rDNA Copy Number Variants Are Frequent Passenger Mutations in <i>Saccharomyces cerevisiae</i> Deletion Collections and <i>de Novo</i> Transformants

Kwan, Elizabeth; Wang, Xiaobin S; Amemiya, Haley M.; Brewer, Bonita J.; Raghuraman, M. K.

doi:10.1534/g3.116.030296

Cited by 54 publications

(43 citation statements)

References 34 publications

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“…Fungal species vary in the copy number of ribosomal DNA , and this number is independent of genome size or ecological roles, such as guild or trophic mode 91 . Strains of the same fungal species, especially yeast, can exhibit high variation of rDNA copy number 92,93 . Relative abundances of reads are sometimes used as a proxy for the relative biomass contributions of some species 94 .…”

Section: Discussionmentioning

confidence: 99%

Can root-associated fungi mediate the impact of abiotic conditions on the growth of a High Arctic herb?

Wutkowska

Ehrich

Mundra

et al. 2020

Preprint

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Arctic plants are affected by many stressors. Root-associated fungi are thought to influence plant performance in stressful environmental conditions. However, the relationships are not transparent; do the number of fungal partners, their ecological functions and community composition mediate the impact of environmental conditions and/or influence host plant performance? To address these questions, we used a common arctic plant as a model system: Bistorta vivipara . Whole plants (including root system) were collected from nine locations in Spitsbergen (n=214). Morphometric features were measured as a proxy for performance and combined with metabarcoding datasets of their root-associated fungi (amplicon sequence variants, ASVs), edaphic and meteorological variables. Seven biological hypotheses regarding fungal influence on plant measures were tested using structural equation modelling. The best-fitting model revealed that local temperature affected plants both directly (negatively aboveground and positively below-ground) and indirectly -mediated by fungal richness and the ratio of symbio-and saprotrophic ASVs. Fungal community composition did not impact plant measurements and plant reproductive investment did not depend on any fungal parameters. The lack of impact of fungal community composition on plant performance suggests that the functional importance of fungi is more important than their identity. The influence of temperature on host plants is therefore complex and should be examined further.

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Section: Discussionmentioning

confidence: 99%

Can root-associated fungi mediate the impact of abiotic conditions on the growth of a High Arctic herb?

Wutkowska

Ehrich

Mundra

et al. 2020

Preprint

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“…Cells were incubated at 30°C in 1M lithium acetate, 50% PEG-3350, 100μg salmon sperm single-stranded DNA, and 200-700 ng of insertion-targeting DNA for 30 minutes prior to a 15 minute heat-shock at 42°C. Due to changes in rDNA size upon lithium acetate transformation (Kwan et al, 2016), electroporation was used for transforming high-copy rDNA plasmid (pRDN-hyg1 leu2-D) into a ~90-repeat strain (Fig. 2C-D).…”

Section: Methods Detailsmentioning

confidence: 99%

Genomic copy-number loss is rescued by self-limiting production of DNA circles

Mansisidor

Molinar

Srivastava

et al. 2018

Preprint

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SummaryCopy-number changes generate phenotypic variability in health and disease. Whether organisms protect against copy-number changes is largely unknown. Here, we show that Saccharomyces cerevisiae monitors the copy number of its ribosomal DNA (rDNA) and rapidly responds to copy-number loss with the clonal amplification of extrachromosomal rDNA circles (ERCs) from chromosomal repeats. ERC production is proportional to repeat loss and reaches a dynamic steady state that responds to the addition of exogenous rDNA copies. ERC levels are also modulated by RNAPI activity and diet, suggesting that rDNA copy number is calibrated against the cellular demand for rRNA. Lastly, we show that ERCs reinsert into the genome in a dosage-dependent manner, indicating that they provide a reservoir for ultimately increasing rDNA array length. Our results reveal a DNA-based mechanism for rapidly restoring copy number in response to catastrophic gene loss that shares fundamental features with unscheduled copy-number amplifications in cancer cells.

