M., Auron P. E., Fenton M. J. The functional importance of a cap site-proximal region of the human prointerleukin 1 beta gene is defined by viral protein trans-activation. Mol Cell Biol. 1992 Aug;12(8):3439–3448. doi: 10.1128/mcb.12.8.3439. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Jeor S. C., Albrecht T. B., Funk F. D., Rapp F. Stimulation of cellular DNA synthesis by human cytomegalovirus. J Virol. 1974 Feb;13(2):353–362. doi: 10.1128/jvi.13.2.353-362.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Jupp R., Hoffmann S., Stenberg R. M., Nelson J. A., Ghazal P. Human cytomegalovirus IE86 protein interacts with promoter-bound TATA-binding protein via a specific region distinct from the autorepression domain. J Virol. 1993 Dec;67(12):7539–7546. doi: 10.1128/jvi.67.12.7539-7546.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kaelin W. G., Jr, Krek W., Sellers W. R., DeCaprio J. A., Ajchenbaum F., Fuchs C. S., Chittenden T., Li Y., Farnham P. J., Blanar M. A. Expression cloning of a cDNA encoding a retinoblastoma-binding protein with E2F-like properties. Cell. 1992 Jul 24;70(2):351–364. doi: 10.1016/0092-8674(92)90108-o. [DOI] [PubMed] [Google Scholar]
  24. Kerry J. A., Priddy M. A., Stenberg R. M. Identification of sequence elements in the human cytomegalovirus DNA polymerase gene promoter required for activation by viral gene products. J Virol. 1994 Jul;68(7):4167–4176. doi: 10.1128/jvi.68.7.4167-4176.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Lafemina R. L., Pizzorno M. C., Mosca J. D., Hayward G. S. Expression of the acidic nuclear immediate-early protein (IE1) of human cytomegalovirus in stable cell lines and its preferential association with metaphase chromosomes. Virology. 1989 Oct;172(2):584–600. doi: 10.1016/0042-6822(89)90201-8. [DOI] [PubMed] [Google Scholar]
  26. Lang D., Stamminger T. The 86-kilodalton IE-2 protein of human cytomegalovirus is a sequence-specific DNA-binding protein that interacts directly with the negative autoregulatory response element located near the cap site of the IE-1/2 enhancer-promoter. J Virol. 1993 Jan;67(1):323–331. doi: 10.1128/jvi.67.1.323-331.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Lum L. S., Hsu S., Vaewhongs M., Wu B. The hsp70 gene CCAAT-binding factor mediates transcriptional activation by the adenovirus E1a protein. Mol Cell Biol. 1992 Jun;12(6):2599–2605. doi: 10.1128/mcb.12.6.2599. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Macias M. P., Stinski M. F. An in vitro system for human cytomegalovirus immediate early 2 protein (IE2)-mediated site-dependent repression of transcription and direct binding of IE2 to the major immediate early promoter. Proc Natl Acad Sci U S A. 1993 Jan 15;90(2):707–711. doi: 10.1073/pnas.90.2.707. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Malone C. L., Vesole D. H., Stinski M. F. Transactivation of a human cytomegalovirus early promoter by gene products from the immediate-early gene IE2 and augmentation by IE1: mutational analysis of the viral proteins. J Virol. 1990 Apr;64(4):1498–1506. doi: 10.1128/jvi.64.4.1498-1506.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Mermod N., O'Neill E. A., Kelly T. J., Tjian R. The proline-rich transcriptional activator of CTF/NF-I is distinct from the replication and DNA binding domain. Cell. 1989 Aug 25;58(4):741–753. doi: 10.1016/0092-8674(89)90108-6. [DOI] [PubMed] [Google Scholar]
