Discovery of a New Family of Mammalian DNA Glycosylases-orthologs of E.coli MutM/Nei

Three candidate proteins were identified and named NEIL (Nei-like) based on substrate preference and sequence homology with E. coli Nei/MutM.

A model for APE and PNK-dependent BER pathways in mammalian cells

Three BER pathways (I, II & III) defined by the type and reaction mechanism of DNA glycosylases are shown. Monofunctional glycosylases (M) generate AP sites which are cleaved by APE1 to leave a 5’-dRP terminus. It is removed by Pol β producing a single nucleotide gap for nucleotide addition (II). When NTH1 and OGG1 carry out elimination, APE1 removal of the resulting 3 dRP generates a single nucleotide gap with a 3’ OH (I). With NEI1s as the initial glycosylase, a 3’ phosphate terminus is generated, which is then removed by PNK (II).

NEIL2 co-localizes with Pol II, especially under oxidative stress. (Co-localization was visualized by superimposition of green and red images.)

Confocal image of NEIL2 and RNA pol II co-localization.

DNA Repair & Mutagenesis Core

DNA repair and mutagenesis includes research relating to the induction of DNA damage, cellular responses to DNA damage, and biological outcomes resulting from the combination of damage and cellular response. DNA is damaged continuously as a result of endogenous biochemical processes that modify DNA structure, or generate free radicals and reactive electrophiles, which can chemically modify DNA. Exposures to exogenous agents occurring naturally in the environment or result from human activity add to the burden of spontaneous DNA damage. Many carcinogens produce their toxic effects through mechanisms involving DNA damage. A critical goal of the environmental health sciences is to understand the mechanisms by which DNA-damaging agents produce adverse health effects, in order to accurately assess the risks posed by human exposure to them. It is becoming clear the cellular response to DNA damage is part of a complex web of regulatory activities that must be understood to appreciate the mechanisms maintaining cellular homeostasis or result in pathology or cell death. Within the scope of this goal, it is important to understand how genotoxic agents in the environment are absorbed and metabolized and how they damage DNA. It is also important to understand how cells respond to DNA damage repair processes, control of cell replication, and apoptosis, and how the resultant effects, including gene mutation and heritable chromosome changes, are induced. The modification of these effects by interindividual allelic variations in genes controlling these processes is becoming recognized as an important factor in human sensitivity to genotoxic agents.

Exposure to genotoxic agents is clearly associated with carcinogenic effects. Genotoxins are implicated in pathologies resulting in adverse reproductive outcomes and accelerated aging processes. The NIEHS Center at the UTMB is located in the geographic center of one of the world's heaviest concentrations of synthetic chemical manufacturing and use. The upper Texas Gulf coast and neighboring Louisiana coastal areas have some of the highest volumes of chemicals released to air and water in the US. Large numbers of hazardous waste sites and population of workers handling mutagenic chemicals, are present in this area, plus intense sunlight in this area results in significant exposures to UV light and a high incidence of skin cancer.

Research Highlights

Core Director and Members

Sankar Mitra, Ph.D., Director, DNA Repair and Mutagenesis Resarch Core, NIEHS Center , Professor, Department of Biochemistry and Molecular Biology

Sherif Abdel-Rahman, Ph.D., Member, DNA Repair and Mutagenesis Research Core, Professor, Department of Preventive Medicine and Community Health

Thomas B. Albrecht, Ph.D., Member, DNA Repair and Mutagenesis Research Core, Professor, Department of Microbiology and Immunology

Werner Braun, Ph.D., Member, DNA Repair and Mutagenesis Research Core, Professor, Department of Biochemistry and Molecular Biology

Wlodzimierz M. Bujalowski, Ph.D., Member, DNA Repair and Mutagenesis Research Core, Professor, Department of Biochemistry and Molecular Biology

David G. Gorenstein, Ph.D., Member, DNA Repair and Mutagenesis Research Core, Associate Dean of Research, Professor, Department of Biochemistry and Molecular Biology

Tapas Hazra, Ph.D., Member, DNA Repair and Mutagenesis Research Core, Assistant Professor, Department of Biochemistry and Molecular Biology

Louise Prakash, Ph.D., Member, DNA Repair and Mutagenesis Research Core, Professor, Department of Biochemistry and Molecular Biology

Satya Prakash, Ph.D., Member, DNA Repair and Mutagenesis Research Core, Professor, Department of Biochemistry and Molecular Biology

