Local chromosome context is a major determinant of crossover pathway biochemistry during budding yeast meiosis
- National Cancer Institute, United States
- The University of Sheffield, United Kingdom
Figures
Figure 1 with 1 supplement
Inserts used to monitor VDE-initiated meiotic recombination.
The HIS4 and URA3 loci are denoted throughout this paper in red and blue, respectively, and are in Red1/Hop1 enriched and depleted regions, respectively (see Figure 4A and Figure 4—figure supplement …
Figure 1—figure supplement 1
Spo11-initiated events at the two insert loci.
(A) Spo11-catalyzed DSBs are more frequent at HIS4 that at URA3. Left—Southern blots of EcoRI digests of DNA from vde∆ strains, probed with pBR322 sequences, showing Spo11-DSBs in the Parent 2 …
Figure 2 with 1 supplement
VDE-initiated recombination occurs at similar levels at the two insert loci.
(A) Cumulative DSB levels are similar at the two insert loci. The fraction of uncut VRS-containing chromosomes (Parent 1) was determined by subtracting the amount of the NCO band in HindIII + VDE …
Figure 2—figure supplement 1
70–80% of VDE-DSBs are repaired.
(A) Fraction of inserts remaining, calculated using HindIII digests (see Figure 1). For the arg4-VRS103 insert, the ratio (Parent 2 + CO2)/ (0.5 x LC) was calculated at 9 hr, and was then normalized …
Figure 3 with 2 supplements
Different resolvase-dependence of crossover formation at the two insert loci.
(A) Crossover frequencies (average of CO1 and CO2) measured as in Figure 2C from HIS4 insert-containing mutants lacking MutLγ (mlh3), structure-selective nucleases (mms4-md yen1 slx1) or both …
Figure 3—figure supplement 1
VDE-DSB and NCO frequencies in resolvase mutants.
(A) VDE-DSB frequencies (top), measured as in Figure 2B, and NCO frequencies (bottom), measured as in Figure 2C, from HIS4 insert-containing strains. (B) As panel A, with strains containing inserts …
Figure 3—figure supplement 2
Southern blots of HindIII and HindIII-VDE digests of DNA from HIS4 insert-containing strains (top) and from URA3 insert-contaning strains (bottom).
Probes and gel labels are as in Figure 1; JM—joint molecule recombination intermediates.
Figure 4 with 2 supplements
pch2∆ mutants display altered Hop1 occupancy and crossover MutLγ-dependence.
(A) Hop1 occupancy at insert loci, determined by chromatin immunoprecipitation and quantitative PCR. Top—cartoon of insert loci, showing the location of primer pairs used. Bottom—relative Hop1 …
Figure 4—figure supplement 1
Hop1 occupancy at non-insert loci, DSBs and NCOs in pch2∆ mutants.
(A) Hop1 occupancy at corresponding loci lacking inserts, determined as in Figure 4A. Occupancy at HIS4 is from strains with inserts at URA3, and vice versa. (B) DSBs and NCOs in inserts at HIS4, …
Figure 4—figure supplement 2
Southern blots of HindIII and HindIII-VDE digests of DNA from HIS4 insert-containing strains (top) and from URA3 insert-contaning strains (bottom).
Gel labels are as in Figure 1; JM—joint molecule recombination intermediates. In the gel with HinDIII digests of samples from a pch2∆ mm4-mn yen1∆ slx1∆ strain with inserts at URA3, the 9 hr sample …
Figure 5 with 2 supplements
VDE-initiated COs are reduced and are MutLγ-independent in the absence of Spo11 activity.
(A) VDE-initiated crossover frequencies, measured as in Figure 2C in spo11-Y135F strains (dark solid lines) in inserts at HIS4 (red) and at URA3 (blue). Data from the corresponding SPO11 strains …
Figure 5—figure supplement 1
DSBs and recombinant products in spo11 strains.
(A) Cumulative DSB levels, expressed as loss of the VRS-containing insert, calculated as in Figure 2A. (B) Relative recovery of recombination products, calculated as in Figure 2—figure supplement 1B.…
Figure 5—figure supplement 2
Southern blots of HindIII and HindIII-VDE digests of DNA from spo11 strains with inserts at HIS4 (top) and at URA3 (bottom).
Gel labels are as in Figure 1; JM—joint molecule recombination intermediates.
Figure 6
Different resolvase functions in different genome domains.
(A) Early crossover decision model for meiotic recombination (Bishop and Zickler, 2004; Hollingsworth and Brill, 2004) illustrating early noncrossover formation, a major pathway where recombination …
Additional files
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Supplementary file 1
Yeast strains.
- https://doi.org/10.7554/eLife.19669.017
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Supplementary file 2
Primary data for graphs in all figures.
- https://doi.org/10.7554/eLife.19669.018
