A 700 bp cis-Acting Region Controls Mating-Type Dependent Recombination Along the Entire Left Arm of Yeast Chromosome III

AbstractHomothallic switching of the mating-type MAT a gene in Saccharomyces cerevisiae results from replacement by gene conversion of MAT-Ya DNA with Yα sequences copied from one of two unexpressed donors. MAT a preferentially recombines with HMLα, located near the left end of chromosome III, but can use HMRα, near the right chromosome end. MAT a donor preference depends on a 700 bp orientation-independent cis-acting recombination enhancer, located 17 kb proximal to HML. Deletion of this element markedly reduces MAT a's use of a donor inserted at any of four different locations along the leftmost 92 kb of chromosome III. This enhancer is sufficient for donor activation, since it stimulates use of the “wrong” donor, when it is inserted 7 kb proximal to HMR

Implications of the Partial Width Z->bb for Supersymmetry Searches and Model-Building

Assuming that the actual values of the top quark mass at FNAL and of the ratio of partial widths Z->bb/Z->hadrons at LEP are within their current one-sigma reported ranges, we present a No-Lose Theorem for superpartner searches at LEP II and an upgraded Tevatron. We impose only two theoretical assumptions: the Lagrangian is that of the Minimal Supersymmetric Standard Model with arbitrary soft-breaking terms, and all couplings remain perturbative up to scales of order 10^16 GeV; there are no assumptions about the soft SUSY breaking parameters, proton decay, cosmology, etc. In particular, if the LEP and FNAL values hold up and supersymmetry is responsible for the discrepancy with the SM prediction of the partial width of Z->bb, then we must have charginos and/or top squarks observable at the upgraded machines. Furthermore, little deviation from the SM is predicted within "super-unified" SUSY. Finally, it appears to be extremely difficult to find any unified MSSM model, regardless of the form of soft SUSY breaking, that can explain the partial width for large tan(beta); in particular, no model with top-bottom-tau Yukawa coupling unification appears to be consistent with the experiments.Comment: 15 pages, University of Michigan preprint UM-TH-94-23. LaTeX file with 4 uuencoded figures sent separately. Compressed PS file (114Kb) available by anonymous FTP from 141.211.96.66 in /pub/preprints/UM-TH-94-23.ps.

Evidence for Domesticated and Wild Populations of Saccharomyces cerevisiae

Saccharomyces cerevisiae is predominantly found in association with human activities, particularly the production of alcoholic beverages. S. paradoxus, the closest known relative of S. cerevisiae, is commonly found on exudates and bark of deciduous trees and in associated soils. This has lead to the idea that S. cerevisiae is a domesticated species, specialized for the fermentation of alcoholic beverages, and isolates of S. cerevisiae from other sources simply represent migrants from these fermentations. We have surveyed DNA sequence diversity at five loci in 81 strains of S. cerevisiae that were isolated from a variety of human and natural fermentations as well as sources unrelated to alcoholic beverage production, such as tree exudates and immunocompromised patients. Diversity within vineyard strains and within saké strains is low, consistent with their status as domesticated stocks. The oldest lineages and the majority of variation are found in strains from sources unrelated to wine production. We propose a model whereby two specialized breeds of S. cerevisiae have been created, one for the production of grape wine and one for the production of saké wine. We estimate that these two breeds have remained isolated from one another for thousands of years, consistent with the earliest archeological evidence for winemaking. We conclude that although there are clearly strains of S. cerevisiae specialized for the production of alcoholic beverages, these have been derived from natural populations unassociated with alcoholic beverage production, rather than the opposite

Mapping Meiotic Single-Strand DNA Reveals a New Landscape of DNA Double-Strand Breaks in Saccharomyces cerevisiae

DNA double-strand breaks (DSBs), which are formed by the Spo11 protein, initiate meiotic recombination. Previous DSB-mapping studies have used rad50S or sae2Δ mutants, which are defective in break processing, to accumulate Spo11-linked DSBs, and report large (≥ 50 kb) “DSB-hot” regions that are separated by “DSB-cold” domains of similar size. Substantial recombination occurs in some DSB-cold regions, suggesting that DSB patterns are not normal in rad50S or sae2Δ mutants. We therefore developed a novel method to map genome-wide, single-strand DNA (ssDNA)–associated DSBs that accumulate in processing-capable, repair-defective dmc1Δ and dmc1Δ rad51Δ mutants. DSBs were observed at known hot spots, but also in most previously identified “DSB-cold” regions, including near centromeres and telomeres. Although approximately 40% of the genome is DSB-cold in rad50S mutants, analysis of meiotic ssDNA from dmc1Δ shows that most of these regions have substantial DSB activity. Southern blot assays of DSBs in selected regions in dmc1Δ, rad50S, and wild-type cells confirm these findings. Thus, DSBs are distributed much more uniformly than was previously believed. Comparisons of DSB signals in dmc1, dmc1 rad51, and dmc1 spo11 mutant strains identify Dmc1 as a critical strand-exchange activity genome-wide, and confirm previous conclusions that Spo11-induced lesions initiate all meiotic recombination

Genome-Wide Analysis of Rad52 Foci Reveals Diverse Mechanisms Impacting Recombination

To investigate the DNA damage response, we undertook a genome-wide study in Saccharomyces cerevisiae and identified 86 gene deletions that lead to increased levels of spontaneous Rad52 foci in proliferating diploid cells. More than half of the genes are conserved across species ranging from yeast to humans. Along with genes involved in DNA replication, repair, and chromatin remodeling, we found 22 previously uncharacterized open reading frames. Analysis of recombination rates and synthetic genetic interactions with rad52Δ suggests that multiple mechanisms are responsible for elevated levels of spontaneous Rad52 foci, including increased production of recombinogenic lesions, sister chromatid recombination defects, and improper focus assembly/disassembly. Our cell biological approach demonstrates the diversity of processes that converge on homologous recombination, protect against spontaneous DNA damage, and facilitate efficient repair

Mre11–Rad50–Nbs1-dependent processing of DNA breaks generates oligonucleotides that stimulate ATM activity

DNA double-strand breaks (DSBs) can be processed by the Mre11–Rad50–Nbs1 (MRN) complex, which is essential to promote ataxia telangiectasia-mutated (ATM) activation. However, the molecular mechanisms linking MRN activity to ATM are not fully understood. Here, using Xenopus laevis egg extract we show that MRN-dependent processing of DSBs leads to the accumulation of short single-stranded DNA oligonucleotides (ssDNA oligos). The MRN complex isolated from the extract containing DSBs is bound to ssDNA oligos and stimulates ATM activity. Elimination of ssDNA oligos results in rapid extinction of ATM activity. Significantly, ssDNA oligos can be isolated from human cells damaged with ionizing radiation and injection of small synthetic ssDNA oligos into undamaged cells also induces ATM activation. These results suggest that MRN-dependent generation of ssDNA oligos, which constitute a unique signal of ongoing DSB repair not encountered in normal DNA metabolism, stimulates ATM activity