wtf genes are prolific dual poison-antidote meiotic drivers
- Stowers Institute for Medical Research, United States
- Fred Hutchinson Cancer Research Center, United States
- University of Kansas Medical Center, United States
- Howard Hughes Medical Institute, Fred Hutchinson Cancer Research Center, United States
Figures
Figure 1 with 1 supplement
A complex meiotic drive landscape on Sk and Sp chromosome 3 is revealed by recombination mapping.
(A) A cross between Sk and Sp generates a heterozygote that has low fertility and preferentially transmits Sk alleles on all three chromosomes into viable gametes (Zanders et al., 2014). (B) …
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Figure 1—source data 1
Breakpoints between Sp and Sk-derived DNA sequences.
The introgression strains used in diploids 1–8 were sequenced and genotyped for single-nucleotide polymorphisms (SNPs) that reliably distinguish Sk and Sp as in (Zanders et al., 2014). The SNPs flanking the recombination event (left and right boundaries) that generated each breakpoint between Sp and Sk DNA for each introgression strain are shown. The coordinates refer to the position of the SNP on Sp chromosome 3. For the introgression used in diploid 9, SNPs were genotyped via PCR and Sanger sequencing.
- https://doi.org/10.7554/eLife.26033.004
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Figure 1—source data 2
Raw data underlying Figure 1C.
Diploids 1–10 (column 1) were generated by crossing the indicated haploid strains (columns 2 and 4). The diploid numbers correspond to those in Figure 1 and the text. All strains are rec12∆ and transmission of chromosome 3 was followed using heterozygous markers at the ade6 locus (columns 3 and 5). hphMX4 confers resistance to hygromycin (HygR). The number of viable progeny inheriting one or both ade6 markers is indicated (columns 6–8), as are the percentage of the progeny that inherited both markers (column 10). These strains have two copies of chromosome 3, so we refer to them as disomes, although other homozygous disomes could be present in the Ade+ HygS and Ade- HygR classes as well. Amongst the progeny that inherit only one ade6 marker, we show the percent that inherit allele 2 (column 12), which is the allele from the pure Sk chromosome. For the statistical analyses (G-tests), we compared the observed heterozygous disomy and allele 2 transmission to the values observed in diploid 10, which is a pure Sk rec12∆ control (columns 11 and 13). The last column indicates the number of independent diploids that were generated and assayed of each genotype.
- https://doi.org/10.7554/eLife.26033.005
Figure 1—figure supplement 1
Generation of mosaic chromosome 3 used in Figure 1B.
The goal of these crosses was to generate a strain containing mostly Sp-derived DNA on chromosome 3 in an otherwise Sk background. This effort was complicated by the different karyotypes of Sp and Sk…
Figure 2
Sk wtf4 is a self-sufficient meiotic driver that kills gametes that do not inherit the gene.
(A) Allele transmission and propidium iodide (PI) staining phenotypes of diploids 11–19. Sk-derived DNA is purple, Sp-derived DNA is green. The cartoons depict chromosome 3. Chromosomes 1 and 2 are …
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Figure 2—source data 1
PI staining correlates with viable spore yield as a measure of fertility in wild-type and wtf heterozygous crosses.
The fertility of the indicated diploids was assayed using both the established viable spore yield assay and by PI staining. We avoided tetrad dissection because we found that it was complicated by disintegration of spores destroyed by drive. The viable spore yield assay is a plating assay that measures the number of viable spores produced per viable diploid induced to undergo meiosis (Smith, 2009). PI is excluded from viable spores, but enters those destroyed by wtf drivers (Figure 2B). Although PI staining likely will not detect spore death by other causes that do not disrupt membrane integrity, the percent of PI-excluding cells correlates with viable spore yield in wild-type and wtf heterozygous crosses. Diploids of four genotypes are shown. Three of the diploids are used elsewhere in the paper (diploids 17, 22, and 27) and the diploid number (column 1) corresponds to the diploid numbers used in the main text and figures. The two strains that were mated to generate the diploids are shown in columns 2 and 4. The drive-relevant genotypes of these parental strains is shown in columns 3 and 5. The last two columns show the PI-staining and viable spore yield phenotypes of each diploid.
- https://doi.org/10.7554/eLife.26033.008
Figure 3 with 1 supplement
Sk wtf4 has the capacity to make two proteins and Wtf4-GFP shows a dual localization pattern.
