T-ANTIGEN OF SV40 BLOCKS P53 TRANSACTIVATION BUT NOT P53 SPECIFIC BINDING TO DNA

  • Authors:
    • Y IMAI
    • AM CHUMAKOV
    • CW MILLER
    • J SAID
    • HP KOEFFLER
  • View Affiliations

  • Published online on: October 1, 1994     https://doi.org/10.3892/ijo.5.4.945
  • Pages: 945-953
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Abstract

The p53 is a DNA binding phosphoprotein that can act as a transcriptional activator through high affinity DNA binding sequences (HBS). The large T antigen (LT-ag) of SV40 virus can bind p53 and their association is considered important for transforming activities of the virus. In this study, we investigated the effects of LT-ag on transcriptional transactivating function of p53 using cotransfection assays and DNA-binding electrophoretic mobility shift assays. A reporter gene containing a minimal TK promoter and two copies of HBS for p53 was cotransfected with p53 and LT-ag expression vector into human SKOV3 cells (p53 non-expressor). The LT-ag inhibited in a dose-dependent fashion transactivation by wild-type p53. The LT-ag was unable to inhibit transactivation of a reporter gene containing a similar promoter (TK). The LT-ag mutants defective for binding to p53, failed to inhibit transactivation. The LT-ag inhibited the transactivation of a CAT reporter gene containing the GAL4-DNA recognition sequences by the p53 protein which was fused to the heterologous DNA binding domain (amino acids 1-147 of GAL4) in cotransfected cells showing that inhibition of p53 activities by LT-ag was not restricted to the p53 HBS-dependent reporter. LT-ag failed to inhibit GAL4-p53 fragment containing the transactivating, but non-LT-ag binding region of p53, showing the importance of LT-ag binding to p53 in order to restrict p53 transactivation. Immunohistochemical analysis showed that in SKOV3, nuclear localization of wild type p53 was unaffected by coexpressed LT-ag. Gel shift analysis determined that nuclear extract from cells cotransfected with p53 and LT-ag expression vectors contained p53 not associated with LT-ag; this free p53 was able to bind to the HBS. These results suggest that LT-ag of SV40 preferentially binds the transcriptionally active p53, preventing it from transactivating through p53-HBS; the transcriptionally inactive p53 in these cells can still bind p53-HBS.

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October 1994
Volume 5 Issue 4

Print ISSN: 1019-6439
Online ISSN:1791-2423

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APA
IMAI, Y., CHUMAKOV, A., MILLER, C., SAID, J., & KOEFFLER, H. (1994). T-ANTIGEN OF SV40 BLOCKS P53 TRANSACTIVATION BUT NOT P53 SPECIFIC BINDING TO DNA. International Journal of Oncology, 5, 945-953. https://doi.org/10.3892/ijo.5.4.945
MLA
IMAI, Y., CHUMAKOV, A., MILLER, C., SAID, J., KOEFFLER, H."T-ANTIGEN OF SV40 BLOCKS P53 TRANSACTIVATION BUT NOT P53 SPECIFIC BINDING TO DNA". International Journal of Oncology 5.4 (1994): 945-953.
Chicago
IMAI, Y., CHUMAKOV, A., MILLER, C., SAID, J., KOEFFLER, H."T-ANTIGEN OF SV40 BLOCKS P53 TRANSACTIVATION BUT NOT P53 SPECIFIC BINDING TO DNA". International Journal of Oncology 5, no. 4 (1994): 945-953. https://doi.org/10.3892/ijo.5.4.945