title: Expression of human poly (ADP-ribose) polymerase 1 in Saccharomyces cerevisiae: Effect on survival, homologous recombination and identification of genes involved in intracellular localization.
abstract: The poly (ADP-ribose) polymerase 1 (PARP-1) actively participates in a series of functions within the cell that include: mitosis, intracellular signaling, cell cycle regulation, transcription and DNA damage repair. Therefore, inhibition of PARP1 has a great potential for use in cancer therapy. As resistance to PARP inhibitors is starting to be observed in patients, thus the function of PARP-1 needs to be studied in depth in order to find new therapeutic targets. To gain more information on the PARP-1 activity, we expressed PARP-1 in yeast and investigated its effect on cell growth and UV induced homologous recombination. To identify candidate genes affecting PARP-1 activity and cellular localization, we also developed a yeast genome wide genetic screen. We found that PARP-1 strongly inhibited yeast growth, but when yeast was exposed to the PARP-1 inhibitor 6(5-H) phenantridinone (PHE), it recovered from the growth suppression. Moreover, we showed that PARP-1 produced PAR products in yeast and we demonstrated that PARP-1 reduced UV-induced homologous recombination. By genome wide screening, we identified 99 mutants that suppressed PARP-1 growth inhibition. Orthologues of human genes were found for 41 of these yeast genes. We determined whether the PARP-1 protein level was altered in strains which are deleted for the transcription regulator GAL3, the histone H1 gene HHO1, the HUL4 gene, the deubiquitination enzyme gene OTU1, the nuclear pore protein POM152 and the SNT1 that encodes for the Set3C subunit of the histone deacetylase complex. In these strains the PARP-1 level was roughly the same as in the wild type. PARP-1 localized in the nucleus more in the snt1Δ than in the wild type strain; after UV radiation, PARP-1 localized in the nucleus more in hho1 and pom152 deletion strains than in the wild type indicating that these functions may have a role on regulating PARP-1 level and activity in the nucleus.
authors.affiliation: Yeast Genetics and Genomics, Institute of Clinical Physiology, National Council of Research (CNR), via Moruzzi 1, 56122 Pisa, Italy.
authors.forename: Marco
authors.initials: M
authors.lastname: La Ferla
authors.affiliation: Yeast Genetics and Genomics, Institute of Clinical Physiology, National Council of Research (CNR), via Moruzzi 1, 56122 Pisa, Italy.
authors.forename: Alberto
authors.initials: A
authors.lastname: Mercatanti
authors.affiliation: Yeast Genetics and Genomics, Institute of Clinical Physiology, National Council of Research (CNR), via Moruzzi 1, 56122 Pisa, Italy.
authors.forename: Giulia
authors.initials: G
authors.lastname: Rocchi
authors.affiliation: Yeast Genetics and Genomics, Institute of Clinical Physiology, National Council of Research (CNR), via Moruzzi 1, 56122 Pisa, Italy.
authors.forename: Samuele
authors.initials: S
authors.lastname: Lodovichi
authors.affiliation: Yeast Genetics and Genomics, Institute of Clinical Physiology, National Council of Research (CNR), via Moruzzi 1, 56122 Pisa, Italy.
authors.forename: Tiziana
authors.initials: T
authors.lastname: Cervelli
authors.affiliation: Yeast Genetics and Genomics, Institute of Clinical Physiology, National Council of Research (CNR), via Moruzzi 1, 56122 Pisa, Italy.
authors.forename: Luca
authors.initials: L
authors.lastname: Pignata
authors.affiliation: Section of Genetic Oncology, University Hospital and University of Pisa, via Roma 57, 56125 Pisa, Italy.
authors.forename: Maria Adelaide
authors.initials: MA
authors.lastname: Caligo
authors.affiliation: Yeast Genetics and Genomics, Institute of Clinical Physiology, National Council of Research (CNR), via Moruzzi 1, 56122 Pisa, Italy. Electronic address: alvaro.galli@ifc.cnr.it.
authors.forename: Alvaro
authors.initials: A
authors.lastname: Galli
mesh_terms: D015153:Blotting, Western; D002467:Cell Nucleus / Q000378:metabolism; D010450:Endopeptidases / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D016681:Genome, Fungal / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006657:Histones / Q000235:genetics / Q000378:metabolism; D059765:Homologous Recombination / Q000235:genetics* / Q000528:radiation effects; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D009154:Mutation; D022022:Nuclear Pore / Q000235:genetics / Q000378:metabolism; D010616:Phenanthrenes / Q000494:pharmacology; D000071137:Poly (ADP-Ribose) Polymerase-1; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D011065:Poly(ADP-ribose) Polymerases / Q000235:genetics* / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D019076:Transgenes / Q000235:genetics*; D014466:Ultraviolet Rays
chemical_list: C099289:Gal3 protein, S cerevisiae; C480035:HHO1 protein, S cerevisiae; D006657:Histones; D008562:Membrane Glycoproteins; C086449:POM152 protein, S cerevisiae; D010616:Phenanthrenes; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D049452:Green Fluorescent Proteins; C040601:phenanthridone; C499320:PARP1 protein, human; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; D010450:Endopeptidases; C000592929:Otu1 protein, S cerevisiae
keywords: Homologous recombination; PARP-1 localization; Poly (ADP-ribose) polymerase 1; Saccharomyces cerevisiae; UV; Yeast genome wide screening