title: Human MUS81 complexes stimulate flap endonuclease 1.
abstract: The yeast heterodimeric Mus81-Mms4 complex possesses a structure-specific endonuclease activity that is critical for the restart of stalled replication forks and removal of toxic recombination intermediates. Previously, we reported that Mus81-Mms4 and Rad27 (yeast FEN1, another structure-specific endonuclease) showed mutual stimulation of nuclease activity. In this study, we investigated the interactions between human FEN1 and MUS81-EME1 or MUS81-EME2, the human homologs of the yeast Mus81-Mms4 complex. We found that both MUS81-EME1 and MUS81-EME2 increased the activity of FEN1, but FEN1 did not stimulate the activity of MUS81-EME1/EME2. The MUS81 subunit alone and its N-terminal half were able to bind to FEN1 and stimulate its endonuclease activity. A truncated FEN1 fragment lacking the C-terminal region that retained catalytic activity was not stimulated by MUS81. Michaelis-Menten kinetic analysis revealed that MUS81 increased the interaction between FEN1 and its substrates, resulting in increased turnover. We also showed that, after DNA damage in human cells, FEN1 co-localizes with MUS81. These findings indicate that the human proteins and yeast homologs act similarly, except that the human FEN1 does not stimulate the nuclease activities of MUS81-EME1 or MUS81-EME2. Thus, the mammalian MUS81 complexes and FEN1 collaborate to remove the various flap structures that arise during many DNA transactions, including Okazaki fragment processing.
authors.affiliation: Department of Biological Sciences, Center for DNA Replication and Genome Instability, Korea Advanced Institute of Science and Technology, Daejeon, Korea.
authors.forename: Yong-Keol
authors.initials: YK
authors.lastname: Shin
authors.forename: Tamir
authors.initials: T
authors.lastname: Amangyeld
authors.forename: Tuan A
authors.initials: TA
authors.lastname: Nguyen
authors.forename: Palinda R
authors.initials: PR
authors.lastname: Munashingha
authors.forename: Yeon-Soo
authors.initials: YS
authors.lastname: Seo
mesh_terms: D015153:Blotting, Western; D004249:DNA Damage; D017931:DNA Primers / Q000737:chemistry; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D004706:Endodeoxyribonucleases / Q000235:genetics / Q000378:metabolism*; D004720:Endonucleases / Q000235:genetics / Q000378:metabolism*; D045585:Flap Endonucleases / Q000235:genetics / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D006801:Humans; D007700:Kinetics; D010957:Plasmids; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D011995:Recombination, Genetic; D013379:Substrate Specificity
chemical_list: D017931:DNA Primers; D004268:DNA-Binding Proteins; D011994:Recombinant Proteins; C507947:Eme1 protein, human; D004706:Endodeoxyribonucleases; D004720:Endonucleases; D045585:Flap Endonucleases; C498772:MUS81 protein, human; C468317:FEN1 protein, human