Abstract
The protein Snm1B plays a key role in interstrand crosslink (ICL) repair. In a yeast two-hybrid screen we identified the protein PSF2 to bind Snm1B. PSF2 is a member of the GINS complex involved in replication initiation and elongation, and is known to play a role in ICL repair. Snm1B was shown to bind PSF2 in human cells through two regions, strongly to a 144 amino acid N-terminal region and weakly to a second smaller 37 amino acid C-terminal region. Ectopic expression of PSF2 increased the amount of Mus81, a protein component of the endonucleolytic complex involved in ICL repair, co-immunoprecipitating with Snm1B. Moreover, deleting the N-terminal, but not C-terminal region of Snm1B reduced the amount of co-immunoprecipitated Mus81. Conversely, the telomere-binding protein TRF2 competed with PSF2 for binding to the C-terminus of Snm1B, and deletion of this region, but not the N-terminal region, reduced Snm1B chromatin association. We speculate that the N-terminal region of Snm1B forms a complex containing PSF2 and Mus81, while the C-terminal region is important for PSF2-mediated chromatin association.
Highlights
Interstrand crosslinks (ICLs) are toxic lesions that covalently attach opposite strands of DNA [1]
Snm1B is required for proper ICL repair, as knockdown of this protein leads to sensitivity of cells to ICLs [2,3,4,13] and blocks the formation of double-strand breaks (DSBs) that occur as an intermediate in ICL repair [3,16]
One of these potential interacting proteins was the PSF2 subunit of the GINS complex, which was of particular interest given that PSF2, like Snm1B, is required for ICL repair [28,30]
Summary
Interstrand crosslinks (ICLs) are toxic lesions that covalently attach opposite strands of DNA [1]. Snm1B is a 60 kDa protein belonging to the b-CASP family of proteins, which contains Snm1A (Dclre1A) and Snm1C (Artemis/ Dclre1C) [8] These proteins are characterized by b-CASP and Metallo-b-Lactamase domains responsible for nucleic acid hydrolysis [8], and all three proteins have inherent 59-39 DNA exonuclease activity [6,9,10,11]. Mus and Eme form the structure-specific endonuclease complex Mus81/Eme1 [17,18] that is important for cleavage of replication fork substrates in vitro [17,18], and for the formation of DSBs after ICL formation during replication in vivo [19] These results suggest that association of Snm1B with Mus is important for ICL repair during DNA replication [3]
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