Budding yeast ATM/ATR control meiotic double-strand break (DSB) levels by down-regulating Rec114, an essential component of the DSB-machinery

An essential feature of meiosis is Spo11 catalysis of programmed DNA double strand breaks (DSBs). Evidence suggests that the number of DSBs generated per meiosis is genetically determined and that this ability to maintain a pre-determined DSB level, or "DSB homeostasis", might be a property of the meiotic program. Here, we present direct evidence that Rec114, an evolutionarily conserved essential component of the meiotic DSB-machinery, interacts with DSB hotspot DNA, and that Tel1 and Mec1, the budding yeast ATM and ATR, respectively, down-regulate Rec114 upon meiotic DSB formation through phosphorylation. Mimicking constitutive phosphorylation reduces the interaction between Rec114 and DSB hotspot DNA, resulting in a reduction and/or delay in DSB formation. Conversely, a non-phosphorylatable rec114 allele confers a genome-wide increase in both DSB levels and in the interaction between Rec114 and the DSB hotspot DNA. These observations strongly suggest that Tel1 and/or Mec1 phosphorylation of Rec114 following Spo11 catalysis down-regulates DSB formation by limiting the interaction between Rec114 and DSB hotspots. We also present evidence that Ndt80, a meiosis specific transcription factor, contributes to Rec114 degradation, consistent with its requirement for complete cessation of DSB formation. Loss of Rec114 foci from chromatin is associated with homolog synapsis but independent of Ndt80 or Tel1/Mec1 phosphorylation. Taken together, we present evidence for three independent ways of regulating Rec114 activity, which likely contribute to meiotic DSBs-homeostasis in maintaining genetically determined levels of breaks

Temperature-Dependent Modulation of Chromosome Segregation in msh4 Mutants of Budding Yeast

BACKGROUND:In many organisms, homologous chromosomes rely upon recombination-mediated linkages, termed crossovers, to promote their accurate segregation at meiosis I. In budding yeast, the evolutionarily conserved mismatch-repair paralogues, Msh4 and Msh5, promote crossover formation in conjunction with several other proteins, collectively termed the Synapsis Initiation Complex (SIC) proteins or 'ZMM's (Zip1-Zip2-Zip3-Zip4-Spo16, Msh4-Msh5, Mer3). zmm mutants show decreased levels of crossovers and increased chromosome missegregation, which is thought to cause decreased spore viability. PRINCIPAL FINDINGS:In contrast to other ZMM mutants, msh4 and msh5 mutants show improved spore viability and chromosome segregation in response to elevated temperature (23 degrees C versus 33 degrees C). Crossover frequencies in the population of viable spores in msh4 and msh5 mutants are similar at both temperatures, suggesting that temperature-mediated chromosome segregation does not occur by increasing crossover frequencies. Furthermore, meiotic progression defects at elevated temperature do not select for a subpopulation of cells with improved segregation. Instead, another ZMM protein, Zip1, is important for the temperature-dependent improvement in spore viability. CONCLUSIONS:Our data demonstrate interactions between genetic (zmm status) and environmental factors in determining chromosome segregation

Broad-Scale Recombination Patterns Underlying Proper Disjunction in Humans

Although recombination is essential to the successful completion of human meiosis, it remains unclear how tightly the process is regulated and over what scale. To assess the nature and stringency of constraints on human recombination, we examined crossover patterns in transmissions to viable, non-trisomic offspring, using dense genotyping data collected in a large set of pedigrees. Our analysis supports a requirement for one chiasma per chromosome rather than per arm to ensure proper disjunction, with additional chiasmata occurring in proportion to physical length. The requirement is not absolute, however, as chromosome 21 seems to be frequently transmitted properly in the absence of a chiasma in females, a finding that raises the possibility of a back-up mechanism aiding in its correct segregation. We also found a set of double crossovers in surprisingly close proximity, as expected from a second pathway that is not subject to crossover interference. These findings point to multiple mechanisms that shape the distribution of crossovers, influencing proper disjunction in humans

Genome-Wide Analysis of Heteroduplex DNA in Mismatch Repair–Deficient Yeast Cells Reveals Novel Properties of Meiotic Recombination Pathways

