scholarly journals Genome-wide haploinsufficiency screen reveals a novel role for γ-TuSC in spindle organization and genome stability

2013 ◽  
Vol 24 (17) ◽  
pp. 2753-2763 ◽  
Author(s):  
John S. Choy ◽  
Eileen O'Toole ◽  
Breanna M. Schuster ◽  
Matthew J. Crisp ◽  
Tatiana S. Karpova ◽  
...  
Keyword(s):  
Chromosome Segregation ◽  
Genome Stability ◽  
Genome Instability ◽  
Electron Tomography ◽  
Gene Dosage ◽  
Microtubule Assembly ◽  
Chromosome Loss ◽  
Genome Wide ◽  
A Genome ◽  
And Function

How subunit dosage contributes to the assembly and function of multimeric complexes is an important question with implications in understanding biochemical, evolutionary, and disease mechanisms. Toward identifying pathways that are susceptible to decreased gene dosage, we performed a genome-wide screen for haploinsufficient (HI) genes that guard against genome instability in Saccharomyces cerevisiae. This led to the identification of all three genes (SPC97, SPC98, and TUB4) encoding the evolutionarily conserved γ-tubulin small complex (γ-TuSC), which nucleates microtubule assembly. We found that hemizygous γ-TuSC mutants exhibit higher rates of chromosome loss and increases in anaphase spindle length and elongation velocities. Fluorescence microscopy, fluorescence recovery after photobleaching, electron tomography, and model convolution simulation of spc98/+ mutants revealed improper regulation of interpolar (iMT) and kinetochore (kMT) microtubules in anaphase. The underlying cause is likely due to reduced levels of Tub4, as overexpression of TUB4 suppressed the spindle and chromosome segregation defects in spc98/+ mutants. We propose that γ-TuSC is crucial for balanced assembly between iMTs and kMTs for spindle organization and accurate chromosome segregation. Taken together, the results show how gene dosage studies provide critical insights into the assembly and function of multisubunit complexes that may not be revealed by using traditional studies with haploid gene deletion or conditional alleles.

scholarly journals A genome-wide screen identifies genes that suppress the accumulation of spontaneous mutations in young and aged yeast cells

2018 ◽  
Author(s):  
Daniele Novarina ◽  
Georges Janssens ◽  
Koen Bokern ◽  
Tim Schut ◽  
Noor van Oerle ◽  
...  
Keyword(s):  
Mutation Rate ◽  
Genome Stability ◽  
Genome Instability ◽  
Spontaneous Mutation ◽  
Yeast Cells ◽  
Spontaneous Mutations ◽  
Genome Maintenance ◽  
Spontaneous Mutation Rate ◽  
Genome Wide ◽  
A Genome

To ensure proper transmission of genetic information, cells need to preserve and faithfully replicate their genome, and failure to do so leads to genome instability, a hallmark of both cancer and aging. Defects in genes involved in guarding genome stability cause several human progeroid syndromes, and an age-dependent accumulation of mutations has been observed in different organisms, from yeast to mammals. However, it is unclear if the spontaneous mutation rate changes during aging, and if specific pathways are important for genome maintenance in old cells. We developed a high-throughput replica-pinning approach to screen for genes important to suppress the accumulation of spontaneous mutations during yeast replicative aging. We found 13 known mutation suppression genes, and 31 genes that had no previous link to spontaneous mutagenesis, and all acted independently of age. Importantly, we identified PEX19, encoding an evolutionarily conserved peroxisome biogenesis factor, as an age-specific mutation suppression gene. While wild-type and pex19Δ young cells have similar spontaneous mutation rates, aged cells lacking PEX19 display an elevated mutation rate. This finding suggests that functional peroxisomes are important to preserve genome integrity specifically in old cells, possibly due to their role in reactive oxygen species metabolism.

scholarly journals Elongation factor ELOF1 drives transcription-coupled repair and prevents genome instability

