Stimulation of Flap Endonuclease-1 by the Bloom's Syndrome Protein

Sudha Sharma,Joshua A Sommers,Leonard Wu,Vilhelm A Bohr,Ian D Hickson,Robert M Brosh

doi:10.1074/jbc.m309898200

Abstract

Bloom's syndrome (BS) is a rare autosomal recessive genetic disorder associated with genomic instability and an elevated risk of cancer. Cellular features of BS include an accumulation of abnormal replication intermediates and increased sister chromatid exchange. Although it has been suggested that the underlying defect responsible for hyper-recombination in BS cells is a temporal delay in the maturation of DNA replication intermediates, the precise role of the BS gene product, BLM, in DNA metabolism remains elusive. We report here a novel interaction of the BLM protein with the human 5'-flap endonuclease/5'-3' exonuclease (FEN-1), a genome stability factor involved in Okazaki fragment processing and DNA repair. BLM protein stimulates both the endonucleolytic and exonucleolytic cleavage activity of FEN-1 and this functional interaction is independent of BLM catalytic activity. BLM and FEN-1 are associated with each other in human nuclei as shown by their reciprocal co-immunoprecipitation from HeLa nuclear extracts. The BLM-FEN-1 physical interaction is mediated through a region of the BLM C-terminal domain that shares homology with the FEN-1 interaction domain of the Werner syndrome protein, a RecQ helicase family member homologous to BLM. This study provides the first evidence for a direct interaction of BLM with a human nucleolytic enzyme. We suggest that functional interactions between RecQ helicases and Rad2 family nucleases serve to process DNA substrates that are intermediates in DNA replication and repair.

Highlights

Bloom’s syndrome (BS) is a rare autosomal recessive genetic disorder associated with genomic instability and an elevated risk of cancer
Physical Interaction between BLM and FEN-1—Previously, we reported a physical interaction between the human WRN
FEN-1 failed to be precipitated by the BLM antibody (Fig. 1A, lower panel, lane 5) despite its presence in BLMϪ/Ϫ nuclear extracts (Fig. 1A, lower panel, lane 4)

Summary

Introduction

Bloom’s syndrome (BS) is a rare autosomal recessive genetic disorder associated with genomic instability and an elevated risk of cancer. We suggest that functional interactions between RecQ helicases and Rad family nucleases serve to process DNA substrates that are intermediates in DNA replication and repair. The gene mutated in BS, designated BLM, encodes a 1417 amino acid protein with a central region (residues 649 –1000) containing seven conserved motifs characteristic of DNA helicases [6]. The region of BLM that contains these seven motifs exhibits ϳ40% homology to other proteins in the RecQ family of DNA helicases. Of the human RecQ homologs, three, including BLM, are associated with genetic disorders characterized by chromosomal instability. The linkage of mutations in human RecQ helicases with genomic instability, cancer, and premature aging suggests that this class of enzymes have important caretaker roles in specialized pathways of DNA metabolism [5].

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