title: 129-Derived Mouse Strains Express an Unstable but Catalytically Active DNA Polymerase Iota Variant.
abstract: Mice derived from the 129 strain have a nonsense codon mutation in exon 2 of the polymerase iota (Polι) gene and are therefore considered Polι deficient. When we amplified Polι mRNA from 129/SvJ or 129/Ola testes, only a small fraction of the full-length cDNA contained the nonsense mutation; the major fraction corresponded to a variant Polι isoform lacking exon 2. Polι mRNA lacking exon 2 contains an open reading frame, and the corresponding protein was detected using a polyclonal antibody raised against the C terminus of the murine Polι protein. The identity of the corresponding protein was further confirmed by mass spectrometry. Although the variant protein was expressed at only 5 to 10% of the level of wild-type Polι, it retained de novo DNA synthesis activity, the capacity to form replication foci following UV irradiation, and the ability to rescue UV light sensitivity in Polι(-/-) embryonic fibroblasts derived from a new, fully deficient Polι knockout (KO) mouse line. Furthermore, in vivo treatment of 129-derived male mice with Velcade, a drug that inhibits proteasome function, stabilized and restored a substantial amount of the variant Polι in these animals, indicating that its turnover is controlled by the proteasome. An analysis of two xeroderma pigmentosum-variant (XPV) cases corresponding to missense mutants of Polη, a related translesion synthesis (TLS) polymerase in the same family, similarly showed a destabilization of the catalytically active mutant protein by the proteasome. Collectively, these data challenge the prevailing hypothesis that 129-derived strains of mice are completely deficient in Polι activity. The data also document, both for 129-derived mouse strains and for some XPV patients, new cases of genetic defects corresponding to the destabilization of an otherwise functional protein, the phenotype of which is reversible by proteasome inhibition.
authors.affiliation: Centre National de la Recherche Scientifique UMR 8200, Institut Gustave Roussy, Villejuif, and Université Paris-Sud, Orsay, France said.aoufouchi@gustaveroussy.fr.
authors.forename: Said
authors.initials: S
authors.lastname: Aoufouchi
authors.affiliation: Institut Necker-Enfants Malades, INSERM U1151-CNRS UMR 8253, Sorbonne Paris Cité, Université Paris Descartes, Faculté de Médecine-Site Broussais, Paris, France.
authors.forename: Annie
authors.initials: A
authors.lastname: De Smet
authors.affiliation: Institut Necker-Enfants Malades, INSERM U1151-CNRS UMR 8253, Sorbonne Paris Cité, Université Paris Descartes, Faculté de Médecine-Site Broussais, Paris, France.
authors.forename: Frédéric
authors.initials: F
authors.lastname: Delbos
authors.affiliation: Centre National de la Recherche Scientifique UMR 8200, Institut Gustave Roussy, Villejuif, and Université Paris-Sud, Orsay, France.
authors.forename: Camille
authors.initials: C
authors.lastname: Gelot
authors.affiliation: Plateau Protéomes Necker, PPN, Université Paris Descartes, Structure Fédérative de Recherche Necker, Paris, France.
authors.forename: Ida Chiara
authors.initials: IC
authors.lastname: Guerrera
authors.affiliation: Institut Necker-Enfants Malades, INSERM U1151-CNRS UMR 8253, Sorbonne Paris Cité, Université Paris Descartes, Faculté de Médecine-Site Broussais, Paris, France.
authors.forename: Jean-Claude
authors.initials: JC
authors.lastname: Weill
authors.affiliation: Institut Necker-Enfants Malades, INSERM U1151-CNRS UMR 8253, Sorbonne Paris Cité, Université Paris Descartes, Faculté de Médecine-Site Broussais, Paris, France.
authors.forename: Claude-Agnès
authors.initials: CA
authors.lastname: Reynaud
mesh_terms: D000818:Animals; D001483:Base Sequence; D001897:Boronic Acids / Q000494:pharmacology; D000069286:Bortezomib; D002460:Cell Line; D004249:DNA Damage / Q000235:genetics; D004260:DNA Repair / Q000235:genetics*; D004261:DNA Replication / Q000235:genetics*; D004259:DNA-Directed DNA Polymerase / Q000235:genetics* / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D046988:Proteasome Endopeptidase Complex / Q000187:drug effects; D061988:Proteasome Inhibitors / Q000494:pharmacology; D020033:Protein Isoforms / Q000096:biosynthesis / Q000235:genetics; D011719:Pyrazines / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D017422:Sequence Analysis, DNA; D054875:Ubiquitination; D014983:Xeroderma Pigmentosum / Q000235:genetics*; D000094769:DNA Polymerase iota; D000099270:Y-Family DNA Polymerases
chemical_list: D001897:Boronic Acids; D000069286:Bortezomib; D004259:DNA-Directed DNA Polymerase; D046988:Proteasome Endopeptidase Complex; D061988:Proteasome Inhibitors; D020033:Protein Isoforms; D011719:Pyrazines; D012333:RNA, Messenger; D000094769:DNA Polymerase iota; C112338:Rad30 protein; D000099270:Y-Family DNA Polymerases