Jonathan B. Ward, Jr., Ph.D., Center Deputy Director and Co-Director, DNA Repair and Mutagenesis Research Core, Professor, Department of Preventive Medicine and Community Health

Key Words

DNA Polymerase, DNA Repair, DNA Repair Enzymes , DNA Replication, Genetic Biomonitoring, Genetic Susceptibility, Mutagenesis, Oxidative Stress, Structural Biology, Ultraviolet Light

2005 Publications

Deng CZ, Fons MP, Rosenblatt J, El-Zein RA, Abdel-Rahman SZ, Albrecht T.  2005 Dec 2. Nickel potentiates the genotoxic effect of benzo[a]pyrene in Chinese hamster lung V79 cells.Environ Mol Mutagen.  PMID: 16329104.

El-Zein RA, Abdel-Rahman SZ, Morris D, Hayes M, Bondy M, and Legator MS. 2005. Cytogenetic effects of Ritalin treatment in children. Cancer Lett. 230, 284-291.

El-Zein RA, Hastings-Smith DA, Ammenheuser MM,Treinen-Moslen M,Gulland FM and Ward Jr, JB. 2005. Evaluation of two different biomarkersfor use in the assessment of toxic chemical exposure in California Sea Lions(Zalophus Californianus). Marine Pollution Bulletin PMID: 16298399.

Galletto R, Jezewska MJ, Bujalowski W. 2005 Feb. Kinetic Mechanism of Rat Polymerase b - dsDNA Interactions. Fluorescence Stopped-Flow Analysis of the Cooperative Ligand Binding to a Two-Site One-Dimensional Lattice. Biochemistry (44):1251-67.

Galletto R, Jezewska MJ, Bujalowski W. 2005. Kinetics of Allosteric Conformational Transition of a Macromolecule Prior to Ligand Binding. Analysis of Stopped-flow Kinetic Experiments. Cell Biochemistry and Biophysics (42):121-44.

Galletto R, Jezewska MJ, Maillard R, Bujalowski W. 2005. The Nucleotide-Binding Site of the Escherichia coli DnaC Protein. Molecular Topography of DnaC Protein – Nucleotide Cofactor Complexes. Cell Biochemistry and Biophysics (43):331-53.

Haracska L, Acharya N, Unk I, Johnson RE, Hurwitz J, Prakash L, Prakash S.  2005 Feb. A single domain in human DNA polymerase i mediates interaction with PCNA: implications for translesion DNA synthesis.  Mol Cell Biol. 25(3):1183-1190.

Haracska L, Johnson RE, Prakash L, Prakash S. 2005 Nov. Trf4 and Trf5 proteins of Saccharomyces cerevisiae exhibit poly(A) RNA polymerase activity but no DNA polymerase activity.  Mol Cell Biol. 25(22):10183-10189.

Hill CE, Affatato AA, Wolfe KJ, Lopez MS, Hallberg C, Canistro D and Abdel-Rahman, SZ. 2005. Gender differences in genetic damage induced by the tobacco-specific nitrosamine NNK and the influence of the Thr241Met polymorphism in the XRCC3gene. Environ. Mol. Mutagen., 46, 22-29.

Hill CE, Wickliffe JK, Affatato AA, Wolfe KJ,  Kinslow CJ, Lopez MS and Abdel- Rahman SZ. 2005.The L84F and the I143V polymorphisms in the O6-Methylguanine-DNA-Methyltransferase (MGMT) gene increase human sensitivity to the genotoxic effects of the tobacco-specific nitrosamine carcinogen NNK. Pharmacogenet. Genomics,15(8):571-578.

Izumi T, Brown DB, Naidu CV, Bhakat KK, MacInnes MA, Saito H, Chen^| DJ and Mitra S. 2005. Two essential but distinct functions of the mammalian AP-endonuclease. Proc. Natl. Acad. Sci.  (102):5739-43.

Jackson EB, Theriot TA, Chattopadhyay R, Mitra S, and Izumi T.  2005. Analysis of nuclear transport signals in the human apurinic/apyrimidinic endonuclease (APE1/Ref1).  Nucleic Acids Research (33):3303-12.

Jezewska MJ, Lucius AL, Bujalowski W. 2005 Mar. Binding of Six Nucleotide Cofactors to the Hexameric Helicase RepA Protein of Plasmid RSF1010.  I. Direct Evidence of Cooperative Interactions Between the Nucleotide-Binding Sites of a Hexameric Helicase. Biochemistry (44):3865-76.

