An investigation into the function of sumoylation in genomic stability in schizosaccharomyces pombe

Sumoylation is an essential posttranslational modification involved in many cellular processes such as DNA replication, chromosomal stability, cytokinesis, DNA damage responses and many others. The process of sumoylation is conserved in all eukaryotic organisms. This study involves the analysis of various aspects of sumoylation in the unicellular model organism Schizosaccharomyces pombe. The first part of this study is concerned with elucidating the functional and structural importance of a SUMO-like domain (SLD) and a putative SUMO-binding domain (SBM) present in the essential protein Rad60. Biochemical and genetical analysis reveals that SLD2 is required for the DNA damage response function of Rad60 but that the putative SBM3 is a key structural feature of the hydrophobic core of SLD2 and therefore unlikely to function as a SUMO-interacting motif. As Rad60 interacts with the SUMO E3 ligase Pli1, which facilitates overall sumoylation and SUMO chain formation, further analysis was undertaken to identify the function of SUMO chain formation and the function of Pli1 in maintaining chromosomal stability. A SUMO chain mutant, Pmt3-K14R; K30R, was characterized and shown to be sensitive to the DNA replication inhibitor hydroxyurea. Analysis of the pli1 null mutant reveals that Pli1 dependent sumoylation has multiple functions at the centromeric repetitive sequences as the mutant displays increased gene conversion at centromeric regions and increased loss of an artificial minichromosome rates compared to a wild type strain. The second part of this study is concerned with identifying specific modified lysine residues in the sumoylation pathway components Fub2, Hus5 and Nse2 and the target proteins Rtf2 and PCNA. After identification of in vitro sumoylation sites, an analysis of the sumoylation of the SUMO conjugating enzyme Hus5 and the sumoylation of the Rtf2 protein is carried out. In vivo and genetical analysis of the hus5-K50R mutant suggests that the sumoylation of the conjugating enzyme is required for maintaining the homeostasis of the pathway and is essential for cell viability when the homologous recombination machinery is impaired. Sumoylation of Rtf2 protein is required for the response to the DNA alkylating agent MMS and, like the sumoylation of Hus5, is essential for cell viability in homologous recombination mutant backgrounds

Increasing Collective Teacher Efficacy through Inuit Qaujimajatuqangit and Coaching: A Community-Based Approach

Abstract Instructional coaching in one remote northern community focuses on teacher mentorship and professional learning and prioritizes the implementation of prepackaged literacy resources. This focus on the implementation of prepackaged programs and data collection, promotes neo-liberal priorities and the privatization of education, over the methods that support the inclusion of Inuit cultural knowledge. Kostogriz (2011) states that we should be cautious about accepting the neo-liberal discourse of the literacy crisis and standardized reforms, the author suggests that low levels of literacy are not a large-scale problem so much as a result of under-provision of a socially just education. This Problem of Practice investigates the role of the learning coach in increasing collective teacher efficacy in accordance with the principals of Inuit Qaujimajatuqangit (IQ). Coaching to improve collective teacher efficacy through the integration of IQ may show greater educational improvement than literacy coaching alone. This vision for change includes an analysis coaching philosophy, leading from the middle, critical theory, organizational learning, the practice of decolonizing education and the positive effects of improving collective teacher efficacy. A PESTE analysis of the political, socio-economic factors; Bolman and Deal’s (2008) human resource frame and Nadler and Tushman’s (1980) congruence model will guide the critical organizational analysis. The leadership approaches to change are transformational, transformative and include ethical and decolonizing methods. The Change Path Model of Cawsey, Deszca, and Ingols (2016) is the chosen organizational framework for leading change in this Organizational Improvement Plan. Analysis of relevant internal data presents information on student educational attainment levels, and external data describes the positive effects of increasing collective efficacy on student achievement and school improvement. Hall and Hord’s (2006) six functions guide the coaching process and Change Path Model implementation planning (Cawsey et al., (2016). Implementation is presented through a series of Plan Do Study Act cycles and supported with a plan to communicate change (Conzemius & O’Neill, 2014). Coaching to influence teacher collaboration with a greater emphasis on IQ approaches and knowledge is seen as a key pathway in the leadership model to increase collective efficacy beliefs. Keywords: collective teacher efficacy, learning coach, Inuit Qaujimajatuqangit (IQ), transformational leadership, transformative leadership, ethical leadership, decolonizing education, Change Path Model