Meiotic DNA double-strand breaks (DSBs) initiate crossover (CO) recombination, which is necessary for accurate chromosome segregation, but DSBs may also repair as non-crossovers (NCOs). Multiple recombination pathways with specific intermediates are expected to lead to COs and NCOs. We revisited the mechanisms of meiotic DSB repair and the regulation of CO formation, by conducting a genome-wide analysis of strand-transfer intermediates associated with recombination events. We performed this analysis in a SK1 × S288C Saccharomyces cerevisiae hybrid lacking the mismatch repair (MMR) protein Msh2, to allow efficient detection of heteroduplex DNAs (hDNAs). First, we observed that the anti-recombinogenic activity of MMR is responsible for a 20% drop in CO number, suggesting that in MMR–proficient cells some DSBs are repaired using the sister chromatid as a template when polymorphisms are present. Second, we observed that a large fraction of NCOs were associated with trans–hDNA tracts constrained to a single chromatid. This unexpected finding is compatible with dissolution of double Holliday junctions (dHJs) during repair, and it suggests the existence of a novel control point for CO formation at the level of the dHJ intermediate, in addition to the previously described control point before the dHJ formation step. Finally, we observed that COs are associated with complex hDNA patterns, confirming that the canonical double-strand break repair model is not sufficient to explain the formation of most COs. We propose that multiple factors contribute to the complexity of recombination intermediates. These factors include repair of nicks and double-stranded gaps, template switches between non-sister and sister chromatids, and HJ branch migration. Finally, the good correlation between the strand transfer properties observed in the absence of and in the presence of Msh2 suggests that the intermediates detected in the absence of Msh2 reflect normal intermediates

The pch2Δ Mutation in Baker's Yeast Alters Meiotic Crossover Levels and Confers a Defect in Crossover Interference

Pch2 is a widely conserved protein that is required in baker's yeast for the organization of meiotic chromosome axes into specific domains. We provide four lines of evidence suggesting that it regulates the formation and distribution of crossover events required to promote chromosome segregation at Meiosis I. First, pch2Δ mutants display wild-type crossover levels on a small (III) chromosome, but increased levels on larger (VII, VIII, XV) chromosomes. Second, pch2Δ mutants show defects in crossover interference. Third, crossovers observed in pch2Δ require both Msh4-Msh5 and Mms4-Mus81 functions. Lastly, the pch2Δ mutation decreases spore viability and disrupts crossover interference in spo11 hypomorph strains that have reduced levels of meiosis-induced double-strand breaks. Based on these and previous observations, we propose a model in which Pch2 functions at an early step in crossover control to ensure that every homolog pair receives an obligate crossover

The Synaptonemal Complex Protein Zip1 Promotes Bi-Orientation of Centromeres at Meiosis I

In meiosis I, homologous chromosomes become paired and then separate from one another to opposite poles of the spindle. In humans, errors in this process are a leading cause of birth defects, mental retardation, and infertility. In most organisms, crossing-over, or exchange, between the homologous partners provides a link that promotes their proper, bipolar, attachment to the spindle. Attachment of both partners to the same pole can sometimes be corrected during a delay that is triggered by the spindle checkpoint. Studies of non-exchange chromosomes have shown that centromere pairing serves as an alternative to exchange by orienting the centromeres for proper microtubule attachment. Here, we demonstrate a new role for the synaptonemal complex protein Zip1. Zip1 localizes to the centromeres of non-exchange chromosomes in pachytene and mediates centromere pairing and segregation of the partners at meiosis I. Exchange chromosomes were also found to experience Zip1-dependent pairing at their centromeres. Zip1 was found to persist at centromeres, after synaptonemal complex disassembly, remaining there until microtubule attachment. Disruption of this centromere pairing, in spindle checkpoint mutants, randomized the segregation of exchange chromosomes. These results demonstrate that Zip1-mediated pairing of exchange chromosome centromeres promotes an initial, bipolar attachment of microtubules. This activity of Zip1 lessens the load on the spindle checkpoint, greatly reducing the chance that the cell will exit the checkpoint delay with an improperly oriented chromosome pair. Thus exchange, the spindle checkpoint, and centromere pairing are complementary mechanisms that ensure the proper segregation of homologous partners at meiosis I

A Fine-Structure Map of Spontaneous Mitotic Crossovers in the Yeast Saccharomyces cerevisiae

