scholarly journals A natural variant of the essential host gene MMS21 restricts the parasitic 2-micron plasmid in Saccharomyces cerevisiae

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Michelle Hays ◽  
Janet M Young ◽  
Paula F Levan ◽  
Harmit S Malik
Keyword(s):  
Chromosome Segregation ◽  
Natural Variation ◽  
Plasmid Stability ◽  
Live Cells ◽  
Selfish Genetic Elements ◽  
Plasmid Loss ◽  
Chromatid Cohesion ◽  
Natural Variant ◽  
Trait Locus ◽  
2 Micron Plasmid

Antagonistic coevolution with selfish genetic elements (SGEs) can drive evolution of host resistance. Here, we investigated host suppression of 2-micron (2μ) plasmids, multicopy nuclear parasites that have co-evolved with budding yeasts. We developed SCAMPR (Single-Cell Assay for Measuring Plasmid Retention) to measure copy number heterogeneity and 2μ plasmid loss in live cells. We identified three S. cerevisiae strains that lack endogenous 2μ plasmids and reproducibly inhibit mitotic plasmid stability. Focusing on the Y9 ragi strain, we determined that plasmid restriction is heritable and dominant. Using bulk segregant analysis, we identified a high-confidence Quantitative Trait Locus (QTL) with a single variant of MMS21 associated with increased 2μ instability. MMS21 encodes a SUMO E3 ligase and an essential component of the Smc5/6 complex, involved in sister chromatid cohesion, chromosome segregation, and DNA repair. Our analyses leverage natural variation to uncover a novel means by which budding yeasts can overcome highly successful genetic parasites.

scholarly journals A natural variant of the essential host gene MMS21 restricts the parasitic 2-micron plasmid in Saccharomyces cerevisiae

2020 ◽  
Author(s):  
Michelle Hays ◽  
Janet M. Young ◽  
Paula Levan ◽  
Harmit S. Malik
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Natural Variation ◽  
Plasmid Stability ◽  
Essential Gene ◽  
Single Amino Acid ◽  
Live Cells ◽  
Selfish Genetic Elements ◽  
Single Amino Acid Change ◽  
Natural Variant ◽  
Trait Locus

AbstractOngoing antagonistic coevolution with selfish genetic elements (SGEs) can drive the evolution of host genomes. Here, we investigated whether natural variation allows some Saccharomyces cerevisiae strains to suppress 2-micron (2μ) plasmids, multicopy nuclear parasites that have co-evolved with budding yeasts. To quantitatively measure plasmid stability, we developed a new method, Single-Cell Assay for Measuring Plasmid Retention (SCAMPR) that measures copy number heterogeneity and 2μ plasmid loss in live cells. Next, in a survey of 52 natural S. cerevisiae isolates we identified three strains that lack endogenous 2μ plasmids and reproducibly inhibit mitotic plasmid stability. Thus, their lack of endogenous 2μ plasmids is genetically determined, rather than the result of stochastic loss. Focusing on one isolate (Y9 ragi strain), we determined that plasmid restriction is heritable and dominant. Using bulk segregant analysis, we identified a high-confidence Quantitative Trait Locus (QTL) for mitotic plasmid instability on Y9 chromosome V. We show that a single amino acid change in MMS21 is associated with increased 2-micron instability. MMS21 is an essential gene, encoding a SUMO E3 ligase and a member of the Smc5/6 complex, which is involved in sister chromatid cohesion, chromosome segregation, and DNA repair. Our analyses leverage natural variation to uncover a novel means by which budding yeasts can overcome a highly successful genetic parasite.

scholarly journals The 2 micron plasmid purloins the yeast cohesin complex

2002 ◽  
Vol 158 (4) ◽  
pp. 625-637 ◽  
Author(s):  
Shwetal Mehta ◽  
Xian Mei Yang ◽  
Clarence S. Chan ◽  
Melanie J. Dobson ◽  
Makkuni Jayaram ◽  
...  
Keyword(s):  
Chromosome Segregation ◽  
Plasmid Stability ◽  
High Fidelity ◽  
Cohesin Complex ◽  
Plasmid Segregation ◽  
Rep Proteins ◽  
Dna Plasmid ◽  
2 Micron Plasmid ◽  
Acting In Concert

