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We study how chromosome architecture contributes to faithful genome segregation. Genome stability relies on the fact that at each round of cell division, the genetic information encoded in the DNA molecules is properly segregated into the two daughter cells. Proper completion of this process, in turn, depends on two major changes in chromosome organization: 1) The two-sister DNA molecules remain tightly associated with each other from the moment of DNA replication until the later stages of the subsequent mitosis; 2) At the onset of nuclear division, chromatin is converted into compact structures with the right mechanical properties (size, flexibility, and rigidity) to facilitate their segregation; Our laboratory adopts a multidisciplinary approach, combining Drosophila genetics, acute protein inactivation, 4D-live cell imaging and biophysical/mathematical modelling to evaluate how dynamic mitotic chromosomes are assembled and how their morphology influences the mechanical aspects of chromosome movement and cell cycle checkpoint signalling. In parallel we aim to dissect how different cells respond to compromised chromosome cohesion and condensation, both at the cellular and organism level. By studying the contribution of chromosome structure in the mechanics of nuclear division we aim to identify novel routes to aneuploidy that underlie several human conditions, including developmental diseases, cancer and infertility. Informal enquiries on research or available positions in our group are welcome and should be sent to Raquel Oliveira. We welcome people from a range of backgrounds, including biology, biochemistry, chemistry, physics, biological engineering and bioinformatics.

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Now showing 1 - 8 of 8
  • Metaphase chromosome structure is dynamically maintained by condensin I-directed DNA (de)catenation
    Publication . Piskadlo, Ewa; Tavares, Alexandra; Oliveira, Raquel A
    Mitotic chromosome assembly remains a big mystery in biology. Condensin complexes are pivotal for chromosome architecture yet how they shape mitotic chromatin remains unknown. Using acute inactivation approaches and live-cell imaging in Drosophila embryos, we dissect the role of condensin I in the maintenance of mitotic chromosome structure with unprecedented temporal resolution. Removal of condensin I from pre-established chromosomes results in rapid disassembly of centromeric regions while most chromatin mass undergoes hyper-compaction. This is accompanied by drastic changes in the degree of sister chromatid intertwines. While wild-type metaphase chromosomes display residual levels of catenations, upon timely removal of condensin I, chromosomes present high levels of de novo Topoisomerase II (TopoII)-dependent re-entanglements, and complete failure in chromosome segregation. TopoII is thus capable of re-intertwining previously separated DNA molecules and condensin I continuously required to counteract this erroneous activity. We propose that maintenance of chromosome resolution is a highly dynamic bidirectional process.
  • Novel insights into mitotic chromosome condensation
    Publication . Piskadlo, Ewa; Oliveira, Raquel A.
    The fidelity of mitosis is essential for life, and successful completion of this process relies on drastic changes in chromosome organization at the onset of nuclear division. The mechanisms that govern chromosome compaction at every cell division cycle are still far from full comprehension, yet recent studies provide novel insights into this problem, challenging classical views on mitotic chromosome assembly. Here, we briefly introduce various models for chromosome assembly and known factors involved in the condensation process (e.g. condensin complexes and topoisomerase II). We will then focus on a few selected studies that have recently brought novel insights into the mysterious way chromosomes are condensed during nuclear division.
  • Dynamical Scenarios for Chromosome Bi-orientation
    Publication . Zhang, Tongli; Oliveira, Raquel A.; Schmierer, Bernhard; Novák, Béla
    Chromosome bi-orientation at the metaphase spindle is essential for precise segregation of the genetic material. The process is error-prone, and error-correction mechanisms exist to switch misaligned chromosomes to the correct, bi-oriented configuration. Here, we analyze several possible dynamical scenarios to explore how cells might achieve correct bi-orientation in an efficient and robust manner. We first illustrate that tension-mediated feedback between the sister kinetochores can give rise to a bistable switch, which allows robust distinction between a loose attachment with low tension and a strong attachment with high tension. However, this mechanism has difficulties in explaining how bi-orientation is initiated starting from unattached kinetochores. We propose four possible mechanisms to overcome this problem (exploiting molecular noise; allowing an efficient attachment of kinetochores already in the absence of tension; a trial-and-error oscillation; and a stochastic bistable switch), and assess their impact on the bi-orientation process. Based on our results and supported by experimental data, we put forward a trial-and-error oscillation and a stochastic bistable switch as two elegant mechanisms with the potential to promote bi-orientation both efficiently and robustly.