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“…We collected 5 mL of culture for each timed sample then embedded the cells in agarose. We ran the samples on a 1% agarose gel on Bio-Rad CHEF electrophoresis apparatus at 100 V for 68 hours, switch time ramped from 300 to 900 seconds as in Kwan et al (2016). We transferred the gels using standard Southern blotting methods to GeneScreen nylon membrane (PerkinElmer) and probed the same blots multiple times for single copy sequences on different chromosomes, stripping between each hybridization.…”

Section: Methodsmentioning

confidence: 99%

The limiting DNA replication initiation factors Sld2 and Sld3 influence origin efficiency independent of origin firing time

Lynch

Kwan

Alvino

et al. 2018

Preprint

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36Chromosome replication in Saccharomyces cerevisiae is initiated from roughly 300 origins that 37 are regulated both by DNA sequence and by the limited abundance of four trans-acting initiation 38 proteins (Sld2, Sld3, Dpb11 and Dbf4, collectively called "SSDD"). We set out to determine how 39 the association of Sld2 or Sld3 at origins contributes to time of origin activation and/or origin 40 efficiency using auxin-induced protein degradation to further decrease their 41 abundance. Depleting cells of either factor slows growth rate, increases S-phase duration, and 42 causes viability defects, without activating the S phase checkpoint. Chr XII is uniquely unstable 43 with breakage occurring specifically within the rDNA locus. The efficiency of the rDNA origin is 44 decreased while the onset of replication initiation is unchanged. We found that origin efficiency 45 is reduced uniformly across the unique portions of the yeast genome. We conclude that the 46 abundance of Sld2 and Sld3 contribute primarily to origin efficiency. 47 93 2013; Vogelauer et al., 2002). However, the discovery that four specific replication initiation 94 factors (Sld2, Sld3, Dpb11, and Dbf4-the "SSDD" factors) are produced at concentrations 95 significantly lower than the rest of the origin/replisome components suggested that they may be 96 rate-limiting for origin initiation (Mantiero et al., 2011). When Mantiero et al. (2011 97 overexpressed all four proteins simultaneously, the temporal firing of origins was compressed, 98 and a subsequent study revealed that origin efficiency increased genome-wide (McGuffee et al.,99 2013). These findings suggest that the varying ability of different parts of the genome to 100 efficiently recruit the SSDD factors results in the pattern of origin use observed during DNA 101 replication. From their observations, Mantiero et al. (2011) developed a model whereby 102Lynch 2018 4 genome features near origins determine how well origins are able to recruit the limiting SSDD 103 factors, with the earliest and most efficient origins being the most competitive for recruiting 104 SSDD factors. Since each of these four proteins is required only for the initiation phase of DNA 105 replication (Bell and Labib, 2016; Deegan et al., 2016; Muramatsu et al., 2010; Tanaka et al., 106 2007; Zegerman and Diffley, 2007), Mantiero et al. (2011) proposed that as early and/or efficient 107 origins become active and release the limiting factors from the assembling replisome, those 108 factors are recycled to origins that are less competitive for recruiting SSDD and thereby promote 109 origin firing at later and/or more inefficient origins. 110 111 More recently, Collart et al. (2013) demonstrated that the same SSDD factors restrict origin 112 firing during early embryonic development in Xenopus laevis. At the mid-blastula transition 113 (MBT), developing embryos transition from relying on stores of maternally-derived transcripts to 114 activating embryonically-driven transcription (Farrell and O'Farrell, 2014; Langley et al....

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rDNA Copy Number Variants Are Frequent Passenger Mutations in Saccharomyces cerevisiae Deletion Collections and de Novo Transformants

Cited by 54 publications

References 34 publications

Can root-associated fungi mediate the impact of abiotic conditions on the growth of a High Arctic herb?

Can root-associated fungi mediate the impact of abiotic conditions on the growth of a High Arctic herb?

Genomic copy-number loss is rescued by self-limiting production of DNA circles

The limiting DNA replication initiation factors Sld2 and Sld3 influence origin efficiency independent of origin firing time

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