| FEN1 activity / context | Primary substrate(s) | Key partners / regulators | Representative 2023–2024 mechanistic or structural findings | Quantitative details | Citations |
|---|---|---|---|---|---|
| Flap endonuclease during Okazaki fragment maturation | Short 5′ RNA/DNA flaps generated by Pol δ strand displacement on lagging-strand intermediates | PCNA; RNaseH2 | FEN1 is the primary nuclease for short 5′ flaps in OFM; cryo-EM captured endogenous PCNA–FEN1 and PCNA–FEN1–RNaseH2 assemblies in multiple primer-removal states, supporting the PCNA “toolbelt” model and indicating that RNaseH2 can promote FEN1-mediated flap cleavage through DNA conformational modulation; product release from FEN1 was identified as rate-limiting | PCNA can stimulate FEN1 activity by ~10–50×; Okazaki primers are ~30 nt total (8–12 nt RNA + 10–20 nt DNA); typical displaced short flaps are ~2–10 nt; cryo-EM resolution 3.5–3.8 Å | (pqac-00000010, pqac-00000016) |
| Structural basis of flap cleavage on PCNA | Nicked/flapped DNA with downstream duplex and 5′ flap | PCNA via PIP-box and IDCL interactions | Structures show one FEN1 bound to one PCNA protomer, with the FEN1 PIP-box inserted into the PCNA hydrophobic pocket; DNA adopts an L-shaped configuration; the cleaved 5′ flap remains transiently bound in an early post-catalytic state, explaining slow product release | One PCNA ring can potentially recruit up to 3 PIP-box proteins; downstream duplex ~12 bp; upstream duplex through the ring 14–19 bp; only ~3 proximal flap nucleotides traceable; catalytic metal-binding residues include E158, E160, D179, D181 | (pqac-00000010, pqac-00000016) |
| Long-patch base excision repair (LP-BER) flap removal | Short DNA flaps produced during repair resynthesis after damaged-base removal | PCNA; BER machinery | FEN1 is a core LP-BER nuclease that removes repair flaps after strand displacement synthesis; recent reviews continue to place FEN1 as the canonical 5′-flap nuclease linking BER and replication, with a two-metal active center characteristic of structure-specific nucleases | LP-BER flaps described as ~2–20 nt; active site uses 2 Mg2+ ions | (pqac-00000013, pqac-00000015, pqac-00000017) |
| 5′ exonuclease proofreading / α-segment error editing | 5′ termini and misincorporated Pol α-generated segments during lagging-strand synthesis | PCNA; mismatch-repair-associated factors | Beyond flap cutting, FEN1’s 5′ exonuclease activity contributes to editing/removal of Pol α errors during Okazaki fragment maturation, helping prevent mutagenesis when nascent lagging strands are processed | No specific 2023–2024 numeric measurement in retrieved text; activity explicitly distinguished from flap endonuclease and gap endonuclease functions | (pqac-00000017, pqac-00000000) |
| Gap endonuclease / stalled fork & structured DNA processing | Gapped or structured DNA intermediates, including secondary structures at difficult-to-replicate regions | Rad9–Rad1–Hus1; SUMOylation; checkpoint signaling | FEN1’s GEN activity is implicated in processing stalled replication forks and DNA secondary structures; SUMO-1 modification promotes switching from replication-associated partners toward the 9-1-1 clamp to counter replication stress | Qualitative evidence in retrieved sources; no specific 2023–2024 numeric value extracted | (pqac-00000014, pqac-00000017) |
| Replication-stress response / partner switching | Flap or fork-associated DNA at stalled replication sites | SUMO-1; Rad9–Rad1–Hus1; phosphorylation | DNA damage-induced sequential phosphorylation and SUMO-1 conjugation facilitate FEN1 interaction with HUS1, helping FEN1 act in repair rather than only canonical OFM; SUMO-defective FEN1 mutants are hypersensitive to fork-stalling agents | Triggering agents cited include UV, hydroxyurea, and mitomycin C; no precise fold-change reported in retrieved excerpt | (pqac-00000014) |
| DPC repair: formaldehyde and TOP2-linked lesions | DPC-conjugated 5′ flaps, including BER-generated 5′ flaps and TOP2-DPC-associated flap substrates | PARP1; PARG; SPRTN; TDP2 | FEN1 can excise non-enzymatic FA-DPCs and enzymatic TOP2-DPCs from 5′-flap-like structures; PARylation promotes recruitment rather than intrinsic catalytic enhancement, placing FEN1 in a PARP1/PARG-regulated DPC-repair axis parallel to SPRTN | Major PARylation site mapped to E285; nuclease-dead comparator D181A; no intrinsic activity increase detected after PARylation in vitro | (pqac-00000001, pqac-00000002, pqac-00000011) |
| DPC repair: TOP3A-DPC pathway choice | Persistent TOP3A-DPC intermediates | PARP1; ubiquitylation machinery; SPRTN/TDP2 | PARP1-driven PARylation recruits FEN1 to TOP3A-DPCs, while ubiquitylation supports an alternative proteolytic pathway; blocking PARylation reduces FEN1–TOP3A interaction and increases TOP3A-DPC burden, indicating PTM-controlled pathway choice | Persistent TOP3A-DPC model used catalytic mutant R364W; no additional quantitative structural values extracted | (pqac-00000012) |
| NER-/R-loop-adjacent cross-talk | 5′-flap substrates with repair gaps; R-loop-associated ssDNA | XPA; RPA | FEN1 forms ternary complexes with XPA and DNA, supporting possible roles beyond canonical BER/replication, including post-incision NER resynthesis or other repair processes; XPA moderately inhibits FEN1 catalytic activity in vitro | Tested DNA substrates carried a 31-nt 5′ flap with 3-, 10-, or 26-nt gaps | (pqac-00000013) |
| Cancer targeting / biomarker relevance | Replication/repair intermediates in stressed or HR-deficient tumor cells | PARP/PARG axis; EXO1; synthetic lethality frameworks | Recent work positions FEN1 as a vulnerability in tumors with defective PAR metabolism or HR deficiency; reviews frame structure-specific nucleases, including FEN1, as therapeutic targets, and PARG-deficient tumor cells show increased dependence on EXO1/FEN1-mediated repair | No single universal effect size given in retrieved excerpt; dependency highlighted in 2024 tumor models | (pqac-00000015) |


*Table: This table summarizes the main biochemical activities of FEN1, their substrates, major cofactors/regulators, and recent 2023–2024 mechanistic advances. It is useful for linking classical functional annotation to current structural biology and repair-pathway models.*