CLMS analysis of fission yeast condensin-DNA samples
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ABSTRACT: Crosslinking-MS analysis of sulfo-SDA crosslinked fission yeast condensin-DNA samples in the initial binding state (absence of nucleotide) and in the DNA gripping state (in the presence of ADP•BeF3)
Project description:Crosslinked DNA from wild type cells or from mutant of the THO complex was analyzed by tiling arrays to precisely detect DNA targets of THO in yeast.
Project description:In this study, we employed the photoactivatable crosslinker (sulfo-SDA) to investigate protein-protein interaction between human Separase-Scc1 fusion protein and SMC1/SMC3 cohesin subunits complex in the presence of DNA.
Project description:This data is from sulfo-SDA crosslinked condensin pentamer. Two datsets, one without atp aand one with ATP. How protein complexes of the SMC family fold DNA into the large loops that are fundamental for the 3D organization of genomes is a central unresolved question of chromosome biology. We used electron cryomicroscopy to investigate the reaction cycle of the SMC complex condensin, which is a key determinant of chromosome morphology and behavior during mitosis. Our structures of the Saccharomyces cerevisiae condensin holo complex at different functional stages suggest that ATP binding induces the transition from a folded-rod SMC conformation into an open architecture and triggers the exchange of the two HEAT-repeat subunits at the SMC ATPase head domains. We propose that these steps result in the interconversion of DNA binding sites in the catalytic core that form the basis of the DNA translocation and loop-extrusion activities of condensin.
Project description:Here, we describe a genome-wide map of uracil lesions arising form deaminated UV-induced cyclobutane pyrimidine dimers (CPDs) in yeast (Saccharomyces cerevisiae) genomic DNA that were UV-irradiated and deaminated as naked DNA in vitro. Deaminated CPDs (dCPDs) were mapped at single nucleotide resolution across the yeast genome using the new dCPD-seq method. We used these data to analyze CPD deamination in different trinucleotide sequence contexts.
Project description:Crosslinking mass spectrometry dataset of the Mpe1 and Cft2 bound to the yeast CPF polymerase module. Sulfo-SDA is the crosslinker used.
Project description:Crosslinked DNA from wild type cells or from mutant of the THO complex was analyzed by tiling arrays to precisely detect DNA targets of THO in yeast. DNA coming from SDS-washed DCF pellet and the supernatant SN2k of 2 strains were compared in the study, WT (W303) and mft1 delta (DLY224). The DNA of each condition (Pellet or Supernatant) was labeled either with Cy3 or Cy5 fluorescent dyes and competitively hybridized on 244k agilent arrays. For each strain, the experiment was performed on 2 biological replicates, with dye swap.
Project description:We use metabolically synchronous, continuously-grown yeast cultures to measure DNA occupancy and track the global patterns with respect to the metabolic state of the culture, showing a genome-wide nucleosome focusing in the reductive phase, followed by clearance of the promoter regions
Project description:DNA topoisomerases are required to resolve DNA topological stress. Despite this essential role, abortive topoisomerase activity generates aberrant protein-linked DNA breaks, jeopardising genome stability. Here, to understand the genomic distribution and mechanisms underpinning topoisomerase-induced DNA breaks, we map Top2 DNA cleavage with strand-specific nucleotide resolution across the S. cerevisiae and human genomes - and use the meiotic Spo11 protein to validate the broad applicability of this method to explore the role of diverse topoisomerase family members. Our data characterises Mre11-dependent repair in yeast, and defines two strikingly different fractions of Top2 activity in humans: tightly localised CTCF-proximal, and broadly distributed transcription-proximal, the latter correlated with gene length and expression. Moreover, single nucleotide accuracy enables us to reveal the influence primary DNA sequence has upon Top2 cleavage - distinguishing canonical DNA double-strand breaks (DSBs) from a major population of DNA single-strand breaks (SSBs) induced by etoposide (VP16) in vivo.