2021 ◽  
Author(s):  
Marit Geijer ◽  
Di Zhou ◽  
Kathiresan Selvam ◽  
Barbara Steurer ◽  
Bastiaan Evers ◽  
...  
Keyword(s):  
Dna Damage ◽  
Rna Polymerase Ii ◽  
Genome Stability ◽  
Genome Instability ◽  
Excision Repair ◽  
Elongation Factor ◽  
Transcription Inhibition ◽  
Genome Wide ◽  
A Genome ◽  
Nucleotide Excision

Correct transcription is crucial for life. However, DNA damage severely impedes elongating RNA Polymerase II (Pol II), causing transcription inhibition and transcription-replication conflicts. Cells are equipped with intricate mechanisms to counteract the severe consequence of these transcription-blocking lesions (TBLs). However, the exact mechanism and factors involved remain largely unknown. Here, using a genome-wide CRISPR/cas9 screen, we identified elongation factor ELOF1 as an important new factor in the transcription stress response upon DNA damage. We show that ELOF1 has an evolutionary conserved role in Transcription-Coupled Nucleotide Excision Repair (TC-NER), where it promotes recruitment of the TC-NER factors UVSSA and TFIIH to efficiently repair TBLs and resume transcription. Additionally, ELOF1 modulates transcription to protect cells from transcription-mediated replication stress, thereby preserving genome stability. Thus, ELOF1 protects the transcription machinery from DNA damage by two distinct mechanisms.

scholarly journals Emerging Technologies for Genome-Wide Profiling of DNA Breakage

2021 ◽  
Vol 11 ◽  
Author(s):  
Matthew J. Rybin ◽  
Melina Ramic ◽  
Natalie R. Ricciardi ◽  
Philipp Kapranov ◽  
Claes Wahlestedt ◽  
...  
Keyword(s):  
Genome Instability ◽  
Dna Double Strand Breaks ◽  
Single Nucleotide ◽  
Strand Breaks ◽  
Single Strand Breaks ◽  
Genome Wide ◽  
A Genome ◽  
Wide Scale ◽  
Nucleotide Resolution ◽  
Genomic Regions

Genome instability is associated with myriad human diseases and is a well-known feature of both cancer and neurodegenerative disease. Until recently, the ability to assess DNA damage—the principal driver of genome instability—was limited to relatively imprecise methods or restricted to studying predefined genomic regions. Recently, new techniques for detecting DNA double strand breaks (DSBs) and single strand breaks (SSBs) with next-generation sequencing on a genome-wide scale with single nucleotide resolution have emerged. With these new tools, efforts are underway to define the “breakome” in normal aging and disease. Here, we compare the relative strengths and weaknesses of these technologies and their potential application to studying neurodegenerative diseases.

scholarly journals An ancestral recombination graph of human, Neanderthal, and Denisovan genomes

2021 ◽  
Vol 7 (29) ◽  
pp. eabc0776
Author(s):  
Nathan K. Schaefer ◽  
Beth Shapiro ◽  
Richard E. Green
Keyword(s):  
Incomplete Lineage Sorting ◽  
Simulated Data ◽  
Modern Human ◽  
Ancestral Recombination Graph ◽  
Lineage Sorting ◽  
Human Genomes ◽  
Genome Wide ◽  
A Genome ◽  
Graph Inference ◽  
And Function

Many humans carry genes from Neanderthals, a legacy of past admixture. Existing methods detect this archaic hominin ancestry within human genomes using patterns of linkage disequilibrium or direct comparison to Neanderthal genomes. Each of these methods is limited in sensitivity and scalability. We describe a new ancestral recombination graph inference algorithm that scales to large genome-wide datasets and demonstrate its accuracy on real and simulated data. We then generate a genome-wide ancestral recombination graph including human and archaic hominin genomes. From this, we generate a map within human genomes of archaic ancestry and of genomic regions not shared with archaic hominins either by admixture or incomplete lineage sorting. We find that only 1.5 to 7% of the modern human genome is uniquely human. We also find evidence of multiple bursts of adaptive changes specific to modern humans within the past 600,000 years involving genes related to brain development and function.

