Genome-Wide Mapping of the Cohesin Complex in the Yeast Saccharomyces cerevisiae

Received December Genome-Wide Mapping of the Cohesin Complex in the Yeast Saccharomyces cerevisiae Earl F. Glynn 0 Paul C. Megee 0 Hong-Guo Yu 0 Cathy Mistrot 0 Elcin Unal 0 Douglas E. Koshland 0 Joseph L. DeRisi 0 Jennifer L. Gerton 0 Bruce Stillman, Cold Spring Harbor Laboratory 0 1 Stowers Institute for Medical Research, Kansas City, Missouri, United States of America, 2 Department of Biochemistry and Molecular Genetics, University of Colorado, Denver, Colorado, United States of America, 3 Howard Hughes Medical Institute, Department of Embryology, Carnegie Institution of Washington , Baltimore , Maryland, United States of America, 4 Department of Biochemistry and Biophysics, University of California , San Francisco, California , United States of America In eukaryotic cells, cohesin holds sister chromatids together until they separate into daughter cells during mitosis. We have used chromatin immunoprecipitation coupled with microarray analysis (ChIP chip) to produce a genome-wide description of cohesin binding to meiotic and mitotic chromosomes of Saccharomyces cerevisiae. A computer program, PeakFinder, enables flexible, automated identification and annotation of cohesin binding peaks in ChIP chip data. Cohesin sites are highly conserved in meiosis and mitosis, suggesting that chromosomes share a common underlying structure during different developmental programs. These sites occur with a semiperiodic spacing of 11 kb that correlates with AT content. The number of sites correlates with chromosome size; however, binding to neighboring sites does not appear to be cooperative. We observed a very strong correlation between cohesin sites and regions between convergent transcription units. The apparent incompatibility between transcription and cohesin binding exists in both meiosis and mitosis. Further experiments reveal that transcript elongation into a cohesin-binding site removes cohesin. A negative correlation between cohesin sites and meiotic recombination sites suggests meiotic exchange is sensitive to the chromosome structure provided by cohesin. The genome-wide view of mitotic and meiotic cohesin binding provides an important framework for the exploration of cohesins and cohesion in other genomes. - Sister chromatid cohesion ensures that daughter cells inherit complete copies of their genome. Cohesion in eukaryotic cells is mediated by a multisubunit protein complex called cohesin. Cohesin consists of four proteins: Smc1, Smc3, Scc1/Mcd1, which is called kleisin and is the target of the protease separase, and Scc3. These proteins have recently been proposed to form a ring structure that encircles sister chromatids (Gruber et al. 2003). Alternately, the ring may act as a snap (Milutinovich and Koshland 2003). Cohesion is established during replication and maintained until metaphase in mitosis (Uhlmann and Nasmyth 1998). All members of the cohesin complex are essential in Saccharomyces cerevisiae, since mutation results in the precocious dissociation of sister chromatids. Cohesion serves at least three roles in the cell with respect to accurate genome transmission. Firstly, cohesion proximal to the centromere facilitates biorientation of chromosomes with respect to the spindle (Tanaka et al. 2000). Secondly, it prevents splitting of chromosomes once bipolar attachments are made (Tanaka et al. 2000). Thirdly, cohesin bound along chromosome arms may be essential for proper chromosome condensation in yeast (Guacci et al. 1997). In meiosis, cohesin at most arm sites disappears prior to the first nuclear division. The meiotic cohesin complex contains Rec8 instead of Scc1/Mcd1 (Klein et al. 1999). Cohesion is maintained distal to crossovers between homologs, which links them and facilitates their biorientation on the meiotic I spindle. Cohesin is also maintained at pericentric regions, which is essential for biorientation of chromosomes at the spindle for the second nuclear division (Buonomo et al. 2000). We are interested in understanding the cis determinants of cohesin binding. Cohesin-associated regions have been identified in yeast using chromatin immunoprecipitation. In these studies cohesin association with chromatin was followed at low resolution along the entire length of Chromosome III (3-kb intervals) or high resolution (300-bp intervals) at limited regions on Chromosome III, V, and XII (Blat and Kleckner 1999; Megee et al. 1999; Tanaka et al. 1999; Laloraya et al. 2000). These studies showed associations of cohesin with specific regions of chromosomes; one of the regions of intense association is centromeres. In addition to the enrichment of cohesin in the pericentric region of Chromosome III, Blat and Kleckner (1999) found a spacing of cohesinassociated regions of 13 kb along the arms of Chromosome III. A similar spacing was observed in a limited region of Chromosome XII (Laloraya et al. 2000). These studies also noted a correlation of cohesins with elevated AT content. The average size of the mapped sites was 0.81 kb (Laloraya et al. 2000). Based on three sites mapped to high resolution, cohesin was proposed to associate with the boundaries of transcriptionally silent regions (Laloraya et al. 2000). Despite these insights into cis determinants of cohesin binding, many aspects of the cohesinDNA interaction remain obscure. The high resolution studies sampled a small portion of the genome, and the low-resolution analysis of Chromosome III does not address questions about the position of cohesin relative to smaller-scale genome features, such as individual transcription units. Furthermore, Chromosome III is the sex chromosome of budding yeast, and, similar to other organisms, it has unusual properties including large domains of repressed recombination, silent mating type loci, and different patterns of replication (Reynolds et al. 1989; Wu et al. 1997). Some discrepancies between high- and low-resolution studies have emerged, including, for example, whether cohesin is found at telomeres (Blat and Kleckner 1999; Laloraya et al. 2000). One approach to better understand cis determinants of cohesin binding is to analyze them across the whole genome. To obtain a genome-wide picture of cohesin binding to S. cerevisiae chromosomes at 12-kb resolution, we used a combination of chromatin immunoprecipitation (ChIP) and microarray methods, often referred to as ChIP chip technology. To aid identification of peaks of cohesin binding we developed a program, PeakFinder, for extraction of peaks from raw ChIP data. We further used this approach to map all the cohesin-binding sites on an ectopic chromosome, a yeast artifical chromosome containing a 334-kb insert from human Chromosome VII. Information from a large number of sites greatly facilitates the assessment of cohesin distribution and of the significance of correlations with many local properties of the genome, such as base composition and coding content. Furthermore, it allows us to evaluate the impact of sever (...truncated)