Principles of Meiotic Chromosome Assembly

Abstract

16During meiotic prophase, chromosomes organise into a series of chromatin loops emanating from 17 a proteinaceous axis, but the mechanisms of assembly remain unclear. Here we elucidate how 18 this elaborate three-dimensional chromosome organisation is underpinned by genomic sequence 19 in Saccharomyces cerevisiae. Entering meiosis, strong cohesin-dependent grid-like Hi-C 20 interaction patterns emerge, reminiscent of mammalian interphase organisation, but with distinct 21 regulation. Meiotic patterns agree with simulations of loop extrusion limited by barriers, yet are 22 patterned by convergent transcription rather than binding of the mammalian interphase factor, 23 CTCF, which is absent in S. cerevisiae-thereby both challenging and extending current 24 paradigms of local chromosome organisation. While grid-like interactions emerge independently 25 of meiotic chromosome synapsis, synapsis itself generates additional compaction that matures 26 differentially according to telomere proximity and chromosome size. Collectively, our results 27 elucidate fundamental principles of chromosome assembly and demonstrate the essential role of 28 cohesin within this evolutionarily conserved process. 29 Schalbetter, Fudenberg et al. 2018 Introduction and Results 30 During meiosis, eukaryotic chromosomes are broken, repaired and paired with their homologs 31 followed by two rounds of segregation-a series of events accompanied by dynamic structural 32 changes of the chromosomes. Most prominent is the paired arrangement of pachytene 33 chromosomes into a dense array of chromatin loops emanating from proteinaceous axes linked 34 by a central core, the synaptonemal complex (SC), which is highly conserved across 35 eukaryotes 1,2 . In S. cerevisiae, structural components include the meiotic cohesin kleisin subunit, 36 Rec8 3 , the transverse filament, Zip1 4 , the axial/lateral elements, Hop1 and Red1 5,6 , and the pro-37 DSB factors Rec114-Mei4-Mer2 (RMM) 7,8 . Much of our understanding of meiotic chromosome 38 structure has been deduced from a combination of electron microscopy, immunofluorescence 39 microscopy, and the genome-wide patterns of protein localisation determined by ChIP. However, 40 the link between key meiotic protein complexes, chromosome conformation, and genomic 41 sequence remains uncharacterized. 42 43 Chromosome conformation capture (3C) techniques generate maps of pairwise contact 44 frequencies that are snapshots of chromosome organisation. 3C methods were originally applied 45 to assay chromosome conformation in S. cerevisiae, including during meiosis 9 . Now they are 46 widely used across a range of organisms and cellular contexts to link 3D organisation directly with 47 53 54Starting with a synchronized G1 population we analysed timepoints encompassing DNA 55 replication, meiotic prophase and both meiotic divisions ( Fig. 1a,b,c, Extended Data Fig. 1a,b,c). 56 In G1, we detect strong centromere clustering ( Fig. 1a,d) and folding back of the arms at the 57 centromeres ( Fig. 1a, Supplementary Fig...

Sub-nucleosomal Genome Structure Reveals Distinct Nucleosome Folding Motifs

“…In contrast to infrequent chromatin looping at M phase ( Figure 3A), previous Hi-C experiments have observed stable chromatin looping during meiotic division (Muller et al, 2018;Schalbetter et al, 2018). This difference in looping frequency may be explained as resulting from specific mechanisms in either of mitosis or meiosis, such as Rec8 mediated sister chromatid cohesion (Watanabe, 2005).…”

Separable roles of the DNA damage response kinase Mec1(ATR) and its activator Rad24(RAD17) within the regulation of meiotic recombination

“…No further increase in recombination was observed in the ndt80AR msh2 Δ double strain (Fig 2A), suggesting that there may be an upper limit to recombination frequency in these backgrounds. Interestingly, extending prophase length, but not MSH2 deletion, skews recombination towards subtelomeres (Fig S3A-B)—regions that are both less compacted as measured by Hi-C [43], and retain disproportionately high levels of DSB formation at pachytene [44].…”

Convergent genes shape budding yeast pericentromeres

“…Hi-C protocol was modified from 20 , 34 , 35 . Cells were cultured, fixed and lysed as described in 34 .…”

“…Lysates were prepared by grinding the frozen pellet in a chilled mortar with a pestle for 15 minutes and 1/10 th of the initial pellet weight (~0.5 g) was taken for further processing. Restriction enzyme digestion ( Dpn II), filling-in, ligation, crosslink reversal, DNA concentration and purification and biotin removal were carried out as described in 35 . DNA was then fragmented on a Bioruptor Plus sonication device (Diagenode) for a total of 2x 30 cycles 30 seconds on/off at High setting.…”