title: Giant yeast cells with nonrecyclable ribonucleotide reductase.
abstract: Ribonucleotide reductase (RNR) catalyzes the reduction of ribonucleotides to deoxyribonucleotides and thereby provides the precursors required for DNA synthesis and repair. In an attempt to test cell resistance to a permanent replicational stress, we constructed a mutant Saccharomyces cerevisiae strain containing exclusively nonrecyclable catalytic subunits of RNR that become inactivated following the reduction of one ribonucleoside diphosphate. In this rnr1C883A rnr3Δ mutant, the synthesis of each deoxyribonucleotide thus requires the production of one Rnr1C883A protein, which means that 26 million Rnr1C883A proteins (half the protein complement of a wild-type cell) have to be produced during each cell cycle. rnr1C883A rnr3Δ cells grow under constant replicational stress, as evidenced by the constitutive activation of the checkpoint effector Rad53, and their S phase is considerably extended compared to the wild type. rnr1C883A rnr3Δ mutants also display additional abnormalities such as a median cell volume increased by a factor of 8, and the presence of massive inclusion bodies. However, they exhibit a good plating efficiency and can be propagated indefinitely. rnr1C883A rnr3Δ cells, which can be used as a protein overexpression system, thus illustrate the robustness of S. cerevisiae to multiple physiological parameters.
authors: Institut de Biologie et de Technologies de Saclay, Service de Biologie Intégrative et de Génétique Moléculaire, Bât. 144, CEA/Saclay, 91191 Gif-sur-Yvette Cedex, France.
authors: Emilie
authors: E
authors: Ma
authors: Arach
authors: A
authors: Goldar
authors: Jean-Marc
authors: JM
authors: Verbavatz
authors: Marie-Claude
authors: MC
authors: Marsolier-Kergoat
mesh_terms: D020134:Catalytic Domain; D002479:Inclusion Bodies / Q000378:metabolism; D009154:Mutation; D012264:Ribonucleotide Reductases / Q000737:chemistry / Q000235:genetics*; D016196:S Phase; D012441:Saccharomyces cerevisiae / Q000166:cytology* / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism
chemical_list: D029701:Saccharomyces cerevisiae Proteins; D012264:Ribonucleotide Reductases