title: Nse1 RING-like domain supports functions of the Smc5-Smc6 holocomplex in genome stability.
abstract: The Smc5-Smc6 holocomplex plays essential but largely enigmatic roles in chromosome segregation, and facilitates DNA repair. The Smc5-Smc6 complex contains six conserved non-SMC subunits. One of these, Nse1, contains a RING-like motif that often confers ubiquitin E3 ligase activity. We have functionally characterized the Nse1 RING-like motif, to determine its contribution to the chromosome segregation and DNA repair roles of Smc5-Smc6. Strikingly, whereas a full deletion of nse1 is lethal, the Nse1 RING-like motif is not essential for cellular viability. However, Nse1 RING mutant cells are hypersensitive to a broad spectrum of genotoxic stresses, indicating that the Nse1 RING motif promotes DNA repair functions of Smc5-Smc6. We tested the ability of both human and yeast Nse1 to mediate ubiquitin E3 ligase activity in vitro and found no detectable activity associated with full-length Nse1 or the isolated RING domains. Interestingly, however, the Nse1 RING-like domain is required for normal Nse1-Nse3-Nse4 trimer formation in vitro and for damage-induced recruitment of Nse4 and Smc5 to subnuclear foci in vivo. Thus, we propose that the Nse1 RING-like motif is a protein-protein interaction domain required for Smc5-Smc6 holocomplex integrity and recruitment to, or retention at, DNA lesions.
authors.affiliation: Department of Molecular Biology, The Scripps Research Institute, La Jolla, CA 92037, USA.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Pebernard
authors.forename: J Jefferson P
authors.initials: JJ
authors.lastname: Perry
authors.forename: John A
authors.initials: JA
authors.lastname: Tainer
authors.forename: Michael N
authors.initials: MN
authors.lastname: Boddy
mesh_terms: D000595:Amino Acid Sequence; D002352:Carrier Proteins / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000378:metabolism*; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism*; D019281:Dimerization; D005656:Fungal Proteins / Q000378:metabolism; D006801:Humans; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D012568:Schizosaccharomyces / Q000235:genetics* / Q000502:physiology*; D029702:Schizosaccharomyces pombe Proteins / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry
chemical_list: D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D002868:Chromosomal Proteins, Non-Histone; D005656:Fungal Proteins; C533270:NSMCE1 protein, human; C498869:Nse1 protein, S pombe; D009687:Nuclear Proteins; C432931:SMC5 protein, human; C432932:SMC6 protein, human; D029702:Schizosaccharomyces pombe Proteins; C486628:Smc5 protein, S pombe; C486630:smc6 protein, S pombe; D044767:Ubiquitin-Protein Ligases