Project description:Chromosome segregation is a fundamental process in all life forms and requires coordination with genome organization, replication and cell division. The mechanism that mediates chromosome segregation in archaea remains enigmatic, despite the centre-stage role assumed by these organisms in the discourse about the origin of eukaryotes. Previously, we identified two proteins, SegA and SegB, which form a minimalist chromosome partition machine in Sulfolobales. Here we uncover patterns and mechanisms that the SegAB system employs to link chromosome organization to genome segregation. Deletion of the genes causes growth and chromosome partition defects. ChIP-seq investigations revealed that SegB binds to multiple sites scattered across the chromosome, but mainly localised close to the segAB locus in most of the examined archaeal genera. The sites are predominantly present in intragenic regions. Recent chromosome conformation studies have shown that the chromosome of Sulfolobales members is organised into two spatially segregated compartments, designated A and B. Interestingly, SegB sites are predominantly located in the A compartment, whose shaping factors have remained elusive so far. We show that SegB coalesces into multiple foci through the nucleoid, exhibiting a biased localisation towards the cell periphery, which hints at potential tethers to the cell membrane. This clue is further strengthened by the structural similarities of SegB orthologues with predicted b-helix domains to membrane-associate proteins, including the distant bactofilin. Atomic force microscopy experiments provided mechanistic insights into how SegB binds DNA and uncovered short-range DNA compaction and long-range looping of distant sites by SegB. These activities point to a significant role for SegB in chromosome condensation that in turn enables genome segregation. Collectively, our data put forward SegAB as important players in bridging chromosome organization to genome segregation in archaea.
Project description:Chromosome segregation is a vital process for all organisms. The mechanisms underpinning chromosomal partitioning in the archaeal domain remain elusive. Our group has identified the first chromosome segregation system in thermophilic archaea. Sulfolobus solfataricus partition system consists of SegA, an orthologue of bacterial Walker-type ParA proteins; SegB, an archaea-specific DNA binding protein and a cis-acting DNA region. ChIP-seq experiments disclosed multiple SegB binding sites scattered over the chromosome and revealed a novel DNA binding motif.
Project description:Genome/chromosome organization is highly ordered and controls nuclear events. Here, we show that the TATA box-binding protein (TBP) interacts with the Cnd2 kleisin subunit of condensin to mediate interphase and mitotic chromosome organization in fission yeast. TBP recruits condensin onto RNA polymerase III-transcribed (Pol III) genes and highly transcribed Pol II genes; condensin in turn associates these genes with centromeres. Inhibition of the Cnd2-TBP interaction disrupts condensin localization across the genome and the proper assembly of mitotic chromosomes, leading to severe defects in chromosome segregation and eventually causing cellular lethality. We propose that the Cnd2-TBP interaction coordinates transcription with chromosomal architecture by linking dispersed gene loci with centromeres. This chromosome arrangement can contribute to the efficient transmission of physical force at the kinetochore to chromosomal arms, thereby supporting the fidelity of chromosome segregation. Genome-wide distributions of condensin and Pol III factors in fission yeast.
Project description:Research on chromosome organization and cell cycle progression in spherical bacteria, particularly Staphylococcus aureus, remains limited and fragmented. In this study, we established a working model to investigate chromosome dynamics in S. aureus using a Fluorescent Reporter Operator System (FROS), which enabled precise localization of specific chromosomal loci. This approach revealed that the S. aureus cell cycle and chromosome replication cycle are not coupled, with cells exhibiting two segregated origins of replication at the start of the cell cycle. The chromosome has a specific origin-terminus-origin conformation, with origins localizing near the membrane, towards the tip of each hemisphere, or the “cell poles”. We further used this system to assess the role of various proteins with a role in S. aureus chromosome biology, focusing on the ParB-parS and SMC-ScpAB systems. Our results demonstrate that ParB binds five parS chromosomal sequences and the resulting complexes influence chromosome conformation, but play a minor role in chromosome compaction and segregation. In contrast, the SMC-ScpAB complex plays a key role in S. aureus chromosome biology, contributing to chromosome compaction, segregation and spatial organization. Additionally, we systematically assessed the impact of proteins linking chromosome segregation to cell division—Noc, FtsK, SpoIIIE, and XerC—on origin and terminus number and positioning. While these proteins have been studied previously, our work uniquely standardizes comparisons by using the same strain background, growth conditions, and analytical approaches for all mutants. This work provides a comprehensive study of the factors governing chromosome dynamics and organization in S. aureus, contributing to our knowledge on chromosome biology of spherical bacteria.