scholarly journals Heterochromatin: Guardian of the Genome

2018 ◽  
Vol 34 (1) ◽  
pp. 265-288 ◽  
Author(s):  
Aniek Janssen ◽  
Serafin U. Colmenares ◽  
Gary H. Karpen
Keyword(s):  
Chromosome Segregation ◽  
Genome Stability ◽  
Constitutive Heterochromatin ◽  
Repetitive Sequences ◽  
Genome Integrity ◽  
Chromatin Domain ◽  
Cellular Processes ◽  
Eukaryotic Nucleus ◽  
And Function ◽  
Heterochromatin Structure

Constitutive heterochromatin is a major component of the eukaryotic nucleus and is essential for the maintenance of genome stability. Highly concentrated at pericentromeric and telomeric domains, heterochromatin is riddled with repetitive sequences and has evolved specific ways to compartmentalize, silence, and repair repeats. The delicate balance between heterochromatin epigenetic maintenance and cellular processes such as mitosis and DNA repair and replication reveals a highly dynamic and plastic chromatin domain that can be perturbed by multiple mechanisms, with far-reaching consequences for genome integrity. Indeed, heterochromatin dysfunction provokes genetic turmoil by inducing aberrant repeat repair, chromosome segregation errors, transposon activation, and replication stress and is strongly implicated in aging and tumorigenesis. Here, we summarize the general principles of heterochromatin structure and function, discuss the importance of its maintenance for genome integrity, and propose that more comprehensive analyses of heterochromatin roles in tumorigenesis will be integral to future innovations in cancer treatment.

Analysis of MicroRNA Transcriptomes Insights into the miR-148a-3p Inducer of Rumen Development in Goats

2020 ◽  
Author(s):  
Tao Zhong ◽  
Cheng Wang ◽  
Jiangtao Hu ◽  
Xiaoyong Chen ◽  
Lili Niu ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Target Genes ◽  
Genetic Regulation ◽  
Mapk Signaling ◽  
Regulation Mechanism ◽  
Ras Signaling ◽  
Genome Wide ◽  
A Genome ◽  
Differentially Expressed Mirnas ◽  
And Function

Abstract Background: Rumen is an important digestive organ of ruminant. From fetal to adult stage, the morphology, structure and function of rumen have changed significantly. But the intrinsic genetic regulation is still limited. We previously reported a genome-wide expression profile of miRNAs in prenatal goat rumens. In the present study, we rejoined analyzed the transcriptomes of rumen miRNAs during prenatal (E60 and E135) and postnatal (D30 and D150) stages.Results: A total of 66 differentially expressed miRNAs (DEMs) were identified in the rumen tissues from D30 and D150 goats. Of these, 17 DEMs were consistently highly expressed in the rumens at the preweaning stages (E60, E135 and D30), while down-regulated at D150. Noteworthy, annotation analysis revealed that the target genes regulated by the DEMs were mainly enriched in MAPK signaling pathway, Jak-STAT signaling pathway and Ras signaling pathway. Interestingly, the expression of miR-148a-3p was significantly high in the embryonic stage and down-regulated at D150. The potential binding sites between miR-148a-3p and QKI were predicted by the TargetScan and verified by the dual luciferase report assay. The co-localization of miR-148a-3p and QKI was observed not in intestinal tracts but in rumen tissues by in situ hybridization. Moreover, the expression of miR-148a-3p in the epithelium was significantly higher than that in the other layers, suggesting that miR-148a-3p involve in the development of rumen epithelial cells by targeting QKI. Subsequently, miR-148a-3p inhibitor was found to induce the proliferation of GES-1 cells.Conclusions: Taken together, these results identified the DEMs involved in the development of rumen and provided an insight into the regulation mechanism of goat rumens during development.

scholarly journals Identification and function of hypoxia-response genes in Drosophila melanogaster

2006 ◽  
Vol 25 (1) ◽  
pp. 134-141 ◽  
Author(s):  
Guowen Liu ◽  
Julianne Roy ◽  
Eric A. Johnson
Keyword(s):  
Recovery Time ◽  
Normal Activity ◽  
Mrna Levels ◽  
Low Oxygen ◽  
Hypoxia Response ◽  
Oxygen Treatment ◽  
Genome Wide ◽  
A Genome ◽  
And Function ◽  
Genome Wide Study

