Cisplatin-induced ubiquitination of RNA polymerase II large subunit and suppression of induction by 7-hydroxystaurosporine (UCN-01)

  • Authors:
    • Li-Ying Yang
    • Hong Jiang
    • Kelly M. Rangel
    • William Plunkett
  • View Affiliations

  • Published online on: September 1, 2003     https://doi.org/10.3892/or.10.5.1489
  • Pages: 1489-1495
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Abstract

Exposure of cells to DNA-damaging agents induces hyperphosphorylation of the C-terminal domain (CTD) of mammalian RNA polymerase II (RNAP II) large subunit (LS); the hyperphosphorylated RNAP II is then ubiquitinated. The purpose of this study was to verify that cisplatin-induced RNAP II ubiquitination is transcription dependent in living cells and to determine whether 7-hydroxystaurosporine (UCN-01) inhibits the ubiquitination induced by cisplatin. Cisplatin at clinically achievable concentrations (2.5-10 µM) induced the ubiquitination of RNAP II in exponentially growing A2780 human ovarian tumor cells; the effect was drug-dose and exposure-time dependent. Such induction, however, was not observed in colcemid-selected mitotic cells. When detergent extraction was applied, the ubiquitinated RNAP II was recovered in the detergent-insoluble fraction, indicating that the protein was tightly bound to DNA. In an in vitro transcription reaction that consists of nuclear extracts and an immobilized DNA template containing a site-specific cisplatin lesion, the elongating RNAP II that was stalled at a cisplatin lesion site on the template was targeted by ubiquitins. Together, our results indicate that the ubiquitination is associated with transcription-coupled repair. We previously showed that the Ser/The kinase-inhibitor UCN-01 inhibits nucleotide excision repair. Here, we further determined the effect of UCN-01 on the phosphorylation and ubiquitination of RNAP II LS in a whole-cell system. Immunoblotting results showed that UCN-01 suppressed the cisplatin-induced ubiquitination and the cisplatin-induced shift from the hypophosphorylated IIa to the hyperphosphorylated IIo, without affecting the basal levels of the IIo and IIa forms of the RNAP II CTD, suggesting that UCN-01 acts by suppressing cisplatin-mediated induction of the one or more kinases that is responsible for the conversion of the IIo that is important for ubiquitination.

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September-October 2003
Volume 10 Issue 5

Print ISSN: 1021-335X
Online ISSN:1791-2431

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Spandidos Publications style
Yang L, Jiang H, Rangel KM and Plunkett W: Cisplatin-induced ubiquitination of RNA polymerase II large subunit and suppression of induction by 7-hydroxystaurosporine (UCN-01). Oncol Rep 10: 1489-1495, 2003
APA
Yang, L., Jiang, H., Rangel, K.M., & Plunkett, W. (2003). Cisplatin-induced ubiquitination of RNA polymerase II large subunit and suppression of induction by 7-hydroxystaurosporine (UCN-01). Oncology Reports, 10, 1489-1495. https://doi.org/10.3892/or.10.5.1489
MLA
Yang, L., Jiang, H., Rangel, K. M., Plunkett, W."Cisplatin-induced ubiquitination of RNA polymerase II large subunit and suppression of induction by 7-hydroxystaurosporine (UCN-01)". Oncology Reports 10.5 (2003): 1489-1495.
Chicago
Yang, L., Jiang, H., Rangel, K. M., Plunkett, W."Cisplatin-induced ubiquitination of RNA polymerase II large subunit and suppression of induction by 7-hydroxystaurosporine (UCN-01)". Oncology Reports 10, no. 5 (2003): 1489-1495. https://doi.org/10.3892/or.10.5.1489