BLM Helicase Facilitates Mus81 Endonuclease Activity in Human Cells

Ran Zhang,Nozomu Yanaihara,John Bradsher,Clare H Mcgowan,Steven P Linke,Veronique Blais,Curtis C Harris,Sagar Sengupta,Qin Yang

doi:10.1158/0008-5472.can-04-2421

Ran Zhang, Nozomu Yanaihara + Show 7 more

Open Access

https://doi.org/10.1158/0008-5472.can-04-2421

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Abstract

Bloom syndrome is a rare, autosomal recessive inherited disorder in humans. The product of the Bloom syndrome mutated gene, designated BLM, is a member of the RecQ helicase family. BLM has been proposed to function at the interface of replication and recombination, and to facilitate the repair of DNA damage. Here, we report in vivo physical interaction and colocalization of BLM and a DNA structure-specific endonuclease, Mus81, at sites of stalled replication forks outside the promyelocytic leukemia nuclear bodies during the S-phase arrest of the cell cycle. Amino acids 125 to 244 of Mus81 interact with the C-terminal region (amino acids 1,007-1,417) of BLM. Whereas Mus81 does not have any effect on the helicase activity of BLM, BLM can stimulate Mus81 endonuclease activity on the nicked Holliday junctions and 3' flap. This stimulation is due to enhanced binding of Mus81 to the DNA substrates. These data suggest a new function of BLM in cooperating with Mus81 during processing and restoration of stalled replication forks.

Highlights

Mus81, a highly conserved DNA structure–specific endonuclease, which shares homology with the XPF/Rad1 family of proteins involved in DNA nucleotide excision repair, is required for the survival of cells undergoing aberrant replication
RNA interference experiments show that Mus81 is required for mitotic recombination in somatic human cells [2]
In the absence of replication stress, BLM was mostly present in the nucleolus and promyelocytic leukemia (PML) nuclear bodies

Summary

Introduction

Mus81, a highly conserved DNA structure–specific endonuclease, which shares homology with the XPF/Rad1 family of proteins involved in DNA nucleotide excision repair, is required for the survival of cells undergoing aberrant replication. These results indicate that BLM-Mus81 interaction may have a direct consequence on the processing of different types of DNA junctions, and indirectly may serve as a platform for diverse functions like damage recognition, signal transduction, DNA repair, and recombination. Equivalent amounts of immunoprecipitated Mus81 from NHF, determined by quantitative Western blot done using purified recombinant GSTMus81 protein titration (data not shown), were used for these assays (Fig. 3A and C, bottom).

Results

Conclusion