Molecular Mechanisms for the Repair and Replication of DNA-Protein Crosslinks

Background and Previous Work

Environmental and endogenous exposure to chemicals that produce DNA-protein and DNA-peptide crosslinks are correlated with an increased risk of several cancers, asthma, and other diseases.  Currently many thousands of individuals in the US population are exposed to one of the most common DNA-protein crosslink-inducing agents, formaldehyde.  These exposures take place in both occupational and in-home settings and affected individuals often experience multi-year chronic exposures that are far in excess of typical indoor air quality standards.  Although little is currently known concerning the cellular repair and response mechanisms for this class of DNA lesions, our prior investigations have both rigorously established synthetic chemical procedures to create and utilize DNAs containing site-specifically modified DNA-protein crosslinks and determined dose-dependent, genome-wide assays that identify genes whose products function to limit DNA-protein crosslink-induced cytotoxicity.  These investigations have generated a series of hypotheses which postulate that eukaryotic cells exposed to chronic, low levels of DNA-protein crosslinking agents minimize cytotoxicity and mutagenesis through homologous recombination, while following acute high dose exposure, cells will shift to pathways initiated by nuclear proteasome-dependent degradation of covalently linked proteins that can then be processed via either nucleotide excision repair or translesion DNA polymerases.

Most Recent Work

To accomplish our objective of determining the fundamental pathways for the repair and tolerance of DNA-protein crosslinks under chronic and acute exposures, gene-specific deletion, or siRNA screening will identify the constellation of genes and interrelated pathways that are critical in limiting cellular toxicity and mutagenesis.  The roles of individual gene products in modulating cellular response to DNA-protein crosslinks will include repair, recombination, translesion synthesis, cell cycle check points, chromatin remodeling and proteolytic pathways.  Biochemical analyses of repair intermediates and the activities of translesion synthesis polymerases will be established using DNAs containing site-specific DNA-peptide crosslinks and randomly adducted DPCs. Collectively, these investigations will yield comprehensive analyses of repair and tolerance of this class of lesions.


Kumari A, Lim YX, Newell AH, Olson SB, McCullough AK. Formaldehyde-induced genome instability is suppressed by an XPF-dependent pathway. DNA Repair (Amst). 11(3):236-46, 2012. PMID: 22186232

Yamanaka K, Minko IG, Finkel SE, Goodman MF, Lloyd RS. The role of high fidelity E. coli DNA polymerase I in the replication bypass of deoxyadenosine DNA-peptide cross-link. J Bacteriol. 193(15):3815-21, 2011. PMID: 21622737

Huang H, Kozekov ID, Kozekova A, Wang H, Lloyd RS, Rizzo CJ, Stone MP. DNA cross-link induced by trans-4-hydroxynonenal. Environ Mol Mutagen. Jul;51(6): 625-34, 2010. PMID 20577992

Huang H, Kozekov ID, Kozekova A, Rizzo CJ, McCullough AK, Lloyd RS, Stone MP. Minor groove orientation of the KWKK peptide tethered via the N-terminal amine to the acrolein-derived 1,N2-gamma-hydroxypropanodeoxyguanosine lesion with a trimethylene linkage. Biochemistry. 27;49(29): 6155-64, 2010. PMID: 20604523

Kumari A, Minko IG, Smith RL, Lloyd RS, McCullough AK. Modulation of UvrD helicase activity by covalent DNA-protein crosslinks. J Biol Chem. 285(28): 21313-22, 2010. PMID: 20444702

Yamanaka K, Minko IG, Takata K, Kolbanovskiy A, Kozekov ID, Wood RD, Rizzo CJ, Lloyd RS. Novel enzymatic function of DNA polymerase nu in translesion DNA synthesis past major groove DNA-peptide and DNA-DNA cross-links. Chem Res Toxicol. 23(3): 689-95, 2010. PMID: 20102227

de Graaf B, Clore A, McCullough AK. Cellular pathways for DNA repair and damage tolerance of formaldehyde-induced DNA-protein crosslinks. DNA Repair. 8(10): 1207-14, 2009. PMID19625222

Minko IG, Kozekov ID, Harris TM, Rizzo CJ, Lloyd RS, Stone MP. Chemistry and biology of DNA containing 1,N(2)-deoxyguanosine adducts of the alpha, beta-unsaturated aldehydes acrolein, crontonaldeyde, and 4-hydroxynoneal. Chem Res Toxicol 22(5):759-78, 2009. PMID: 19397281

Minko IG, Kozekov ID, Kozekova A, Harris TM, Rizzo CJ, Lloyd RS.  Mutagenic potential of DNA-peptide crosslinks mediated by acrolein-derived DNA adducts. Mutat Res. 637 (1-2):161-172, 2008. PMID: 17868748

Minko IG, Yamanaka, K, Kozekov ID, Kozekova A, Indiani, C, O’Donnell, ME, Jiang, Q, Goodman, MF, Rizzo CJ, Lloyd, RS. Replication Bypass of the Acrolein-mediated Deoxyguanine DNA –Peptide Cross-links by DNA polymerases of the DinB Family. Chem Res Toxicol. 21(10):1983-90, 2008. PMID: 18788757

Minko IG, Kurtz AJ, Croteau DL, Van Houten B, Harris TM, Lloyd RS. Initiation of repair of DNA-polypeptide cross-links by the UvrABC nuclease. Biochemistry. 44: 3000-3009, 2005. PMID: 15723543

Kurtz AJ, and Lloyd RS. 1,N(2)-Deoxyguanosine Adducts of Acrolein, Crotonaldehyde, and trans-4-Hydroxynonenal Cross-link to Peptides via Schiff Base Linkage. J Biol Chem. 278(8):5970-5976, 2003. PMID: 12502710

Kurtz AJ, Dodson ML, Lloyd RS. Evidence for multiple imino intermediates and identification of reactive nucleophiles in peptide-catalyzed beta-elimination at abasic sites. Biochemistry. 41(22):7054-7064, 2002. PMID: 12033939

Minko IG, Zou Y, Lloyd RS. Incision of DNA-protein crosslinks by UvrABC nuclease suggests a potential repair pathway involving nucleotide excision repair. Proc Natl Acad Sci USA. 99(4):1905-1909, 2002. PMID: 11842222