G. aculeatus BioNano Irys optical genome map

G. aculeatus BioNano Irys optical genome ma

Unassigned contigs narrowed to linkage groups

1,263 previously unassigned contigs narrowed to linkage group

The Fitness Consequences of Aneuploidy Are Driven by Condition-Dependent Gene Effects

<div><p>Aneuploidy is a hallmark of tumor cells, and yet the precise relationship between aneuploidy and a cell’s proliferative ability, or cellular fitness, has remained elusive. In this study, we have combined a detailed analysis of aneuploid clones isolated from laboratory-evolved populations of <i>Saccharomyces cerevisiae</i> with a systematic, genome-wide screen for the fitness effects of telomeric amplifications to address the relationship between aneuploidy and cellular fitness. We found that aneuploid clones rise to high population frequencies in nutrient-limited evolution experiments and show increased fitness relative to wild type. Direct competition experiments confirmed that three out of four aneuploid events isolated from evolved populations were themselves sufficient to improve fitness. To expand the scope beyond this small number of exemplars, we created a genome-wide collection of >1,800 diploid yeast strains, each containing a different telomeric amplicon (Tamp), ranging in size from 0.4 to 1,000 kb. Using pooled competition experiments in nutrient-limited chemostats followed by high-throughput sequencing of strain-identifying barcodes, we determined the fitness effects of these >1,800 Tamps under three different conditions. Our data revealed that the fitness landscape explored by telomeric amplifications is much broader than that explored by single-gene amplifications. As also observed in the evolved clones, we found the fitness effects of most Tamps to be condition specific, with a minority showing common effects in all three conditions. By integrating our data with previous work that examined the fitness effects of single-gene amplifications genome-wide, we found that a small number of genes within each Tamp are centrally responsible for each Tamp’s fitness effects. Our genome-wide Tamp screen confirmed that telomeric amplifications identified in laboratory-evolved populations generally increased fitness. Our results show that Tamps are mutations that produce large, typically condition-dependent changes in fitness that are important drivers of increased fitness in asexually evolving populations.</p></div

Aneuploidy variably affects fitness of evolved clones.

<p>The individual fitness effects of point mutations and aneuploid events were determined for all mutations identified in the evolved clones S8c2 (A), P5c3 (B), and P6c1 (C). The supernumerary chromosome(s) in each clone are labeled by their identifying translocation or, in the case of the chromosome XIII disomy, as “Chr XIII.” Aneuploid and euploid clones are color-coded according to the legend. As described in the text, “All” for P5c3 and P6c1 and “None” for P5c3 represent backcrossed segregants that contain all or none of the mutations present in the original evolved clone. “No <i>SUL1</i> amp” for S8c2 is a backcrossed segregant that contains all the mutations identified in S8c2 except for the <i>SUL1</i> amplicon. Raw data can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1002155#pbio.1002155.s010" target="_blank">S2 Table</a>.</p

Aneuploid events variably affect fitness under alternative conditions.

<p>In both A) and B), the name of each aneuploid event or aneuploid clone, respectively, is highlighted with the color corresponding to the condition from which it was originally isolated, while the color of the bar indicates the conditions under which the relative fitness was determined: glucose- (red), sulfate- (blue) and phosphate-limitation (green). A) Aneuploid events identified in evolution experiments and isolated into an otherwise wild-type background show divergent fitness effects under the three conditions examined. The supernumerary chromosomes are labeled according to their identifying translocation (i.e., V_R t X_CEN) and the chromosome XIII disomy is labeled as “Chr XIII.” The missense mutation in <i>PHO84</i> identified in the phosphate-limited evolution experiment P6 only affects fitness under phosphate-limiting conditions and has no effect on fitness under glucose- or sulfate-limiting conditions. B) Evolved aneuploid clones have divergent fitness effects under the three conditions examined here. G, P, and S indicate evolved clones isolated from glucose-, phosphate-, and sulfate-limited evolution experiments, respectively. Raw data can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1002155#pbio.1002155.s010" target="_blank">S2 Table</a>.</p

Experimental design for genome-wide screen for the fitness effects of Tamps.