(A) Model for meiotic drive of Sk wtf4 via a poison-antidote mechanism. (B) wtf4 creates a long and an alternative short transcript. See Figure 3—figure supplement 1 for a depiction of the long-read …
Figure 3—figure supplement 1
Sp wtf4 has alternate transcriptional start sites.
Our annotation of the wtf4 gene with alternate start sites predicted is shown at the top in the same format as Figures 3–5. The PomBase annotation for Sp wtf4 is shown below that in blue. The …
Figure 4
Sk wtf4 creates two proteins using alternate transcripts: an antidote and a gamete-killing poison.
(A) Separation of function wtf4 alleles. The red stars indicate start codon mutations. (B) Allele transmission and PI staining phenotypes of Sp diploids with the indicated Sk wtf4 alleles integrated …
Figure 5 with 1 supplement
Wtf4 antidote is spore-specific and Wtf4 poison spreads throughout the ascus.
(A) Constructs tagging either the Wtf4 antidote (top) or poison (bottom) proteins. The red stars indicate start codon mutations. (B) Allele transmission and PI staining phenotypes for tagged …
Figure 5—figure supplement 1
Spectral unmixing verifies true signal.
Wtf4 poison (cyan) and antidote (magenta) protein localization in a mature ascus processed using linear unmixing [top] and unprocessed [bottom]. Scale bar represents three microns.
Additional files
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Supplementary file 1
Raw data of allele transmission from Figures 2–5.
Each horizontal entry represents the relevant genotype and allele transmission of the indicated diploids. The first column represents the diploid number, which matches the numbers shown in Figures 2–5. In columns 2–5, the two haploid parent strains (SZY#s) are indicated as are the alleles contributed by those parent strains at the experimental locus monitored for drive. Alleles derived from Sp are green whereas those from Sk are purple. For diploids 11–15, transmission of the wtf4 locus was followed using alleles of ura4, which is fortuitously closely linked to wtf4. Columns 9 and 10 indicate which phenotypes were followed at the drive loci and the number of progeny that displayed each phenotype. Some progeny inherited both alleles at a given drive locus and when the markers were codominant we could detect those disomes. The number of those disomes, which are likely heterozygous diploids or aneuploids, are shown in column 11 and their overall frequency is shown in column 12. If we did not have codominant markers, columns 11 and 12 are filled with ---. Column 13 is a measure of meiotic drive. It shows the fraction of the non-disomic progeny that inherited allele 1 (column 3). Column 14 shows the total number of progeny assayed for each diploid and column 15 is the p value calculated from a G-test comparing the allele transmission of allele 1 to a control. Diploid 13 served as the control for diploids 11, 12, 14 and 15. Diploid 16 served as the control for the rest of the diploids. Columns 6–8 are internal controls for each diploid. These controls represent an additional heterozygous locus unlinked to the meiotic drive locus that should undergo Mendelian allele transmission. The lys locus is lys4, the ade locus is ade6, and the ura locus is ura4. The final column indicates the number of independently generated diploids that were tested for each genotype.
- https://doi.org/10.7554/eLife.26033.014
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Supplementary file 2
Raw data for PI-staining phenotypes from Figures 2–5.
Each horizontal entry represents the relevant genotype and allele transmission of the indicated diploids. The first column represents the diploid number, which matches the numbers shown in Figures 2–5. In columns 2–5, the two haploid parent strains (SZY#s) are indicated as are the alleles contributed by those parent strains at the experimental locus monitored for drive. Alleles derived from Sp are green whereas those from Sk are purple. Columns 6 and 7 indicate the number of spores that stained with PI (dead spores) and those that did not (likely living spores) and column 8 shows the percentage of spores that did not stain with PI. Column 9 shows the number of stained asci that were scored for each diploid type. Spores not contained within 4-spore asci were not scored. Column 10 shows the p value from a G test comparing the number of stained and unstained spores for each diploid to a control diploid. Diploid 13 served as a control for diploids 11, 12, 14 and 15. Diploid 16 served as a control for all other diploids. The number of independently generated diploid strains that were tested is indicated in the last column.
- https://doi.org/10.7554/eLife.26033.015
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Supplementary file 3
Yeast strains.
- https://doi.org/10.7554/eLife.26033.016
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Supplementary file 4
Plasmids.
- https://doi.org/10.7554/eLife.26033.017
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Supplementary file 5
Oligos.
- https://doi.org/10.7554/eLife.26033.018
