Fob1-dependent condensin recruitment and loop extrusion on yeast chromosome III

Abstract

Despite recent advances in single-molecule and structural analysis of condensin activity in vitro, mechanisms of functional condensin loading and loop extrusion that lead to specific chromosomal organization remain unclear. In Saccharomyces cerevisiae, the most prominent condensin loading site is the rDNA locus on chromosome XII, but its repetitiveness deters rigorous analysis of individual genes. An equally prominent non-rDNA condensin site is located on chromosome III (chrIII). It lies in the promoter of a putative non-coding RNA gene called RDT1, which is in a segment of the recombination enhancer (RE) that dictates MATa-specific chrIII organization. Here, we unexpectedly find that condensin is recruited to the RDT1 promoter in MATa cells through hierarchical interactions with Fob1, Tof2, and cohibin (Lrs4/Csm1), a set of nucleolar factors that also recruit condensin to the rDNA. Fob1 directly binds to this locus in vitro, while its binding in vivo depends on an adjacent Mcm1/α2 binding site that provides MATa cell specificity. We also uncover evidence for condensin-driven loop extrusion anchored by Fob1 and cohibin at RDT1 that unidirectionally extends toward MATa on the right arm of chrIII, supporting donor preference during mating-type switching. S. cerevisiae chrIII therefore provides a new platform for the study of programmed condensin-mediated chromosome conformation.

DNA Methylation-Based High-Resolution Mapping of Long-Distance Chromosomal Interactions in Nucleosome-Depleted Regions

“…In terms of mechanism, published ChIP-seq data shows strong binding of condensin on the right side of RE near the ARS304 and RDT1 gene 40 , and deletion of this region abolishes the mating typedependent Hi-C pattern of ChrIII 36 . Consistent with this observation, we found that the depletion of condensin subunit Smc2 compromises the MTAC signal between MATa and RE.Functionally, condensin depletion impairs mating-type switch efficiency and donor preference40,42 . Together, this data provides strong support for the model that condensin mediates the looping between MATa and RE to promote the usage of HML as the sequence donor.Finally, by comparing the proximity of NDRs to different VPs, a few major spatial patterns start to emerge.…”

“…Previous studies indicate that RE, Sir2, and condensin are important for MATa interactions 40,42 . To test if these elements indeed lead to changes of MTAC signals, we constructed strains containing a RE or SIR2 gene deletion, or auxin-degradable Smc2 (Smc2-AID) ( Materials and Methods ).…”

“…Consistent with this observation, we found that the depletion of condensin subunit Smc2 compromises the MTAC signal between MATa and RE. Functionally, condensin depletion impairs mating-type switch efficiency and donor preference 40,42 . Together, this data provides strong support for the model that condensin mediates the looping between MATa and RE to promote the usage of HML as the sequence donor.…”

“…The interaction patterns are consistent with a mixed population: cells that were originally MATa lose most of their long-distance contacts, while those originally MATα gain interactions with a few telomeres (Fig S5D and E). These findings confirm the interactions with MAT found in Fig 5B and illustrate their dynamic changes during the mating type switching process.Previous studies indicate that RE, Sir2, and condensin are important for MATa interactions40,42 .To test if these elements indeed lead to changes of MTAC signals, we constructed strains containing a RE or SIR2 gene deletion, or auxin-degradable Smc2 (Smc2-AID) (Materials and Methods). Since Smc2-AID shows a partial degradation (reduced to ~50% of wt level) in the presence of ubiquitin ligase OsTIR1 even without auxin (FigS5F), we compared Smc2-AID strains with or without OsTIR1.…”

Molecular basis for condensin enrichment at pericentromeres

“…In fission yeast and Candida albicans , monopolin proteins (Lrs4-Csm1/Mde4-Pcs1) promote the centromeric enrichment of condensin and to prevent merotely 34–36 . Similarly, monopolin (Lrs4-Csm1) is important for condensin localization and function within the rDNA and mating locus in budding yeast 37,38 . In Xenopus egg extracts, the general transcription factor TFIIH promotes the enrichment of condensin on chromatin, though this may be indirect 39 .…”