The yeast 2 micron plasmid achieves high fidelity segregation by coupling its partitioning pathway to that of the chromosomes. Mutations affecting distinct steps of chromosome segregation cause the plasmid to missegregate in tandem with the chromosomes. In the absence of the plasmid stability system, consisting of the Rep1 and Rep2 proteins and the STB DNA, plasmid and chromosome segregations are uncoupled. The Rep proteins, acting in concert, recruit the yeast cohesin complex to the STB locus. The periodicity of cohesin association and dissociation is nearly identical for the plasmid and the chromosomes. The timely disassembly of cohesin is a prerequisite for plasmid segregation. Cohesin-mediated pairing and unpairing likely provides a counting mechanism for evenly partitioning plasmids either in association with or independently of the chromosomes.

scholarly journals Multiple Subunits of the Caenorhabditis elegans Anaphase-Promoting Complex Are Required for Chromosome Segregation During Meiosis I

Genetics ◽  
2002 ◽  
Vol 160 (2) ◽  
pp. 805-813 ◽  
Author(s):  
Edward S Davis ◽  
Lucia Wille ◽  
Barry A Chestnut ◽  
Penny L Sadler ◽  
Diane C Shakes ◽  
...  
Keyword(s):  
Rna Interference ◽  
Caenorhabditis Elegans ◽  
Chromosome Segregation ◽  
Sister Chromatid ◽  
Sister Chromatid Cohesion ◽  
Anaphase Promoting Complex ◽  
Genetic Screens ◽  
Chromatid Cohesion ◽  
Interference Studies ◽  
Meiosis I

Abstract Two genes, originally identified in genetic screens for Caenorhabditis elegans mutants that arrest in metaphase of meiosis I, prove to encode subunits of the anaphase-promoting complex or cyclosome (APC/C). RNA interference studies reveal that these and other APC/C subunits are essential for the segregation of chromosomal homologs during meiosis I. Further, chromosome segregation during meiosis I requires APC/C functions in addition to the release of sister chromatid cohesion.

scholarly journals Double or nothing: a Drosophila mutation affecting meiotic chromosome segregation in both females and males.

Genetics ◽  
1994 ◽  
Vol 136 (3) ◽  
pp. 953-964 ◽  
Author(s):  
D P Moore ◽  
W Y Miyazaki ◽  
J E Tomkiel ◽  
T L Orr-Weaver
Keyword(s):  
Cell Death ◽  
Chromosome Segregation ◽  
Sister Chromatid ◽  
Meiotic Chromosome ◽  
Sister Chromatid Cohesion ◽  
Aberrant Chromosome ◽  
Chromatid Cohesion ◽  
Meiotic Nondisjunction ◽  
Lethal Allele ◽  
Meiosis I

Abstract We describe a Drosophila mutation, Double or nothing (Dub), that causes meiotic nondisjunction in a conditional, dominant manner. Previously isolated mutations in Drosophila specifically affect meiosis either in females or males, with the exception of the mei-S332 and ord genes which are required for proper sister-chromatid cohesion. Dub is unusual in that it causes aberrant chromosome segregation almost exclusively in meiosis I in both sexes. In Dub mutant females both nonexchange and exchange chromosomes undergo nondisjunction, but the effect of Dub on nonexchange chromosomes is more pronounced. Dub reduces recombination levels slightly. Multiple nondisjoined chromosomes frequently cosegregate to the same pole. Dub results in nondisjunction of all chromosomes in meiosis I of males, although the levels are lower than in females. When homozygous, Dub is a conditional lethal allele and exhibits phenotypes consistent with cell death.

scholarly journals Identification of a novel nucleolar protein complex required for mitotic chromosome segregation through centromeric accumulation of Aurora B

2020 ◽  
Vol 48 (12) ◽  
pp. 6583-6596
Author(s):  
Akiko Fujimura ◽  
Yuki Hayashi ◽  
Kazashi Kato ◽  
Yuichiro Kogure ◽  
Mutsuro Kameyama ◽  
...  
Keyword(s):  
Chromosome Segregation ◽  
Protein Complex ◽  
Metaphase Plate ◽  
Nucleolar Protein ◽  
Aurora B ◽  
Mitotic Progression ◽  
Mitotic Chromosomes ◽  
Nucleolar Proteins ◽  
Chromatid Cohesion ◽  
Wd Repeat