Characterisation of the SUMO-like domains of Schizosaccharomyces pombe Rad60

The S. pombe Rad60 protein is required for the repair of DNA double strand breaks, recovery from replication arrest, and is essential for cell viability. It has two SUMO-like domains (SLDs) at its C-terminus, an SXS motif and three sequences that have been proposed to be SUMO-binding motifs (SBMs). SMB1 is located in the middle of the protein, SBM2 is in SLD1 and SBM3 is at the C-terminus of SLD2. We have probed the functions of the two SUMO-like domains, SLD1 and SLD2, and the putative SBMs. SLD1 is essential for viability, while SLD2 is not. rad60-SLD2Δ cells are sensitive to DNA damaging agents and hydroxyurea. Neither ubiquitin nor SUMO can replace SLD1 or SLD2. Cells in which either SBM1 or SBM2 has been mutated are viable and are wild type for response to MMS and HU. In contrast mutation of SBM3 results in significant sensitivity to MMS and HU. These results indicate that the lethality resulting from deletion of SLD1 is not due to loss of SBM2, but that mutation of SBM3 produces a more severe phenotype than does deletion of SLD2. Using chemical denaturation studies, FPLC and dynamic light scattering we show this is likely due to the destabilisation of SLD2. Thus we propose that the region corresponding to the putative SBM3 forms part of the hydrophobic core of SLD2 and is not a SUMO-interacting motif. Over-expression of Hus5, which is the SUMO conjugating enzyme and known to interact with Rad60, does not rescue rad60-SLD2Δ, implying that as well as having a role in the sumoylation process as previously described [1], Rad60 has a Hus5-independent function

Dynamic optimal taxation with human capital.

This paper revisits the dynamic optimal taxation results of Jones, Manuelli, and Rossi (1993, 1997). They use a growth model with human capital and find that optimal taxes on both capital income and labor income converge to zero in steady state. For one of the models under consideration, I show that the representative household's problem does not have an interior solution. This raises concerns since these corners are inconsistent with aggregate data. Interiority is restored if preferences are modified so that human capital augments the value of leisure time. With this change, the optimal tax problem is analyzed and, reassuringly, the Jones, Manuelli, and Rossi results are confirmed: neither capital income nor labor income should be taxed in steady state

SUMO chain formation is required for response to replication arrest in S. pombe

SUMO is a ubiquitin-like protein that is post-translationally attached to one or more lysine residues on target proteins. Despite having only 18% sequence identity with ubiquitin, SUMO contains the conserved betabetaalphabetabetaalphabeta fold present in ubiquitin. However, SUMO differs from ubiquitin in having an extended N-terminus. In S. pombe the N-terminus of SUMO/Pmt3 is significantly longer than those of SUMO in S. cerevisiae, human and Drosophila. Here we investigate the role of this N-terminal region. We have used two dimensional gel electrophoresis to demonstrate that S. pombe SUMO/Pmt3 is phosphorylated, and that this occurs on serine residues at the extreme N-terminus of the protein. Mutation of these residues (in pmt3-1) results in a dramatic reduction in both the levels of high Mr SUMO-containing species and of total SUMO/Pmt3, indicating that phosphorylation of SUMO/Pmt3 is required for its stability. Despite the significant reduction in high Mr SUMO-containing species, pmt3-1 cells do not display an aberrant cell morphology or sensitivity to genotoxins or stress. Additionally, we demonstrate that two lysine residues in the N-terminus of S. pombe SUMO/Pmt3 (K14 and K30) can act as acceptor sites for SUMO chain formation in vitro. Inability to form SUMO chains results in aberrant cell and nuclear morphologies, including stretched and fragmented chromatin. SUMO chain mutants are sensitive to the DNA synthesis inhibitor, hydroxyurea (HU), but not to other genotoxins, such as UV, MMS or CPT. This implies a role for SUMO chains in the response to replication arrest in S. pomb

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

Effectiveness of a national quality improvement programme to improve survival after emergency abdominal surgery (EPOCH): a stepped-wedge cluster-randomised trial

Background: Emergency abdominal surgery is associated with poor patient outcomes. We studied the effectiveness of a national quality improvement (QI) programme to implement a care pathway to improve survival for these patients. Methods: We did a stepped-wedge cluster-randomised trial of patients aged 40 years or older undergoing emergency open major abdominal surgery. Eligible UK National Health Service (NHS) hospitals (those that had an emergency general surgical service, a substantial volume of emergency abdominal surgery cases, and contributed data to the National Emergency Laparotomy Audit) were organised into 15 geographical clusters and commenced the QI programme in a random order, based on a computer-generated random sequence, over an 85-week period with one geographical cluster commencing the intervention every 5 weeks from the second to the 16th time period. Patients were masked to the study group, but it was not possible to mask hospital staff or investigators. The primary outcome measure was mortality within 90 days of surgery. Analyses were done on an intention-to-treat basis. This study is registered with the ISRCTN registry, number ISRCTN80682973. Findings: Treatment took place between March 3, 2014, and Oct 19, 2015. 22 754 patients were assessed for elegibility. Of 15 873 eligible patients from 93 NHS hospitals, primary outcome data were analysed for 8482 patients in the usual care group and 7374 in the QI group. Eight patients in the usual care group and nine patients in the QI group were not included in the analysis because of missing primary outcome data. The primary outcome of 90-day mortality occurred in 1210 (16%) patients in the QI group compared with 1393 (16%) patients in the usual care group (HR 1·11, 0·96–1·28). Interpretation: No survival benefit was observed from this QI programme to implement a care pathway for patients undergoing emergency abdominal surgery. Future QI programmes should ensure that teams have both the time and resources needed to improve patient care. Funding: National Institute for Health Research Health Services and Delivery Research Programme