53. In a human cell line, a large fraction of double-strand DNA breaks are repaired by non-homologous end joining (NHEJ). An inhibitor of FLAP endonuclease will affect

(1) recruitment of DNA—dependent kinase

(2) gap trimming

(3) DNA unwinding

(4) pairing of micro-homology regions.

Introduction

Double-strand DNA breaks (DSBs) are critical lesions that threaten genomic integrity. In human cells, one major repair pathway for DSBs is non-homologous end joining (NHEJ), which directly ligates broken DNA ends without requiring a homologous template. During NHEJ, DNA ends often require processing to create ligatable termini. A key player in this processing is flap endonuclease 1 (FEN-1), an enzyme that removes 5′ overhanging flaps and trims DNA ends to facilitate repair. Understanding how inhibition of FEN-1 affects NHEJ provides insight into the molecular mechanics of DNA repair.

Role of Flap Endonuclease (FEN-1) in NHEJ

• FEN-1 Function:
  FEN-1 is a structure-specific nuclease that recognizes and cleaves 5′ flap structures—single-stranded DNA overhangs that arise during DNA replication and repair.

• In NHEJ:
  When DNA ends are not perfectly compatible, small flaps or gaps can form. FEN-1 processes these flaps by trimming them, generating clean, ligatable ends.

• Gap Trimming:
  This trimming is essential for preparing DNA ends for subsequent ligation by DNA ligase IV, ensuring efficient and accurate repair.

Effect of FEN-1 Inhibition on NHEJ

• Inhibiting FEN-1 impairs gap trimming, leading to accumulation of unprocessed DNA flaps.

• This results in reduced efficiency of NHEJ because DNA ends remain incompatible for ligation.

• Other nucleases may partially compensate, but FEN-1 is a major contributor to flap removal during NHEJ.

Why Other Options Are Incorrect

• (1) Recruitment of DNA-dependent kinase:
  DNA-PK recruitment is primarily mediated by Ku proteins binding DNA ends, independent of FEN-1 activity.

• (3) DNA unwinding:
  DNA unwinding during NHEJ is not a major step; helicases may play roles elsewhere, but FEN-1 does not unwind DNA.

• (4) Pairing of micro-homology regions:
  Microhomology pairing is a separate step involving alignment of short homologous sequences and is not directly dependent on FEN-1.

Summary Table

Final Answer

(2) gap trimming

Keywords

flap endonuclease, FEN-1, non-homologous end joining, NHEJ, DNA double-strand break repair, gap trimming, DNA repair enzymes, DNA ligation, DNA end processing, genome stability

Conclusion

Flap endonuclease 1 (FEN-1) is crucial for processing DNA ends during non-homologous end joining by trimming 5′ flap structures and gaps. Inhibiting FEN-1 disrupts this gap trimming step, hindering the generation of ligatable DNA ends and thereby impairing efficient DSB repair via NHEJ. This highlights the essential role of FEN-1 in maintaining genomic integrity through DNA repair pathways.