Mark Hedglin | Eberly College of Science

The genetic information (i.e., genome) of a human cell is encoded in strands of DNA that assemble into an antiparallel DNA double helix. Each time a cell divides, the genome must be faithfully replicated and transferred to a daughter cell for genetic inheritance. The former occurs during S-phase of the cell cycle and relies on high-fidelity, i.e. “replicative,” DNA polymerases that read template DNA strands and synthesize their complementary DNA. Additional “core” proteins and enzymes are also involved and the basic mechanism of human DNA replication has been deciphered. However, it is currently unknown how DNA replication is achieved on genomic DNA within a human cell. For example, the majority of proteins and enzymes implicated in human DNA replication are dynamically modified by chemical and protein moieties in vivo. Currently, the functional role and regulation of many of these modifications is unknown. Furthermore, genomic DNA is continuously subjected to damage from reactive metabolites and environmental mutagens. Prominent examples are modifications (lesions) to the native template DNA bases that alter or eliminate their base pairing capability. It is unclear how DNA lesions are accommodated during S-phase without compromising the fidelity of DNA replication. We aspire to decipher how efficient and faithful replication of the human genome is achieved within the highly-complex, dynamic, and reactive cellular environment. To do so, we employ a multi-disciplinary, collaborative approach, combining biophysical, biochemical, and molecular and cellular biology techniques to; 1) identify cellular factors involved in various aspects of human DNA replication and; 2) re-constitute human DNA replication in various biological scenarios and at various levels of complexity.

Websites

Hedglin Lab Website

@hedglinlab.bsky.social

@Hedglin_Lab X

Selected Publications (Hedglin Lab Members in BOLD, Undergraduate Hedglin Lab Members are also Underlined)

Shin Y, Hedglin M, Murakami KS. Cryo-EM structure of apo-form human DNA polymerase δ elucidates its minimal DNA synthesis activity without PCNA. J Biol Chem. 2025 Feb 22;301(4):108342.

Norris JL, Hedglin M. (Corresponding Author) Direct, ensemble FRET approaches to monitor transient state kinetics of human DNA polymerase δ holoenzyme assembly and initiation of DNA synthesis. Methods Enzymol. 2024;705:271-309.

Norris JL, Rogers LO, Pytko KG, Dannenberg RL, Perreault S, Kaushik V, Kuppa S, Antony E, Hedglin M. (Corresponding Author) Replication protein A dynamically re-organizes on primer/template junctions to permit DNA polymerase δ holoenzyme assembly and initiation of DNA synthesis. Nucleic Acids Res. 2024 Jul 22;52(13):7650-7664.

Pytko KG, Dannenberg RL, Eckert KA, Hedglin M. (Corresponding Author) Replication of [AT/TA]₂₅ Microsatellite Sequences by Human DNA Polymerase δ Holoenzymes Is Dependent on dNTP and RPA Levels. Biochemistry. 2024 Apr 16;63(8):969-983.

Muoio D, Laspata N, Dannenberg RL, Curry C, Darkoa-Larbi S, Hedglin M, Uttam S, Fouquerel E. PARP2 promotes Break Induced Replication-mediated telomere fragility in response to replication stress. Nat Commun. 2024 Apr 2;15(1):2857.

Stroik S, Carvajal-Garcia J, Gupta D, Edwards A, Luthman A, Wyatt DW, Dannenberg RL, Feng W, Kunkel TA, Gupta GP, Hedglin M, Wood R, Doublié S, Rothenberg E, Ramsden DA. Stepwise requirements for polymerases δ and θ in theta-mediated end joining. Nature. 2023 Nov;623(7988):836-841.

Hoitsma NM, Norris J, Khoang TH, Kaushik V, Chadda R, Antony E, Hedglin M, Freudenthal BD. Mechanistic insight into AP-endonuclease 1 cleavage of abasic sites at stalled replication fork mimics. Nucleic Acids Res. 2023 Jul 21;51(13):6738-6753.

Dannenberg RL, Cardina JA, Pytko KG, Hedglin M. (Corresponding Author) Tracking of progressing human DNA polymerase δ holoenzymes reveals distributions of DNA lesion bypass activities. Nucleic Acids Res. 2022 Sep 23;50(17):9893-9908.

Li M, Sengupta B, Benkovic SJ, Lee TH, Hedglin M. (Corresponding Author) PCNA Monoubiquitination Is Regulated by Diffusion of Rad6/Rad18 Complexes along RPA Filaments. Biochemistry. 2020 Dec 15;59(49):4694-4702.

Gaubitz C, Liu X, Magrino J, Stone NP, Landeck J, Hedglin M, Kelch BA. Structure of the human clamp loader reveals an autoinhibited conformation of a substrate-bound AAA+ switch. Proc Natl Acad Sci U S A. 2020 Sep 22;117(38):23571-23580.

Shin Y, Hedglin M, Murakami KS. Structural basis of reiterative transcription from the pyrG and pyrBI promoters by bacterial RNA polymerase. Nucleic Acids Res. 2020 Feb 28;48(4):2144-2155.

Li M, Larsen L, Hedglin M. (Corresponding Author) Rad6/Rad18 Competes with DNA Polymerases η and δ for PCNA Encircling DNA. Biochemistry. 2020 Feb 4;59(4):407-416.

Hedglin M (Corresponding Author), Aitha M, Pedley A, and Benkovic, SJ. Replication protein A dynamically regulates monoubiquitination of proliferating cell nuclear antigen. J Biol Chem. 2019 Mar 29; 294(13): 5157-5168.