Hypoxia, an insufficient level of oxygen in the cell, occurs during normal activity and also in pathological conditions such as ischemia and tumorigenesis. Although many hypoxia-response genes have been identified, an understanding of the functional role for these genes in the living animal is lacking. Here we present a genome-wide study of gene expression changes during hypoxia and then functionally test a subset of these genes for roles in survival and recovery from hypoxia. We found 79 genes with increased mRNA levels when adult flies were treated with 0.5% O2 for 6 h. A subset of these genes had detectably increased levels in as short as 1 h of low-oxygen treatment. Mild hypoxia levels resulted in an increase in transcription levels for only 20 genes. Viability during hypoxia and recovery time from hypoxia-induced paralysis was examined in flies with a reduction in activity in hypoxia-response genes. The observed decreased viability and increased recovery time from paralysis in many of the lines demonstrate that the increased transcript levels seen after hypoxia are important for the response to low oxygen.

scholarly journals The yeast core spliceosome maintains genome integrity through R-loop prevention and α-tubulin expression

2018 ◽  
Author(s):  
Annie S. Tam ◽  
Veena Mathew ◽  
Tianna S. Sihota ◽  
Anni Zhang ◽  
Peter C. Stirling
Keyword(s):  
Gene Expression ◽  
Dna Replication ◽  
Chromosome Segregation ◽  
Genome Stability ◽  
Spindle Assembly Checkpoint ◽  
Genome Instability ◽  
Genome Integrity ◽  
Genome Maintenance ◽  
Maintenance Factors ◽  
Spindle Assembly Checkpoint Protein

To achieve genome stability cells must coordinate the action of various DNA transactions including DNA replication, repair, transcription and chromosome segregation. How transcription and RNA processing enable genome stability is only partly understood. Two predominant models have emerged: one involving changes in gene expression that perturb other genome maintenance factors, and another in which genotoxic DNA:RNA hybrids, called R-loops, impair DNA replication. Here we characterize genome instability phenotypes in a panel yeast splicing factor mutants and find that mitotic defects, and in some cases R-loop accumulation, are causes of genome instability. Genome instability in splicing mutants is exacerbated by loss of the spindle-assembly checkpoint protein Mad1. Moreover, removal of the intron from the α-tubulin gene TUB1 restores genome integrity. Thus, while R-loops contribute in some settings, defects in yeast splicing predominantly lead to genome instability through effects on gene expression.

scholarly journals MRE11-RAD50-NBS1 activates Fanconi Anemia R-loop suppression at transcription-replication conflicts

2018 ◽  
Author(s):  
Emily Yun-chia Chang ◽  
James P. Wells ◽  
Shu-Huei Tsai ◽  
Yan Coulombe ◽  
Yujia A. Chan ◽  
...  
Keyword(s):  
Dna Replication ◽  
Fanconi Anemia ◽  
Replication Fork ◽  
Genome Instability ◽  
Human Cancer ◽  
Replication Stress ◽  
Maintenance Factors ◽  
Genome Wide ◽  
A Genome ◽  
Dna Replication Stress

SUMMARYEctopic R-loop accumulation causes DNA replication stress and genome instability. To avoid these outcomes, cells possess a range of anti-R-loop mechanisms, including RNaseH that degrades the RNA moiety in R-loops. To comprehensively identify anti-R-loop mechanisms, we performed a genome-wide trigenic interaction screen in yeast lacking RNH1 and RNH201. We identified >100 genes critical for fitness in the absence of RNaseH, which were enriched for DNA replication fork maintenance factors such as RAD50. We show in yeast and human cells that R-loops accumulate during RAD50 depletion. In human cancer cell models, we find that RAD50 and its partners in the MRE11-RAD50-NBS1 complex regulate R-loop-associated DNA damage and replication stress. We show that a non-nucleolytic function of MRE11 is important for R-loop suppression via activation of PCNA-ubiquitination by RAD18 and recruiting anti-R-loop helicases in the Fanconi Anemia pathway. This work establishes a novel role for MRE11-RAD50-NBS1 in directing tolerance mechanisms of transcription-replication conflicts.

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