Project description:Research on chromosome organization and cell cycle progression in spherical bacteria, particularly Staphylococcus aureus, remains limited and fragmented. In this study, we established a working model to investigate chromosome dynamics in S. aureus using a Fluorescent Reporter Operator System (FROS), which enabled precise localization of specific chromosomal loci. This approach revealed that the S. aureus cell cycle and chromosome replication cycle are not coupled, with cells exhibiting two segregated origins of replication at the start of the cell cycle. The chromosome has a specific origin-terminus-origin conformation, with origins localizing near the membrane, towards the tip of each hemisphere, or the “cell poles”. We further used this system to assess the role of various proteins with a role in S. aureus chromosome biology, focusing on the ParB-parS and SMC-ScpAB systems. Our results demonstrate that ParB binds five parS chromosomal sequences and the resulting complexes influence chromosome conformation, but play a minor role in chromosome compaction and segregation. In contrast, the SMC-ScpAB complex plays a key role in S. aureus chromosome biology, contributing to chromosome compaction, segregation and spatial organization. Additionally, we systematically assessed the impact of proteins linking chromosome segregation to cell division—Noc, FtsK, SpoIIIE, and XerC—on origin and terminus number and positioning. While these proteins have been studied previously, our work uniquely standardizes comparisons by using the same strain background, growth conditions, and analytical approaches for all mutants. This work provides a comprehensive study of the factors governing chromosome dynamics and organization in S. aureus, contributing to our knowledge on chromosome biology of spherical bacteria.
Project description:Genome/chromosome organization is highly ordered and controls nuclear events. Here, we show that the TATA box-binding protein (TBP) interacts with the Cnd2 kleisin subunit of condensin to mediate interphase and mitotic chromosome organization in fission yeast. TBP recruits condensin onto RNA polymerase III-transcribed (Pol III) genes and highly transcribed Pol II genes; condensin in turn associates these genes with centromeres. Inhibition of the Cnd2-TBP interaction disrupts condensin localization across the genome and the proper assembly of mitotic chromosomes, leading to severe defects in chromosome segregation and eventually causing cellular lethality. We propose that the Cnd2-TBP interaction coordinates transcription with chromosomal architecture by linking dispersed gene loci with centromeres. This chromosome arrangement can contribute to the efficient transmission of physical force at the kinetochore to chromosomal arms, thereby supporting the fidelity of chromosome segregation.
Project description:Chromosome segregation has been assumed to be a cell-autonomous process. We tested this assumption by comparing chromosome segregation fidelity in epithelial cells in various contexts and discovered that these cells have increased chromosome missegregation outside of their native tissue. Using organoid culture systems, we show that tissue architecture, specifically integrin function, is required for accurate chromosome segregation in epithelia. We find that tissue architecture enhances the correction of merotelic microtubule-kinetochore attachments, and this is critically important for maintaining chromosome stability in the polyploid liver. Our data lead to the surprising conclusion that chromosome segregation in epithelia is a cell non-autonomous process. We propose that disruption of tissue architecture could underlie the chromosome instability that characterizes and drives epithelial cancers. Moreover, our findings highlight the importance of context for fundamental cellular processes and caution against the exclusive reliance on cell culture systems for deciphering and manipulating mammalian biology.