Homologous Recombination and Translesion DNA Synthesis Play Critical Roles on Tolerating DNA Damage Caused by Trace Levels of Hexavalent Chromium.

Xu Tian,Dayna Neo,Fred A Wright,Yi-Hui Zhou,Jun Nakamura,Youjun Chen,Keyur Patel,Shunichi Takeda,Minoru Takata,Jean Clement,James A Swenberg,Julian Sale,John R Ridpath,Kerstin Borgmann

doi:10.1371/journal.pone.0167503

Abstract

Contamination of potentially carcinogenic hexavalent chromium (Cr(VI)) in the drinking water is a major public health concern worldwide. However, little information is available regarding the biological effects of a nanomoler amount of Cr(VI). Here, we investigated the genotoxic effects of Cr(VI) at nanomoler levels and their repair pathways. We found that DNA damage response analyzed based on differential toxicity of isogenic cells deficient in various DNA repair proteins is observed after a three-day incubation with K2CrO4 in REV1-deficient DT40 cells at 19.2 μg/L or higher as well as in TK6 cells deficient in polymerase delta subunit 3 (POLD3) at 9.8 μg/L or higher. The genotoxicity of Cr(VI) decreased ~3000 times when the incubation time was reduced from three days to ten minutes. TK mutation rate also significantly decreased from 6 day to 1 day exposure to Cr(VI). The DNA damage response analysis suggest that DNA repair pathways, including the homologous recombination and REV1- and POLD3-mediated error-prone translesion synthesis pathways, are critical for the cells to tolerate to DNA damage caused by trace amount of Cr(VI).

Highlights

Chromium (Cr) is a naturally occurring element that exists in a variety of oxidation states between -2 to +6
Chicken DT40 cells deficient in either homologous recombination or the error-prone translesion synthesis pathway are hyper-sensitive to Cr(VI)
At the dose of 100 μg Cr/L, which is the maximum contaminant levels (MCL) for total Cr in drinking water as set by the U.S EPA, DT40 cells showed substantial toxicity with ~50% survival rate compared with vehicle-treated cells

Summary

Introduction

Chromium (Cr) is a naturally occurring element that exists in a variety of oxidation states between -2 to +6. Nucleotide excision repair [19] and Fanconi anemia genes [20] were previously reported to be essential for the removal of Cr(VI)-induced DNA damage or activated in human cells exposed with Cr(VI). In Saccharomyces cerevisiae and HR repair pathways were reported to be involved into the removal of Cr(VI) induced DNA damage in mammalian cells[23]. To further investigate the mechanism of Cr(VI) induced DNA damage and repair pathways, in this study we screened a battery of DT40 mutant cells using genotoxicity profiling and found the DNA repair genes and pathways to be critical for cell survival when these cells were exposed to Cr (VI) at nanomolar concentrations. We confirmed our results using human cancer knockdown cells and further studied the time- and dose- dependent genotoxicity and mutagenicity of nanomolar concentrations of Cr(VI)

Materials and Methods

Results

Discussion