Repair-Resistant DNA Lesions

Abstract

The eukaryotic global genomic nucleotide excision repair (GG-NER) pathway is the major mechanism that removes most bulky and some nonbulky lesions from cellular DNA. There is growing evidence that certain DNA lesions are repaired slowly or are entirely resistant to repair in cells, tissues, and in cell extract model assay systems. It is well established that the eukaryotic DNA lesion-sensing proteins do not detect the damaged nucleotide, but recognize the distortions/destabilizations in the native DNA structure caused by the damaged nucleotides. In this article, the nature of the structural features of certain bulky DNA lesions that render them resistant to NER, or cause them to be repaired slowly, is compared to that of those that are good-to-excellent NER substrates. Understanding the structural features that distinguish NER-resistant DNA lesions from good NER substrates may be useful for interpreting the biological significance of biomarkers of exposure of human populations to genotoxic environmental chemicals. NER-resistant lesions can survive to replication and cause mutations that can initiate cancer and other diseases. Furthermore, NER diminishes the efficacy of certain chemotherapeutic drugs, and the design of more potent pharmaceuticals that resist repair can be advanced through a better understanding of the structural properties of DNA lesions that engender repair-resistance.

Figure 1

Structures of UV photodimers: (A) T^T cyclobutane pyrimidine dimer (CPD) and (B) 6–4 UV photoproduct. (C) Co-crystal structure of yeast Rad4-Rad23 with a T^T CPD (not resolved experimentally) opposite two thymines (blue) in the complementary strand (Min and Pavletich; PDB ID: 2QSG).

Figure 2

(A) Structures and carbon atom numbering systems of bay region benzo[a]pyrene (B[a]P), and fjord region benzo[c]phenanthrene (B[c]Ph), benzo[g]chrysene (B[g]C), and dibenzo[a,l]pyrene (DB[a,l]P). (B) Metabolic activation of PAH by P450 cytochrome and epoxy hydrolases that generate the enantiomeric diol epoxides is shown. The absolute configurations of substituents around the 7,8,9,10 (B[a]P); 4,3,2,1 (B[c]Ph); 11,12,13,14 (DB[a,l]P and B[g]C) of the metabolized aromatic rings, shaded in gray, are also shown.

Figure 3

Stereochemistry-dependent conformational motifs of DNA adducts that result from the reactions of B[a]PDE with (A, B) dG, and with (C) dA in double-stranded DNA (see the text for details).

Figure 4

Steric hindrance due to bulky −OH groups limits the allowed values of the torsion angle β′ and thus the conformational space of the bulky polycyclic aromatic residues in the (+)-trans and (−)-trans-B[a]P-N²–dG adducts in double stranded DNA. The two top structures illustrate the principles of opposite orientations of the PAH residues relative to the planes of dG that result from the binding of enantiomeric PAH diol epoxides to the exocyclic amino groups of purines in DNA. The bottom structure designates the torsion angle β′.

Figure 5

Conformations of DB[a,l]PDE-N⁶–dA (top) and DB[a,l]PDE-N²–dG (bottom) adducts. 14R-N⁶–dA: intercalated on the 5′-side of the intact dA*–dT base pair from the major groove without base displacement; 14S-N⁶–dA: same, but intercalated on the 3′-side of dA*–dT. 14R-N²–dG: intercalated from the minor groove on the 3′-side of the disrupted dG*–dC base pair. 14S-N²–dG: the DB[a,l]PDE residue is positioned in a distorted and widened minor groove on the 5′-side of the dG*–dC base pair.

Figure 6

Relative NER efficiencies in Hela cell extracts of stereoisomeric (+)-trans- and (+)-cis-B[a]PDE-N²–dG adducts. (A) Typical autoradiograph of a gel depicting dual excision products in the 24–32 nucleotide range (size markers shown in lane M) after incubation of 135-mer control duplexes with (+)-trans- (lane 1) or (+)-cis-B[a]PDE-N²–dG adducts (lane 2) for 60 min. (B) Relative incision efficiencies after correcting for loading differences in each lane (data adapted from Mocquet et al.). (C, D) Stereochemistry of the (+)-cis- and (+)-trans- B[a]P-N²–dG adducts, respectively.

Figure 7

Effects of base sequence context on the NER efficiencies of the same 10S (+)-trans-B[a]PDE-N²–dG adduct in HeLa cell extracts.

Figure 8

Dependence of NER efficiencies in HeLa cell extracts of 10R (−)-trans- and 10S (+)-trans-B[a]PDE-N⁶–dA adducts in different sequence contexts (NER data adapted from Buterin et al.).

Figure 9

Deleting the partner C from double-stranded DNA (135-del duplexes) abolishes the NER efficiency that is observed in full (135-mer) duplexes (Full) with C opposite G* ((+)-cis-B[a]PDE-N²–dG). (A) Gel autoradiograph of NER dual incision products incubated in HeLa cell extracts for 0, 10, 20, and 30 min (lanes 1, 2, 3, and 4, respectively; lane M, size markers). (B) Thermal melting curves of 11-mer Del duplexes with and without the adduct (data adapted from Reeves et al.).

Figure 10

(A) Autoradiograph of dual excision products after incubation of stereoisomeric B[a]PDE-N²−dA (G*-10), DB[a,l]PDE-N⁶–dA (A*-14), or DB[a,l]PDE-N²–dG (G*-14) adducts embeded in identical 135-mer base sequence contexts in HeLa cell extracts. (B) Relative NER efficiencies and (C) impact of the same stereoisomeric DNA adducts on thermal stabilities (ΔT_m); the adducts were embedded in the 11-mer duplexes [5′-d(CCATCX*CTACC)]·[5′-d(GGTAGYGATGG)] with X* = DB[a,l]PDE-N⁶–dA or DB[a,l]PDE-N²–dG, and Y = T or C, respectively; these same 11-mers were embedded in the 135-mer duplexes in the NER experiments depicted in panel A.

Figure 11

Summary of stereochemical features of the fjord PAH diol epoxide -N⁶–dA and -N²–dG adducts.

Figure 12

Structures of Aflatoxin B1-derived guanine adducts.

Figure 13

Structures of 2-aminofluorene (AF) and 2-acetylaminofluorene (AAF)-derived guanine adducts.

Figure 14

Structures of 3-nitrobenzanthrone (ABA)-derived guanine and adenine adducts.

Figure 15

Structures of 6-nitrochrysene (6AC)-derived guanine and adenine adducts.

Figure 16

Structures of aristolochic acid-derived adenine adducts. X = COCH₃ for ALI, and X = H for ALII.

Figure 17

Structures of 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) derived guanine adducts.