Human Fanconi Anemia Complementation Group A Protein Stimulates the 5’ Flap Endonuclease Activity of FEN1

Liangyue Qian,Yanbin Zhang,Suyog Padgaonkar,Fenghua Yuan,Paola Rodriguez-Tello,Fenfei Leng

doi:10.1371/journal.pone.0082666

Abstract

In eukaryotic cells, Flap endonuclease 1 (FEN1) is a major structure-specific endonuclease that processes 5’ flapped structures during maturation of lagging strand DNA synthesis, long patch base excision repair, and rescue of stalled replication forks. Here we report that fanconi anemia complementation group A protein (FANCA), a protein that recognizes 5’ flap structures and is involved in DNA repair and maintenance of replication forks, constantly stimulates FEN1-mediated incision of both DNA and RNA flaps. Kinetic analyses indicate that FANCA stimulates FEN1 by increasing the turnover rate of FEN1 and altering its substrate affinity. More importantly, six pathogenic FANCA mutants are significantly less efficient than the wild-type at stimulating FEN1 endonuclease activity, implicating that regulation of FEN1 by FANCA contributes to the maintenance of genomic stability.

Highlights

During DNA replication, various structured DNA intermediates are effectively processed to avoid deleterious consequences such as genome instability and cancer
Initial incubation of the purified protein with the flap structures showed that Flap endonuclease 1 (FEN1) incised both the DNA and RNA flap structures (Figure 1A)
These results clearly demonstrate that mutations in fanconi anemia complementation group A protein (FANCA) significantly affect its ability to stimulate FEN1 and the interaction between FANCA and FEN1 is relevant to the etiology of Fanconi anemia

Summary

Introduction

During DNA replication, various structured DNA intermediates are effectively processed to avoid deleterious consequences such as genome instability and cancer. Replication on the lagging strands is discontinuous and initiated by DNA Pol α, which synthesizes an RNA primer approximately 12-nt long that is further extended with approximately 20-nt of DNA. This RNA-DNA hybrid is believed to be made at low fidelity and subjected to displacement by the high-fidelity DNA Pol δ. The strand displacement activity of Pol δ produces a 5’ single-stranded flap structure that contains the RNA primer and some of the initiator DNA. Defects in FEN1 cause accumulation of mutations, genomic instability, cancer predisposition and chronic inflammation [20,21,22]

Objectives

Methods

Results

Conclusion