<p>A) A genome-wide pool of telomeric amplicon strains (Tamps) was constructed. Each Tamp initiates at the <i>KanMX</i> cassette and extends to the proximal telomere, creating a strain that has two chromosomal copies (2N) at most genomic locations, one copy (1N) in the region replaced by the <i>KanMX</i> cassette in the deletion collection, and three copies (3N) at locations telomeric of the deleted gene. The Tamp BC and a portion of KanMX are also present at 2 copies. Each strain contains two barcodes: one identifying the Tamp and a second identifying the unique biological replicate. A third barcode was incorporated during the generation of the barcode sequencing libraries which allowed for multiplexing of experimental samples. Large black arrows represent telomeres; large black circles represent centromeres. The primers represent the barseq primers used to create libraries for sequencing. B) Genome-wide pooled competition experimental design.</p

Tamps on the right arm of chromosome II increase fitness under sulfate limitation.

<p>A) The chrII-targeted pool of 21 Tamps spanning the right arm of chromosome II identified two driver genes that increased fitness under sulfate-limited conditions: <i>BSD2</i> (green rectangle) and <i>SUL1</i> (red rectangle). Both <i>BSD2</i> and <i>SUL1</i> are associated with a Downstep in the fitness landscape. Grey circles represent the individual Tamp fitnesses +/- SE. Each stacked box represents the average fitness of the Tamps enclosed within it; yellow boxes denote positive fitness and teal boxes denote negative fitness. Raw data can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1002155#pbio.1002155.s013" target="_blank">S5 Table</a>. B) Top panel: The 122 Tamps spanning chromosome II in the genome-wide screen are represented as vertical blue lines. Bottom panel: the fitness of each Tamp (+/- SE) is plotted as a blue dot directly below the blue line representing the corresponding Tamp; see red guider arrow for an example. The yellow and blue boxes represent the average relative fitness, positive or negative respectively, of the Tamps enclosed. The extent of each box, i.e. each fitness breakpoint, was defined by DNAcopy segmentation of the fitness landscape. Lower panel: As discussed in the text, an example of a Downstep and an Upstep is highlighted with a purple arrow. Please note that as the data presented in parts A) and B) are from two different competition experiments with different pools, the relative fitnesses would not be expected to be identical. Raw data can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1002155#pbio.1002155.s014" target="_blank">S6 Table</a>.</p

Aneuploid events rise to high population frequencies in evolution experiments, and aneuploid clones are more fit than their euploid, wild-type ancestor.

<p>A) Each row represents an independent evolution experiment and is named according to [<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1002155#pbio.1002155.ref010" target="_blank">10</a>], where “P” indicates a phosphate-limited evolution experiment, “S” indicates a sulfate-limited evolution experiment, and “G” indicates a glucose-limited evolution experiment. The dotted vertical lines distinguish the 16 yeast chromosomes and the black squares mark the position of each centromere. Aneuploid events are defined as red or green boxes where shades of red indicate amplifications and shades of green boxes indicate deletions. The population frequency of each aneuploid event is represented as a color along the red-green color gradient shown to the right. Grey regions indicate euploid regions. Raw data can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1002155#pbio.1002155.s009" target="_blank">S1 Table</a>. B) The relative fitness of evolved clones isolated from evolution experiments carried out under glucose-limited (red), phosphate-limited (green), or sulfate-limited conditions (blue) are shown as the mean +/- standard error (SE). Aneuploid clones isolated from glucose-limited evolution experiments show a significantly greater fitness than euploid clones (Wilcoxon Rank Sum test, <i>p</i>-value = 0.029). Clones isolated from sulfate-limited evolution experiments show significantly greater fitness than the glucose-limited or phosphate-limited evolved clones (Wilcoxon Rank-Sun test, <i>p</i>-value = 0.036). Raw data can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1002155#pbio.1002155.s010" target="_blank">S2 Table</a>.</p