Abstract The nucleolus is a membrane-less nuclear structure that disassembles when cells undergo mitosis. During mitosis, nucleolar factors are thus released from the nucleolus and dynamically change their subcellular localization; however, their functions remain largely uncharacterised. Here, we found that a nucleolar factor called nucleolar protein 11 (NOL11) forms a protein complex with two tryptophan-aspartic acid (WD) repeat proteins named WD-repeat protein 43 (WDR43) and Cirhin in mitotic cells. This complex, referred to here as the NWC (NOL11-WDR43-Cirhin) complex, exists in nucleoli during interphase and translocates to the periphery of mitotic chromosomes, i.e., perichromosomal regions. During mitotic progression, both the congression of chromosomes to the metaphase plate and sister chromatid cohesion are impaired in the absence of the NWC complex, as it is required for the centromeric enrichment of Aurora B and the associating phosphorylation of histone H3 at threonine 3. These results reveal the characteristics of a novel protein complex consisting of nucleolar proteins, which is required for regulating kinetochores and centromeres to ensure faithful chromosome segregation.

scholarly journals The yeast S phase checkpoint enables replicating chromosomes to bi-orient and restrain spindle extension during S phase distress

2005 ◽  
Vol 168 (7) ◽  
pp. 999-1012 ◽  
Author(s):  
Jeff Bachant ◽  
Shannon R. Jessen ◽  
Sarah E. Kavanaugh ◽  
Candida S. Fielding
Keyword(s):  
Dna Replication ◽  
Chromosome Segregation ◽  
Replication Fork ◽  
S Phase ◽  
Origin Of Replication ◽  
Independent Manner ◽  
Checkpoint Signaling ◽  
Hydroxyurea Treatment ◽  
Chromatid Cohesion ◽  
S Phase Checkpoint

The budding yeast S phase checkpoint responds to hydroxyurea-induced nucleotide depletion by preventing replication fork collapse and the segregation of unreplicated chromosomes. Although the block to chromosome segregation has been thought to occur by inhibiting anaphase, we show checkpoint-defective rad53 mutants undergo cycles of spindle extension and collapse after hydroxyurea treatment that are distinct from anaphase cells. Furthermore, chromatid cohesion, whose dissolution triggers anaphase, is dispensable for S phase checkpoint arrest. Kinetochore–spindle attachments are required to prevent spindle extension during replication blocks, and chromosomes with two centromeres or an origin of replication juxtaposed to a centromere rescue the rad53 checkpoint defect. These observations suggest that checkpoint signaling is required to generate an inward force involved in maintaining preanaphase spindle integrity during DNA replication distress. We propose that by promoting replication fork integrity under these conditions Rad53 ensures centromere duplication. Replicating chromosomes can then bi-orient in a cohesin-independent manner to restrain untimely spindle extension.

scholarly journals Overexpressed Pituitary Tumor-Transforming Gene Causes Aneuploidy in Live Human Cells

Endocrinology ◽  
2003 ◽  
Vol 144 (11) ◽  
pp. 4991-4998 ◽  
Author(s):  
Run Yu ◽  
Wenge Lu ◽  
Jiandong Chen ◽  
Chris J. McCabe ◽  
Shlomo Melmed
Keyword(s):  
Cancer Cells ◽  
Chromosome Segregation ◽  
Pituitary Tumor ◽  
Fluorescent Protein ◽  
Human Cancer ◽  
Live Cells ◽  
Pituitary Tumor Transforming Gene ◽  
Transforming Gene

Abstract The mammalian securin, pituitary tumor-transforming gene (PTTG), is overexpressed in several tumors and transforms cells in vitro and in vivo. To test the hypothesis that PTTG overexpression causes aneuploidy, enhanced green fluorescent protein (EGFP)-tagged PTTG (PTTG-EGFP) was expressed in human H1299 cancer cells (with undetectable endogenous PTTG expression) and mitosis of individual live cells observed. Untransfected cells and cells expressing EGFP alone exhibited appropriate mitosis. PTTG-EGFP markedly prolonged prophase and metaphase, indicating that PTTG blocks progression of mitosis to anaphase. In cells that underwent apparently normal mitosis (35 of 65 cells), PTTG-EGFP was degraded about 1 min before anaphase onset. Cells that failed to degrade PTTG-EGFP exhibited asymmetrical cytokinesis without chromosome segregation (18 of 65 cells) or chromosome decondensation without cytokinesis (9 of 65 cells), resulting in appearance of a macronucleus. Fifty-one of 55 cells expressing a nondegradable mutant PTTG exhibited asymmetrical cytokinesis without chromosome segregation, and some (4 of 55) decondensed chromosomes, both resulting in macronuclear formation. During this abnormal cytokinesis, all chromosomes and spindles and both centrosomes moved to one daughter cell, suggesting potential chaos in the subsequent mitosis. In conclusion, failure of PTTG degradation or enhanced PTTG accumulation, as a consequence of overexpression, inhibits mitosis progression and chromosome segregation but does not directly affect cytokinesis, resulting in aneuploidy. These results demonstrate that PTTG induces aneuploidy in single, live, human cancer cells.

