Environmental insults can lead to DNA base damages and DNA strand breaks, outcomes that have the potential to destabilize the genome if left unrepaired. In order to avoid genome instability and cancer, DNA polymerases play a central role in accurately replicating the genome and repairing damaged DNA. DNA Polymerase e (Pol E) specifically plays important roles not only in replication and repair, but is unique among DNA polymerases in its involvement in coordinating replication progression and mitosis through its checkpoint function. The important, yet not precisely defined roles played by Pol e in replication and repair make the study of its fidelity crucial to understanding how human cells are able to avoid the consequences of DNA damage that, if left unrepaired, can lead to mutagenesis and diseases like cancer. Given its importance, the lack of knowledge of the fidelity of human Pol e stands in sharp contrast to the growing amount of information on the fidelity of the yeast enzyme. One important tool to determine the contribution of a DNA polymerase to genome stability is to identify and characterize mutant derivatives of DNA polymerases that decrease overall DNA synthesis fidelity. This project proposes to identify and characterize mutant alleles of human Pol e that suppress Pol e checkpoint activation in vivo through decreased replication fidelity, altered lesion bypass efficiency and fidelity, or some combination of these activities. In addition, these alleles will be used to probe for Pol e function in replication and repair in vivo. Both yeast and human Pol e have been identified in multi-protein complexes involved in replication, transcription and recombination, and have been observed to colocalize in vivo with regions of active replication. These interactions remain poorly characterized and not well understood, especially for the human enzyme. This project proposes to determine the Pol ? protein interaction network by using recombinant Pol e to probe for Pol e interaction partners and characterize the functional significance of these interactions.

Public Health Relevance

Human Pol t plays an essential role in faithfully duplicating the entire human genome. Environmental agents that interfere with this role can result in mutagenesis and genome instability, and potentially lead to cancer. The proposed studies will provide a better understanding of how human Pol e protects against human disease like cancer, thus aiding the design of therapeutic strategies against these diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of Environmental Health Sciences (NIEHS)
Type
Research Transition Award (R00)
Project #
5R00ES016780-04
Application #
8118527
Study Section
Special Emphasis Panel (NSS)
Program Officer
Shaughnessy, Daniel
Project Start
2009-09-01
Project End
2014-01-31
Budget Start
2011-08-01
Budget End
2014-01-31
Support Year
4
Fiscal Year
2011
Total Cost
$248,816
Indirect Cost
Name
Tulane University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
053785812
City
New Orleans
State
LA
Country
United States
Zip Code
70118
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Shinbrot, Eve; Henninger, Erin E; Weinhold, Nils et al. (2014) Exonuclease mutations in DNA polymerase epsilon reveal replication strand specific mutation patterns and human origins of replication. Genome Res 24:1740-50
Agbor, Anderson Ayuk; Göksenin, A Yasemin; LeCompte, Kimberly G et al. (2013) Human Pol ?-dependent replication errors and the influence of mismatch repair on their correction. DNA Repair (Amst) 12:954-63
Göksenin, A Yasemin; Zahurancik, Walter; LeCompte, Kimberly G et al. (2012) Human DNA polymerase ? is able to efficiently extend from multiple consecutive ribonucleotides. J Biol Chem 287:42675-84
Korona, Dagmara A; Lecompte, Kimberly G; Pursell, Zachary F (2011) The high fidelity and unique error signature of human DNA polymerase epsilon. Nucleic Acids Res 39:1763-73