Scientists Reveal A Mechanism of Efficient and Accurate Translesion Synthesis Past a BPDE-dG lesion
Benzo[a]pyrene (BP) is one of the common environmental pollutants generated during incomplete fuel combustion, in tobacco smoke, and in cooked food. BP can be metabolized in vivo to form the most tumorigenic and mutagenic BPDE, which bind covalently with guanine residues in DNA.
The formation of BPDE-dG lesion has been closely associated with lung cancer. Translesion DNA synthesis (TLS) polymerases can directly incorporate nucleotide opposite and beyond a variety of replication-blocking lesions, thus avoiding replication fork collapse.
TLS can be classified into two categories: error-free TLS and error-prone TLS. Error-prone TLS is one of the fundamental mechanisms for genome mutagenesis. To date, fifteen translesion synthesis polymerases have been identified in mammals, and DNA polymerase κ (Polκ) is the only known DNA polymerase in mouse and human cells that can bypass a BPDE-dG lesion efficiently and accurately, thereby reducing mutation risk. Defective of POLK gene generate increased genome mutation rate associated with exposure to BP. However, it is not known how a BPDE-dG lesion is accommodated and how dCTP is selected and incorporated by Polκ.
Polκ can bypass the BPDE-dG adduct in an error-free manner. However, Dpo4, Polκ’s homolog in archaeal, bypass the same BPDE-dG lesion in an error-prone manner. By superimposing the structure of Polκ with Dpo4, the group led by Prof. GUO Caixia from Beijing Institute of Genomics, CAS, in collaboration with Dr. YANG Wei of NIH, USA, have predicted the structure gap between the active site and the little finger domain as well as the unique N-clasp in Polκ may account for the discrepancy.
By employing various techniques, including biochemistry, molecular biology, synthetic chemistry as well as molecular simulations, scientists revealed for the first time that the structure of gap determines the polymerase’s processivity and fidelity. The in vitro regulation of BPDE-dG bypass by the structural gap has also been verified through in vivo cell survival assays. Deletion of the first 51-residues (Δ51) of the N-clasp in Polκ not only leads to reduced polymerization activity on normal dG template, but complete loss of BPDE bypass activity, which uncovered the key role of the N-clasp in lesion bypass.
The study has revealed the molecular mechanism underlying the accurate bypass of BPDE-dG lesion by Polκ. With the increased environmental pollutions, the incidence of many major human diseases, such as cancer, neurodegenerative diseases, cardiovascular disease, etc. have increased significantly. This study has important implications with regard to further our understanding of the molecular mechanisms associated with major diseases induced from common environmental pollutants, as well as developing preventive measures and treatments for human diseases.
The article has been published online in Proceedings of the National Academy of Sciences, USA (PNAS) in Jan, 2014.The coauthors include first author Dr. LIU Yang, and Dr. TANG Tie-Shan from State Key Laboratory of Biomembrane and Membrane Biotechnology, Institute of Zoology, CAS, etc. The work is supported by CAS, Ministry of Science and Technology and Natural Science Foundation of China.
The normal DNA synthetic and TLS activity by Polκ and predicted structure model of Polκ active site
(Image by GUO's group)