  • Disengaging the Smc3/kleisin interface releases cohesin from Drosophila chromosomes during interphase and mitosis
    Publication . Eichinger, Christian S; Kurze, Alexander; Oliveira, Raquel A; Nasmyth, Kim
    Cohesin's Smc1, Smc3, and kleisin subunits create a tripartite ring within which sister DNAs are entrapped. Evidence suggests that DNA enters through a gate created by transient dissociation of the Smc1/3 interface. Release at the onset of anaphase is triggered by proteolytic cleavage of kleisin. Less well understood is the mechanism of release at other stages of the cell cycle, in particular during prophase when most cohesin dissociates from chromosome arms in a process dependent on the regulatory subunit Wapl. We show here that Wapl-dependent release from salivary gland polytene chromosomes during interphase and from neuroblast chromosome arms during prophase is blocked by translational fusion of Smc3's C-terminus to kleisin's N-terminus. Our findings imply that proteolysis-independent release of cohesin from chromatin is mediated by Wapl-dependent escape of DNAs through a gate created by transient dissociation of the Smc3/kleisin interface. Thus, cohesin's DNA entry and exit gates are distinct.
  • Manipulation of the Quorum Sensing Signal AI-2 Affects the Antibiotic-Treated Gut Microbiota
    Publication . Thompson, Jessica Ann; Oliveira, Rita Almeida; Djukovic, Ana; Ubeda, Carles; Xavier, Karina Bivar
    The mammalian gut microbiota harbors a diverse ecosystem where hundreds of bacterial species interact with each other and their host. Given that bacteria use signals to communicate and regulate group behaviors (quorum sensing), we asked whether such communication between different commensal species can influence the interactions occurring in this environment. We engineered the enteric bacterium, Escherichia coli, to manipulate the levels of the interspecies quorum sensing signal, autoinducer-2 (AI-2), in the mouse intestine and investigated the effect upon antibiotic-induced gut microbiota dysbiosis. E. coli that increased intestinal AI-2 levels altered the composition of the antibiotic-treated gut microbiota, favoring the expansion of the Firmicutes phylum. This significantly increased the Firmicutes/Bacteroidetes ratio, to oppose the strong effect of the antibiotic, which had almost cleared the Firmicutes. This demonstrates that AI-2 levels influence the abundance of the major phyla of the gut microbiota, the balance of which is known to influence human health.
  • Premature Sister Chromatid Separation Is Poorly Detected by the Spindle Assembly Checkpoint as a Result of System-Level Feedback
    Publication . Mirkovic, Mihailo; Hutter, Lukas H.; Novák, Béla; Oliveira, Raquel A.
    Sister chromatid cohesion, mediated by the cohesin complex, is essential for faithful mitosis. Nevertheless, evidence suggests that the surveillance mechanism that governs mitotic fidelity, the spindle assembly checkpoint (SAC), is not robust enough to halt cell division when cohesion loss occurs prematurely. The mechanism behind this poor response is not properly understood. Using developing Drosophila brains, we show that full sister chromatid separation elicits a weak checkpoint response resulting in abnormal mitotic exit after a short delay. Quantitative live-cell imaging approaches combined with mathematical modeling indicate that weak SAC activation upon cohesion loss is caused by weak signal generation. This is further attenuated by several feedback loops in the mitotic signaling network. We propose that multiple feedback loops involving cyclin-dependent kinase 1 (Cdk1) gradually impair error-correction efficiency and accelerate mitotic exit upon premature loss of cohesion. Our findings explain how cohesion defects may escape SAC surveillance.
  • Centromere-Independent Accumulation of Cohesin at Ectopic Heterochromatin Sites Induces Chromosome Stretching during Anaphase
    Publication . Oliveira, Raquel A.; Kotadia, Shaila; Tavares, Alexandra; Mirkovic, Mihailo; Bowlin, Katherine; Eichinger, Christian S.; Nasmyth, Kim; Sullivan, William
    Pericentric heterochromatin, while often considered as "junk" DNA, plays important functions in chromosome biology. It contributes to sister chromatid cohesion, a process mediated by the cohesin complex that ensures proper genome segregation during nuclear division. Long stretches of heterochromatin are almost exclusively placed at centromere-proximal regions but it remains unclear if there is functional (or mechanistic) importance in linking the sites of sister chromatid cohesion to the chromosomal regions that mediate spindle attachment (the centromere). Using engineered chromosomes in Drosophila melanogaster, we demonstrate that cohesin enrichment is dictated by the presence of heterochromatin rather than centromere proximity. This preferential accumulation is caused by an enrichment of the cohesin-loading factor (Nipped-B/NIPBL/Scc2) at dense heterochromatic regions. As a result, chromosome translocations containing ectopic pericentric heterochromatin embedded in euchromatin display additional cohesin-dependent constrictions. These ectopic cohesion sites, placed away from the centromere, disjoin abnormally during anaphase and chromosomes exhibit a significant increase in length during anaphase (termed chromatin stretching). These results provide evidence that long stretches of heterochromatin distant from the centromere, as often found in many cancers, are sufficient to induce abnormal accumulation of cohesin at these sites and thereby compromise the fidelity of chromosome segregation.