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Arrest of replication by mammalian DNA polymerases α and β caused by chromium‐DNA lesions
Author(s) -
Bridgewater Laura C.,
Manning Francis C. R.,
Patierno Steven R.
Publication year - 1998
Publication title -
molecular carcinogenesis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.254
H-Index - 97
eISSN - 1098-2744
pISSN - 0899-1987
DOI - 10.1002/(sici)1098-2744(199812)23:4<201::aid-mc2>3.0.co;2-6
Subject(s) - dna polymerase , dna polymerase ii , dna polymerase i , biology , polymerase , dna polymerase delta , dna clamp , microbiology and biotechnology , dna replication , dna , biochemistry , polymerase chain reaction , gene , reverse transcriptase
We have previously shown that trivalent chromium, and hexavalent chromium in the presence of one of its primary in vivo reductants, ascorbate, can bind to DNA and form interstrand crosslinks capable of obstructing replication. This effect was demonstrated in vitro by using Sequenase Version 2.0 T7 DNA polymerase; its parent enzyme, the unmodified T7 DNA polymerase; and Escherichia coli polymerase I large (Klenow) fragment; and it was demonstrated ex vivo by using Taq polymerase and DNA from chromium‐treated human lung cells as template. This study was performed to determine whether DNA‐bound chromium affects mammalian DNA polymerases in the same manner. Two mammalian enzymes, DNA polymerase α and DNA polymerase β, were used. DNA polymerase α is a processive enzyme believed to be the primary lagging‐stand synthetase, whereas DNA polymerase β is a non‐processive enzyme believed to function in DNA repair by filling single stranded gaps one base at a time. DNA polymerase arrest assays were performed with each of these enzymes to replicate DNA with toxicologically relevant levels of chromium adducts produced by either trivalent chromium or hexavalent chromium and ascorbate. Both enzymes responded to chromium‐DNA damage by arresting replication, and the arrests increased in a dose‐dependent manner. Furthermore, the guanine‐specific pattern of arrests produced when an exonuclease‐free preparation of DNA polymerase β was used corresponded exactly to the arrest patterns produced in vitro by the exonuclease‐free enzyme Sequenase and ex vivo by Taq polymerase. These results suggest that replication arrest may be a common response of polymerases to DNA‐chromium lesions and provide a plausible mechanism for the inhibition of DNA synthesis and S‐phase cell‐cycle delay that occurs in mammalian cells treated with genotoxic chromium compounds. Mol. Carcinog. 23:201–206, 1998. © 1998 Wiley‐Liss, Inc.