Impaired manganese metabolism causes mitotic misregulation.

Manganese is an essential trace element, whose intracellular levels need to be carefully regulated. Mn(2+) acts as a cofactor for many enzymes and excess of Mn(2+) is toxic. Alterations in Mn(2+) homeostasis affect metabolic functions and mutations in the human Mn(2+)/Ca(2+) transporter ATP2C1 have been linked to Hailey-Hailey disease. By deletion of the yeast orthologue PMR1 we have studied the impact of Mn(2+) on cell cycle progression and show that an excess of cytosolic Mn(2+) alters S-phase transit, induces transcriptional up-regulation of cell cycle regulators, bypasses the need for S-phase cell cycle checkpoints and predisposes to genomic instability. On the other hand, we find that depletion of the Golgi Mn(2+) pool requires a functional morphology checkpoint to avoid the formation of polyploid cells.

The ribonucleotide reductase inhibitor, Sml1, is sequentially phosphorylated, ubiquitylated and degraded in response to DNA damage.

Regulation of ribonucleotide reductase (RNR) is important for cell survival and genome integrity in the face of genotoxic stress. The Mec1/Rad53/Dun1 DNA damage response kinase cascade exhibits multifaceted controls over RNR activity including the regulation of the RNR inhibitor, Sml1. After DNA damage, Sml1 is degraded leading to the up-regulation of dNTP pools by RNR. Here, we probe the requirements for Sml1 degradation and identify several sites required for in vivo phosphorylation and degradation of Sml1 in response to DNA damage. Further, in a strain containing a mutation in Rnr1, rnr1-W688G, mutation of these sites in Sml1 causes lethality. Degradation of Sml1 is dependent on the 26S proteasome. We also show that degradation of phosphorylated Sml1 is dependent on the E2 ubiquitin-conjugating enzyme, Rad6, the E3 ubiquitin ligase, Ubr2, and the E2/E3-interacting protein, Mub1, which form a complex previously only implicated in the ubiquitylation of Rpn4.

Nuclear-mitochondrial epistasis for fitness in Saccharomyces cerevisiae.

In addition to the familiar possibility of epistasis between nuclear loci, interactions may evolve between the mitochondrial and nuclear genomes in eukaryotic cells. We looked for such interactions in Saccharomyces cerevisiae genotypes evolved independently and asexually in the laboratory for 2000 generations, and in an ecologically distinct pathogenic S. cerevisiae strain. From these strains we constructed derivatives entirely lacking mitochondrial DNA and then used crosses to construct matched and unmatched pairings of nuclear and mitochondrial genomes. We detected fitness effects of such interactions in an evolved laboratory strain and in crosses between the laboratory and pathogen strains. In both cases, there were significant contributions to progeny fitness of both nuclear and mitochondrial genomes and of their interaction. A second evolved genotype showed incompatibility with the first evolved genotype, but the nuclear and mitochondrial contributions to this incompatibility could not be resolved. These results indicate that cytonuclear interactions analogous to those already known from plants and animals can evolve rapidly on an evolutionary timescale.