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Open Access Short Report

Regulation of PCNA polyubiquitination in human cells

Jan Brun123, Roland K Chiu4, Bradly G Wouters5 and Douglas A Gray12*

Author Affiliations

1 Ottawa Health Research Institute, Ottawa, ON K1H 8L6, Canada

2 Department of Biochemistry, Microbiology, and Immunology, University of Ottawa, Ottawa, Ontario, Canada

3 Apoptosis Research Centre, Children's Hospital of Eastern Ontario, Ottawa, Ontario K1H 8L1, Canada

4 Department of Health Risk and Toxicology, University of Maastricht, 6200MD Maastricht, the Netherlands

5 Ontario Cancer Insitute, Princess Margaret Hospital, Toronto, Ontario M5G 2MG, Canada

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BMC Research Notes 2010, 3:85  doi:10.1186/1756-0500-3-85

Published: 30 March 2010

Abstract

Background

The ubiquitin-based molecular switch dictating error free versus error prone repair has been conserved throughout eukaryotic evolution. A central component of this switch is the homotrimeric clamp PCNA, which is ubiquitinated in response to genotoxic stress allowing recovery of replication forks blocked at sites of DNA damage. The particulars of PCNA ubiquitination have been elucidated in yeast and to a further extent recently in human cells. However, gaps in the detailed mechanism and regulation of PCNA polyubiquitination still persist in human cells.

Findings

We expand upon several studies and show that PCNA is polyubiquitnated in normal skin fibroblasts, and that this ubiquitination is dependant on RAD18. Furthermore we define the types of DNA damage that induce ubiquitination on PCNA. Cisplatin, methylmethane sulphonate and benzo(a)pyrene-diol-epoxide induce the polyubiquitination of PCNA to the same extent as UV while polyubiquitination is not detected after X-ray treatment. Moreover, we show that ubiquitination of PCNA is not regulated by cell cycle checkpoint kinases ATM-Chk2 or ATR-Chk1. Significantly, we report that PCNA polyubiquitination is negatively regulated by USP1.

Conclusions

Our results demonstrate the importance of PCNA polyubiquitination in human cells and define the key regulator of this ubiquitination.