During radiation therapy of human tumors the majority of DNA lesions originate from the radiolysis of water in the vicinity of the DNA molecule. These lesions can block DNA replication, which ultimately can lead to cell death and therapeutic efficacy. Alternatively, DNA damage can be bypassed resulting in mutations and the possibility of secondary tumors at the site of treatment by ionizing radiation. Thus it is the interaction between a DNA polymerase and a DNA lesion that ultimately determines the fate of irradiated cells. In order to maximize therapeutic gain and minimize the risk of secondary tumors at the treatment site it is imperative to have a solid mechanistic understanding of the interactions between a human DNA polymerase and a radiation-induced DNA lesion. We propose to address this central issue by studying two types of human DNA polymerases, replicative and specialized, in the context of ionization-induced lesions such as thymine glycol and abasic sites. DNA polymerase ? is a high-fidelity enzyme that plays a crucial role in DNA replication and repair. Pol ? is only somewhat proficient at inserting and extending past abasic sites and is unable to bypass Tg. In contrast, two specialized low-fidelity enzymes, DNA polymerases ? and ?, are much more efficient at extending past these DNA lesions generated by ionizing radiation. We will use a combination of biochemical and structural methods to uncover the mechanisms underpinning replication block or lesion bypass in the context of a replicative or bypass polymerase, with the ultimate goal to design compounds that inhibit pol ?, a polymerase that is overexpressed in breast cancer tumors.
DNA polymerase is one of the three human replicative DNA polymerase. Beyond its role in DNA replication it also plays a crucial role in DNA repair. The high fidelity of this replicative DNA polymerase is paramount for the maintenance of genome stability and for the avoidance of carcinogenesis. Pol ? and ? are two specialized human DNA polymerases; which are known to bypass ionization-generated lesions such as abasic sites or thymine glycols. Importantly pol ? is overexpressed in breast cancer tumors and knockdown of its gene makes tumor cells more sensitive to radiation; with little effect to normal tissues. The long-term goal of this proposal is to develop inhibitors of pol ? to aid in the radiosensitization of tumor cells.
|Lubula, Mulu Y; Poplawaski, Amanda; Glass, Karen C (2014) Crystallization and preliminary X-ray diffraction analysis of the BRPF1 bromodomain in complex with its H2AK5ac and H4K12ac histone-peptide ligands. Acta Crystallogr F Struct Biol Commun 70:1389-93|
|Eckenroth, Brian E; Fleming, Aaron M; Sweasy, Joann B et al. (2014) Crystal structure of DNA polymerase ? with DNA containing the base lesion spiroiminodihydantoin in a templating position. Biochemistry 53:2075-7|
|Yousefzadeh, Matthew J; Wyatt, David W; Takata, Kei-Ichi et al. (2014) Mechanism of suppression of chromosomal instability by DNA polymerase POLQ. PLoS Genet 10:e1004654|
|Doublié, Sylvie; Zahn, Karl E (2014) Structural insights into eukaryotic DNA replication. Front Microbiol 5:444|
|Zahn, Karl E; Doublié, Sylvie (2014) Look Ma, no PCNA: how DNA polymerase ? synthesizes long stretches of DNA without a processivity factor. Nat Struct Mol Biol 21:12-4|
|Wallace, Susan S (2013) DNA glycosylases search for and remove oxidized DNA bases. Environ Mol Mutagen 54:691-704|
|Zahn, Karl E; Wallace, Susan S; Doublié, Sylvie (2011) DNA polymerases provide a canon of strategies for translesion synthesis past oxidatively generated lesions. Curr Opin Struct Biol 21:358-69|
|Zahn, Karl E; Averill, April; Wallace, Susan S et al. (2011) The miscoding potential of 5-hydroxycytosine arises due to template instability in the replicative polymerase active site. Biochemistry 50:10350-8|
|Aller, Pierre; Duclos, Stéphanie; Wallace, Susan S et al. (2011) A crystallographic study of the role of sequence context in thymine glycol bypass by a replicative DNA polymerase serendipitously sheds light on the exonuclease complex. J Mol Biol 412:22-34|
|Zahn, Karl E; Tchesnokov, Egor P; Gotte, Matthias et al. (2011) Phosphonoformic acid inhibits viral replication by trapping the closed form of the DNA polymerase. J Biol Chem 286:25246-55|
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