Homologous recombination is an important mechanism for the repair of DNA damage in mitotically dividing cells. Mitotic crossovers between homologues with heterozygous alleles can produce two homozygous daughter cells (loss of heterozygosity), whereas crossovers between repeated genes on non-homologous chromosomes can result in translocations. Using a genetic system that allows selection of daughter cells that contain the reciprocal products of mitotic crossing over, we mapped crossovers and gene conversion events at a resolution of about 4 kb in a 120-kb region of chromosome V of Saccharomyces cerevisiae. The gene conversion tracts associated with mitotic crossovers are much longer (averaging about 12 kb) than the conversion tracts associated with meiotic recombination and are non-randomly distributed along the chromosome. In addition, about 40% of the conversion events have patterns of marker segregation that are most simply explained as reflecting the repair of a chromosome that was broken in G1 of the cell cycle

Rapid transcriptional plasticity of duplicated gene clusters enables a clonally reproducing aphid to colonise diverse plant species

Background: The prevailing paradigm of host-parasite evolution is that arms races lead to increasing specialisation via genetic adaptation. Insect herbivores are no exception and the majority have evolved to colonise a small number of closely related host species. Remarkably, the green peach aphid, Myzus persicae, colonises plant species across 40 families and single M. persicae clonal lineages can colonise distantly related plants. This remarkable ability makes M. persicae a highly destructive pest of many important crop species. Results: To investigate the exceptional phenotypic plasticity of M. persicae, we sequenced the M. persicae genome and assessed how one clonal lineage responds to host plant species of different families. We show that genetically identical individuals are able to colonise distantly related host species through the differential regulation of genes belonging to aphid-expanded gene families. Multigene clusters collectively upregulate in single aphids within two days upon host switch. Furthermore, we demonstrate the functional significance of this rapid transcriptional change using RNA interference (RNAi)-mediated knock-down of genes belonging to the cathepsin B gene family. Knock-down of cathepsin B genes reduced aphid fitness, but only on the host that induced upregulation of these genes. Conclusions: Previous research has focused on the role of genetic adaptation of parasites to their hosts. Here we show that the generalist aphid pest M. persicae is able to colonise diverse host plant species in the absence of genetic specialisation. This is achieved through rapid transcriptional plasticity of genes that have duplicated during aphid evolution

A new species of Boehlkea (Characiformes: Characidae: Stevardiinae) from the rio Japurá, Amazon basin, Brazil

ABSTRACT A new species of Boehlkea is described from rio Japurá, Amazon basin. The new species differs from B. fredcochui by the presence of a vertically elongate humeral spot (vs. absence), complete lateral line (vs. incomplete), four rows of scales below lateral line (vs. three), and lower number of branched anal-fin rays (17-21 vs. 22-25), and from B. orcesi by the higher number of maxillary teeth (13-14 vs. 5-12), greater head length (27.9-29.9% vs. 24.3-27.5% of SL), and by the color pattern (basal half of dorsal-fin, distal portion of pelvic-fin, lower caudal-fin lobe and anal-fin with black chromatophores vs. absence of black chromatophores in the fins)

Doyne lecture 2016:intraocular health and the many faces of inflammation

Dogma for reasons of immune privilege including sequestration (sic) of ocular antigen, lack of lymphatic and immune competent cells in the vital tissues of the eye has long evaporated. Maintaining tissue and cellular health to preserve vision requires active immune responses to prevent damage and respond to danger. A priori the eye must contain immune competent cells, undergo immune surveillance to ensure homoeostasis as well as an ability to promote inflammation. By interrogating immune responses in non-infectious uveitis and compare with age-related macular degeneration (AMD), new concepts of intraocular immune health emerge. The role of macrophage polarisation in the two disorders is a tractable start. TNF-alpha regulation of macrophage responses in uveitis has a pivotal role, supported via experimental evidence and validated by recent trial data. Contrast this with the slow, insidious degeneration in atrophic AMD or in neovasular AMD, with the compelling genetic association with innate immunity and complement, highlights an ability to attenuate pathogenic immune responses and despite known inflammasome activation. Yolk sac-derived microglia maintains tissue immune health. The result of immune cell activation is environmentally dependent, for example, on retinal cell bioenergetics status, autophagy and oxidative stress, and alterations that skew interaction between macrophages and retinal pigment epithelium (RPE). For example, dead RPE eliciting macrophage VEGF secretion but exogenous IL-4 liberates an anti-angiogenic macrophage sFLT-1 response. Impaired autophagy or oxidative stress drives inflammasome activation, increases cytotoxicity, and accentuation of neovascular responses, yet exogenous inflammasome-derived cytokines, such as IL-18 and IL-33, attenuate responses