  31. Pearson B. E., Nasheuer H. P., Wang T. S. Human DNA polymerase alpha gene: sequences controlling expression in cycling and serum-stimulated cells. Mol Cell Biol. 1991 Apr;11(4):2081–2095. doi: 10.1128/mcb.11.4.2081. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Pizzorno M. C., Mullen M. A., Chang Y. N., Hayward G. S. The functionally active IE2 immediate-early regulatory protein of human cytomegalovirus is an 80-kilodalton polypeptide that contains two distinct activator domains and a duplicated nuclear localization signal. J Virol. 1991 Jul;65(7):3839–3852. doi: 10.1128/jvi.65.7.3839-3852.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Pizzorno M. C., O'Hare P., Sha L., LaFemina R. L., Hayward G. S. trans-activation and autoregulation of gene expression by the immediate-early region 2 gene products of human cytomegalovirus. J Virol. 1988 Apr;62(4):1167–1179. doi: 10.1128/jvi.62.4.1167-1179.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Plachter B., Britt W., Vornhagen R., Stamminger T., Jahn G. Analysis of proteins encoded by IE regions 1 and 2 of human cytomegalovirus using monoclonal antibodies generated against recombinant antigens. Virology. 1993 Apr;193(2):642–652. doi: 10.1006/viro.1993.1172. [DOI] [PubMed] [Google Scholar]
  35. Sambucetti L. C., Cherrington J. M., Wilkinson G. W., Mocarski E. S. NF-kappa B activation of the cytomegalovirus enhancer is mediated by a viral transactivator and by T cell stimulation. EMBO J. 1989 Dec 20;8(13):4251–4258. doi: 10.1002/j.1460-2075.1989.tb08610.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Schwartz R., Sommer M. H., Scully A., Spector D. H. Site-specific binding of the human cytomegalovirus IE2 86-kilodalton protein to an early gene promoter. J Virol. 1994 Sep;68(9):5613–5622. doi: 10.1128/jvi.68.9.5613-5622.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Stenberg R. M., Fortney J., Barlow S. W., Magrane B. P., Nelson J. A., Ghazal P. Promoter-specific trans activation and repression by human cytomegalovirus immediate-early proteins involves common and unique protein domains. J Virol. 1990 Apr;64(4):1556–1565. doi: 10.1128/jvi.64.4.1556-1565.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Stenberg R. M., Witte P. R., Stinski M. F. Multiple spliced and unspliced transcripts from human cytomegalovirus immediate-early region 2 and evidence for a common initiation site within immediate-early region 1. J Virol. 1985 Dec;56(3):665–675. doi: 10.1128/jvi.56.3.665-675.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Stinski M. F., Thomsen D. R., Stenberg R. M., Goldstein L. C. Organization and expression of the immediate early genes of human cytomegalovirus. J Virol. 1983 Apr;46(1):1–14. doi: 10.1128/jvi.46.1.1-14.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Thomsen D. R., Stenberg R. M., Goins W. F., Stinski M. F. Promoter-regulatory region of the major immediate early gene of human cytomegalovirus. Proc Natl Acad Sci U S A. 1984 Feb;81(3):659–663. doi: 10.1073/pnas.81.3.659. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Wade M., Kowalik T. F., Mudryj M., Huang E. S., Azizkhan J. C. E2F mediates dihydrofolate reductase promoter activation and multiprotein complex formation in human cytomegalovirus infection. Mol Cell Biol. 1992 Oct;12(10):4364–4374. doi: 10.1128/mcb.12.10.4364. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Walker S., Hagemeier C., Sissons J. G., Sinclair J. H. A 10-base-pair element of the human immunodeficiency virus type 1 long terminal repeat (LTR) is an absolute requirement for transactivation by the human cytomegalovirus 72-kilodalton IE1 protein but can be compensated for by other LTR regions in transactivation by the 80-kilodalton IE2 protein. J Virol. 1992 Mar;66(3):1543–1550. doi: 10.1128/jvi.66.3.1543-1550.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Wathen M. W., Stinski M. F. Temporal patterns of human cytomegalovirus transcription: mapping the viral RNAs synthesized at immediate early, early, and late times after infection. J Virol. 1982 Feb;41(2):462–477. doi: 10.1128/jvi.41.2.462-477.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]

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