scholarly journals A quantitative analysis of cohesin decay in mitotic fidelity

2018 ◽  
Vol 217 (10) ◽  
pp. 3343-3353 ◽  
Author(s):  
Sara Carvalhal ◽  
Alexandra Tavares ◽  
Mariana B. Santos ◽  
Mihailo Mirkovic ◽  
Raquel A. Oliveira
Keyword(s):  
Quantitative Analysis ◽  
Chromosome Segregation ◽  
Sister Chromatid ◽  
Mitotic Spindle ◽  
Sister Chromatid Cohesion ◽  
Force Balance ◽  
Chromosome Organization ◽  
Centromeric Chromatin ◽  
Chromatid Cohesion ◽  
Chromosome Alignment

Sister chromatid cohesion mediated by cohesin is essential for mitotic fidelity. It counteracts spindle forces to prevent premature chromatid individualization and random genome segregation. However, it is unclear what effects a partial decline of cohesin may have on chromosome organization. In this study, we provide a quantitative analysis of cohesin decay by inducing acute removal of defined amounts of cohesin from metaphase-arrested chromosomes. We demonstrate that sister chromatid cohesion is very resistant to cohesin loss as chromatid disjunction is only observed when chromosomes lose >80% of bound cohesin. Removal close to this threshold leads to chromosomes that are still cohered but display compromised chromosome alignment and unstable spindle attachments. Partial cohesin decay leads to increased duration of mitosis and susceptibility to errors in chromosome segregation. We propose that high cohesin density ensures centromeric chromatin rigidity necessary to maintain a force balance with the mitotic spindle. Partial cohesin loss may lead to chromosome segregation errors even when sister chromatid cohesion is fulfilled.

scholarly journals ConnectingGCN5’s centromeric SAGA to the mitotic tension-sensing checkpoint

2018 ◽  
Vol 29 (18) ◽  
pp. 2201-2212 ◽  
Author(s):  
Emily L. Petty ◽  
Masha Evpak ◽  
Lorraine Pillus
Keyword(s):  
Chromosome Segregation ◽  
Sister Chromatid ◽  
Spindle Assembly Checkpoint ◽  
Binding Function ◽  
Chromatid Cohesion ◽  
Histone H3 Variant ◽  
Spindle Microtubules ◽  
Low Tension ◽  
Centromere Histone H3 ◽  
Segregation Of Chromosomes

Multiple interdependent mechanisms ensure faithful segregation of chromosomes during cell division. Among these, the spindle assembly checkpoint monitors attachment of spindle microtubules to the centromere of each chromosome, whereas the tension-sensing checkpoint monitors the opposing forces between sister chromatid centromeres for proper biorientation. We report here a new function for the deeply conserved Gcn5 acetyltransferase in the centromeric localization of Rts1, a key player in the tension-sensing checkpoint. Rts1 is a regulatory component of protein phopshatase 2A, a near universal phosphatase complex, which is recruited to centromeres by the Shugoshin (Sgo) checkpoint component under low-tension conditions to maintain sister chromatid cohesion. We report that loss of Gcn5 disrupts centromeric localization of Rts1. Increased RTS1 dosage robustly suppresses gcn5∆ cell cycle and chromosome segregation defects, including restoration of Rts1 to centromeres. Sgo1’s Rts1-binding function also plays a key role in RTS1 dosage suppression of gcn5∆ phenotypes. Notably, we have identified residues of the centromere histone H3 variant Cse4 that function in these chromosome segregation-related roles of RTS1. Together, these findings expand the understanding of the mechanistic roles of Gcn5 and Cse4 in chromosome segregation.

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