Destabilization of the replication fork protection complex disrupts meiotic chromosome segregation

Escorcia, Wilber; Forsburg, Susan L.

doi:10.1091/mbc.e17-02-0101

Cited by 11 publications

(29 citation statements)

References 118 publications

(239 reference statements)

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“…Thus, meiosis is associated with nuclear size fluctuations when homologous chromosomes align and recombine and with nuclear size reductions following two rounds of chromosome separation. Alleles that change these nuclear dynamics are likely associated with genome stability regulation in meiosis 12,13 .…”

Section: Representative Resultsmentioning

confidence: 99%

“…The Rec8-GFP marker is, thus, useful to examine conditions that disrupt the establishment, removal, and overall stability of meiotic cohesion. Paired with a marker of nuclear division such as Hht1-mRFP 5,13 , Rec8-GFP can be employed to address questions of how cohesion timing and stability prior to and during meiosis affect proper chromosome segregation 12,13 . This protocol does not address proteins whose dynamics occur primarily in the cytosol.…”

Section: Representative Resultsmentioning

confidence: 99%

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Examination of Mitotic and Meiotic Fission Yeast Nuclear Dynamics by Fluorescence Live-cell Microscopy

Escorcia

Shen

Yuan

et al. 2019

JoVE

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Live-cell imaging is a microscopy technique used to examine cell and protein dynamics in living cells. This imaging method is not toxic, generally does not interfere with cell physiology, and requires minimal experimental handling. The low levels of technical interference enable researchers to study cells across multiple cycles of mitosis and to observe meiosis from beginning to end. Using fluorescent tags such as Green Fluorescent Protein (GFP) and Red Fluorescent Protein (RFP), researchers can analyze different factors whose functions are important for processes like transcription, DNA replication, cohesion, and segregation. Coupled with data analysis using Fiji (a free, optimized ImageJ version), live-cell imaging offers various ways of assessing protein movement, localization, stability, and timing, as well as nuclear dynamics and chromosome segregation. However, as is the case with other microscopy methods, live-cell imaging is limited by the intrinsic properties of light, which put a limit to the resolution power at high magnifications, and is also sensitive to photobleaching or phototoxicity at high wavelength frequencies. However, with some care, investigators can bypass these physical limitations by carefully choosing the right conditions, strains, and fluorescent markers to allow for the appropriate visualization of mitotic and meiotic events.

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Section: Representative Resultsmentioning

confidence: 99%

Section: Representative Resultsmentioning

confidence: 99%

Examination of Mitotic and Meiotic Fission Yeast Nuclear Dynamics by Fluorescence Live-cell Microscopy

Escorcia

Shen

Yuan

et al. 2019

JoVE

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“…Likewise, perturbed DNA replication (GO:0006260) can indirectly lead to problems with chromosome segregation (GO:0007059), due to the presence of DNA structures that cannot be separated (e.g. [29]. A chromosome segregation phenotype alone therefore does not suffice to confidently annotate a gene product as involved in chromosome segregation.…”

Section: Annotation Error Typesmentioning

confidence: 99%

Term Matrix: A novel Gene Ontology annotation quality control system based on ontology term co-annotation patterns

Wood

Carbon

Harris

et al. 2020

Preprint

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Biological processes are accomplished by the coordinated action of gene products. Gene products often participate in multiple processes, and can therefore be annotated to multiple Gene Ontology (GO) terms. Nevertheless, processes that are functionally, temporally, and/or spatially distant may have few gene products in common, and co-annotation to unrelated processes likely reflects errors in literature curation, ontology structure, or automated annotation pipelines. We have developed an annotation quality control workflow that uses rules based on mutually exclusive processes to detect annotation errors, based on and validated by case studies including the three we present here: fission yeast protein-coding gene annotations over time; annotations for cohesin complex subunits in human and model species; and annotations using a selected set of GO biological process terms in human and five model species. For each case study, we reviewed available GO annotations, identified pairs of biological processes which are unlikely to be correctly co-annotated to the same gene products (e.g., amino acid metabolism and cytokinesis), and traced erroneous annotations to their sources. To date we have generated 107 quality control rules, and corrected 289 manual annotations in eukaryotes and over 2.5 million automatically propagated annotations across all taxa.

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“…There are excellent markers for mitotic landmarks including fluorescently tagged spindle pole body protein Sad1 (King and Drivas 2008) or tubulin (Sawin and Tran 2006), and septation is easily examined under light microscopy (Minet et al 1979). We have developed and employed numerous tools to identify and characterize features of DNA synthesis and replication stress, including fluorescent tagged RPA and Rad52 proteins (Sabatinos et al 2012; Sabatinos and Forsburg 2015a;; Escorcia and Forsburg 2017), and have also examined abnormal mitotic divisions in response to replication stress including nuclear envelope, cell membrane, and histone markers (Sabatinos et al 2012; Escorcia and Forsburg 2017).…”

Section: Introductionmentioning

confidence: 99%

Checkpoint regulation of nuclear Tos4 defines S phase arrest in fission yeast

Kim

Tripathi

Shen

et al. 2019

Preprint

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From yeast to humans, the cell cycle is tightly controlled by regulatory networks that regulate cell proliferation and can be monitored by dynamic visual markers in living cells. We have observed S phase progression by monitoring nuclear accumulation of the FHA-containing DNA binding protein Tos4, which is expressed in the G1/S phase transition. We use Tos4 localization to distinguish three classes of DNA replication mutants: those that arrest with an apparent 1C DNA content and accumulate Tos4 at the restrictive temperature;; those that arrest with an apparent 2C DNA content, that do not accumulate Tos4;; and those that proceed into mitosis despite a 1C DNA content, again without Tos4 accumulation. Our data indicate that Tos4 localization in these conditions is responsive to checkpoint kinases, with activation of the Cds1 checkpoint kinase promoting Tos4 retention in the nucleus, and activation of the Chk1 damage checkpoint promoting its turnover. Tos4 localization therefore allows us to monitor checkpoint-dependent activation that responds to replication failure in early versus late S phase.

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Destabilization of the replication fork protection complex disrupts meiotic chromosome segregation

Cited by 11 publications

References 118 publications

Examination of Mitotic and Meiotic Fission Yeast Nuclear Dynamics by Fluorescence Live-cell Microscopy

Examination of Mitotic and Meiotic Fission Yeast Nuclear Dynamics by Fluorescence Live-cell Microscopy

Term Matrix: A novel Gene Ontology annotation quality control system based on ontology term co-annotation patterns

Checkpoint regulation of nuclear Tos4 defines S phase arrest in fission yeast

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