title: Different non-synonymous polymorphisms modulate the interaction of the WRN protein to its protein partners and its enzymatic activities.
abstract: Werner syndrome (WS) is characterized by the premature onset of several age-associated pathologies including cancer. The protein defective in WS patients (WRN) is a helicase/exonuclease involved in DNA replication and repair. Here, we present the results of a large-scale proteome analysis that has been undertaken to determine protein partners of different polymorphic WRN proteins found with relatively high prevalence in the human population. We expressed different fluorescently tagged-WRN (eYFP-WRN) variants in human 293 embryonic kidney cells (HEK293) and used a combination of affinity-purification and mass spectrometry to identify different compositions of WRN-associated protein complexes. We found that a WRN variant containing a phenylalanine residue at position 1074 and an arginine at position 1367 (eYFP-WRN(F-R)) possesses more affinity for DNA-PKc, KU86, KU70, and PARP1 than a variant containing a leucine at position 1074 and a cysteine at position 1367 (eYFP-WRN(L-C)). Such results were confirmed in a WRN-deficient background using WS fibroblasts. Interestingly, the exonuclase activity of WRN recovered from immunoprecipitated eYFP-WRN(L-C) variant was lower than the eYFP-WRN(F-R) in WS cells. Finally, HEK293 cells and WS fibroblasts overexpressing the eYFP-WRN(F-R) variant were more resistant to the benzene metabolite hydroquinone than cells expressing the eYFP-WRN(L-C) variant. These results indicate that the protein-protein interaction landscape of WRN is subject to modulation by polymorphic amino acids, a characteristic associated with distinctive cell survival outcome.
authors.affiliation: Centre de Recherche du CHU de Québec, Pavillon CHUL Université Laval, Faculté de Médecine, Québec, Canada.
authors.forename: Jean-Philippe
authors.initials: JP
authors.lastname: Gagné
authors.affiliation: Centre de Recherche sur le Cancer de l'Université Laval, Hôpital Hôtel-Dieu de Québec, Québec, Québec, Canada.
authors.forename: Sophie
authors.initials: S
authors.lastname: Lachapelle
authors.affiliation: Centre de Recherche du CHU de Québec, Pavillon CHUL Université Laval, Faculté de Médecine, Québec, Canada.
authors.forename: Chantal
authors.initials: C
authors.lastname: Garand
authors.affiliation: Centre de Recherche du CHU de Québec, Pavillon CHUL Université Laval, Faculté de Médecine, Québec, Canada.
authors.forename: Serges P
authors.initials: SP
authors.lastname: Tsofack
authors.affiliation: Centre de Recherche sur le Cancer de l'Université Laval, Hôpital Hôtel-Dieu de Québec, Québec, Québec, Canada.
authors.forename: Yan
authors.initials: Y
authors.lastname: Coulombe
authors.affiliation: Centre de Recherche sur le Cancer de l'Université Laval, Hôpital Hôtel-Dieu de Québec, Québec, Québec, Canada.
authors.forename: Marie-Christine
authors.initials: MC
authors.lastname: Caron
authors.affiliation: Centre de Recherche du CHU de Québec, Pavillon CHUL Université Laval, Faculté de Médecine, Québec, Canada.
authors.forename: Guy G
authors.initials: GG
authors.lastname: Poirier
authors.affiliation: Centre de Recherche sur le Cancer de l'Université Laval, Hôpital Hôtel-Dieu de Québec, Québec, Québec, Canada.
authors.forename: Jean-Yves
authors.initials: JY
authors.lastname: Masson
authors.affiliation: Centre de Recherche du CHU de Québec, Pavillon CHUL Université Laval, Faculté de Médecine, Québec, Canada.
authors.forename: Michel
authors.initials: M
authors.lastname: Lebel
mesh_terms: D001426:Bacterial Proteins; D004249:DNA Damage; D057809:HEK293 Cells; D006801:Humans; D006873:Hydroquinones / Q000494:pharmacology; D047468:Immunoprecipitation; D008164:Luminescent Proteins; D013058:Mass Spectrometry; D020641:Polymorphism, Single Nucleotide; D020543:Proteome; D000071657:Werner Syndrome Helicase / Q000737:chemistry* / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D006873:Hydroquinones; D008164:Luminescent Proteins; D020543:Proteome; C054562:yellow fluorescent protein, Bacteria; C098785:WRN protein, human; D000071657:Werner Syndrome Helicase; C031927:hydroquinone
keywords: Gerotarget; exonuclease; mass spectrometry; polymorphism; proteomics; werner syndrome