Mechanism of in vivo activation of the MutLgamma-Exo1 complex for meiotic crossover formation

Description: Summary from the GEO: "Programmed DNA double-strand breaks (DSBs) initiate meiotic recombination and their subsequent repair culminates in crossover (CO) formation. COs result from the asymmetric cleavage of double-Holliday junction (dHJ) intermediates, that requires the MutLγ complex together with a non-catalytic function of Exo1, an activity essential for fertility but at risk of generating unwanted chromosome rearrangements. Here we show how crossover formation by MutLγ is activated at the right time and at the right place. MutLγ forms a constitutive complex with Exo1, and in meiotic cells transiently contacts the upstream MutSγ (Msh4-Msh5) heterodimer. MutLγ associates with DSB hotspots at a late step in the recombinational repair, once recombination intermediates have been stabilized and engaged in the crossover repair pathway. MutLγ-Exo1 is recruited to DSB hotspots independently of the polo-like Cdc5 kinase, but to activate dHJ resolution, Cdc5 is recruited to the recombination intermediates and interacts individually with both MutLγ and Exo1, suggesting their direct modification. in vivo, MutLγ occupancy is restrained on recombination intermediates, and genome-wide, MutLγ associates with the vast majority of DSB hotspots, but at a lower frequency in centromeres, consistent with a strategy to reduce at-risk crossover events in these regions, and in late replicating regions. Our data highlight the highly temporally and spatially control of the activity of this constitutive, potentially harmful, nuclease"

Overall design from the GEO: "9 samples total: 2 independent replicates of Mer3-Flag ChIP at 4 h in meiosis and their control untagged anti-Flag ChIP at 4 h in meiosis; 2 Mlh3-Myc8 ChIP at 5 h and 5h30 in pCUP1-IME1-synchronized meiosis and their corresponding controls spo11∆ Mlh3-Myc8 ChIP at 5 h and 5h30 in pCUP1-IME1-synchronized meiosis; 1 exo1∆ Mlh3-Myc8 ChIP at 5h30 in pCUP1-IME1 synchronized meiosis and its corresponding control spo11∆ Mlh3-Myc8 ChIP at 5h30 in pCUP1-IME1 synchronized meiosis."

Subject of Study

Saccharomyces cerevisiae

Subject(s)

DNA Breaks, Double-Stranded

Meiosis

MutL Proteins

Protein Binding

Recombination, Genetic

Access via GEO
TAR of BigWig files of genome binding/occupancy profiling by high throughput sequencing
Accession #: GSE132850

Access via SRA
ChIP sequencing of 9 samples
Accession #: SRP201616

Access via BioProject
Additional information about overall initiative.
Accession #: PRJNA549269
Access Restrictions: Free to All
Access Instructions: The NCBI Gene Expression Omnibus, SRA, and BioProject databases provide open access to these files.
Associated Publications: Sanchez A, Adam C, Rauh F, et al. Exo1 recruits Cdc5 polo kinase to MutLγ to ensure efficient meiotic crossover formation. Proceedings of the National Academy of Sciences of the United States of America. 2020; 117(48):30577-30588.
Data Type: Genomic
Equipment Used: Illumina HiSeq 2500
High throughput sequencing system
Software Used: SRA Toolkit
The NCBI SRA Toolkit enables reading ("dumping") of sequencing files from the SRA database and writing ("loading") files into the .sra format.
Dataset Format(s): Plain Text, SRA, TAR, BIGWIG
Dataset Size: 1.0 GB (TAR of BW), 4.8 Kb (TXT), 8.5 GB (SRA)

Data Catalog Record Updated: 2023-12-07