Johnson RE, Prakash L, Prakash S. 2005 Aug 22. Distinct mechanisms of cis-syn thymine dimer bypass by Dpo4 and DNA polymerase h. Proc Natl Acad Sci.  102(35):12359-12364.

Johnson RE, Prakash L, Prakash S. 2005 Jul 26. Biochemical evidence for the requirement of Hoogsteen base pairing for replication by human DNA polymerase i. Proc Natl Acad Sci. 102(30):10466-10471.

Kang J, Lee MS, Gorenstein DG. 2005. Quantitative analysis of chemiluminescence signals using a cooled charge-coupled device camera.  Analytical Biochemistry 345, 66-71.

Kang J, Lee MS, Gorenstein DG. 2005. The enhancement of PCR amplification of a random sequence DNA library by DMSO and betaine: application to in vitro combinatorial selection of aptamers. Journal of Biochemical and Biophysical Methods, 64, 147-151.

Leary J F, Reece LM, Yang X-B, and Gorenstein DG. 2005. High-Throughput Flow-cytometric Screening of Combinatorial Bead Libraries for Proteomics and Drug Discovery, Advanced Biomedical and Clinical Diagnostic Systems III, edited by Tuan Vo-Dinh, Warren S. Grundfest, David A. Benaron, Gerald E. Cohn, Proc. of SPIE Vol. 5692, 216-223.

Li ZY, Mao H, Kallick DA and Gorenstein DG.2005. The Effects of Thiophosphate Substitutions on Native siRNA Gene Silencing, Biochem. Biophys. Res. Commun., 329, 1026-1030.

Mandavilli BS, Boldogh I and Van Houten B. 2005 Feb 18. 3-Nitropropionic acid induced hydrogen peroxide, mitochondrial DNA damage and cell death are attenuated by Bcl-2 overexpression in PC12 cells. Molecular Brain Research, 133:215-23.

Nair DT, Johnson RE, Prakash L, Prakash S, Aggarwal AK. 2005 Sep 30.  Rev1 employs a novel mechanism of DNA synthesis using a protein template.  Science 309(5744):2219-2222.

Nair DT, Johnson RE, Prakash L, Prakash S, Aggarwal AK. 2005.  Human DNA polymerase i incorporates dCTP opposite template G via a G.C+ Hoogsteen base pair.  Structure 13:1569-1577.

Prakash S, Johnson RE, Prakash L. 2005 Jul. Eukaryotic translesion synthesis DNA polymerases: specificity of structure and function. Annu Rev Biochem. 74:317-353.

Somasunderam AD, Ferguson MR, Rojo DR, Thiviyanathan V, Li X, O’Brien WA, and Gorenstein DG.  2005.  Combinatorial Selection, Inhibition and Antiviral Activity of DNA Thioaptamers Targeting RNase H Domain of HIV-1 Reverse Transcriptase. Biochemistry, 44, 1038-10395.

Szczesny B and Mitra S.  2005.  Effect of aging on intracellular distribution of abasic (AP) endonuclease 1 in the mouse liver. Mech. Ageing Dev. (126):1071-78.

Thiviyanathan V, Somasunderam A, Volk DE and Gorenstein DG. 2005. 5-Hydroxy Uracil Can Form Stable Base Pairs With all Four Bases in a DNA Duplex, Chem. Commun., (3), 400-402.

Wickliffe JK, Galbert LA, Ammenheuser MM, Herring SM, Xie J, Masters OE 3rd, Friedberg EC, Lloyd RS, Ward JB Jr.  2005 Aug 10.  3,4-Epoxy-1-butene, a reactive metabolite of 1,3-butadiene, induces somatic mutations in Xpc-null mice.  Environ Mol Mutagen. 47(1):67-70.

Wolfle WT, Johnson RE, Minko IE, Lloyd RS, Prakash S, Prakash L. 2005 Oct. Human DNA polymerase i promotes replication through a ring-closed minor-groove adduct that adopts a syn conformation in DNA.  Mol Cell Biol. 25(19):8748-8754.

Wolfle WT, Washington MT, Kool ET, Spratt TE, Helquist SA, Prakash L, Prakash S. 2005 Aug. Evidence for a Watson-Crick hydrogen bonding requirement in DNA synthesis by human DNA polymerase k. Mol Cell Biol. 25(16):7137-7143.

Publication Archives

Important Links

NIEHS Annual Report http://www-apps.niehs.nih.gov/centers/Public/res-core/ctr1062-4690.htm