Cyclin-Dependent Kinase regulation and function during meiosis

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Biology, 2010.This electronic version was submitted by the student author. The certified thesis is available in the Institute Archives and Special Collections.Cataloged from student submitted PDF version of thesis.Includes bibliographical references.Meiosis is the process by which haploid gametes are produced from a diploid progenitor cell. Accurate completion of the meiotic divisions requires a variety of modifications to the mitotic chromosome segregation machinery, which allow the reductional meiotic chromosome segregation program to occur. Oscillations in the activity of Cyclin- Dependent Kinases (CDKs) drive virtually every event in the mitotic cell cycle, including events such as cell cycle entry, DNA replication, and chromosome segregation. While much is known about the activity of CDKs, the regulation of CDK activity, and the mechanisms by which CDK activity promotes cell cycle events during vegetative growth in Saccharomyces cerevisiae, relatively little is known about the roles of CDKs during the meiotic divisions. This work examines CDK activity during meiosis, the regulation of CDK activity during meiosis, and mechanisms by which CDKs regulate proper meiotic chromosome segregation. First, a striking diversity in Clb-CDK activity is observed during meiosis, including the identification of Clb1-CDK, and Clb3-CDK as meiosis I and meiosis II specific Clb-CDKs respectively. Second, Clb3 protein is shown to be restricted to meiosis II by translational control mediated by the 5'UTR of the CLB3 message. Finally, premature production of Clb3 results in the premature separation of sister-chromatids during meiosis I.by Thomas M. Carlile.Ph.D

Regulated Formation of an Amyloid-like Translational Repressor Governs Gametogenesis

Message-specific translational control is required for gametogenesis. In yeast, the RNA-binding protein Rim4 mediates translational repression of numerous mRNAs, including the B-type cyclin CLB3, which is essential for establishing the meiotic chromosome segregation pattern. Here, we show that Rim4 forms amyloid-like aggregates and that it is the amyloid-like form of Rim4 that is the active, translationally repressive form of the protein. Our data further show that Rim4 aggregation is a developmentally regulated process. Starvation induces the conversion of monomeric Rim4 into amyloid-like aggregates, thereby activating the protein to bring about repression of translation. At the onset of meiosis II, Rim4 aggregates are abruptly degraded allowing translation to commence. Although amyloids are best known for their role in the etiology of diseases such as Alzheimer’s, Parkinson’s, and diabetes by forming toxic protein aggregates, our findings show that cells can utilize amyloid-like protein aggregates to function as central regulators of gametogenesis.Charles A. King Trust (Postdoctoral Fellowship)American Cancer Society (Fellowship)National Institutes of Health (U.S.) (Grants GM62207, GM77537, and GM094303

Health and Health Care of Mothers and Children in a Suburban Area of Luanda, Angola

Population health data available in Angola are often insufficient to guide the planning of health interventions. To address this gap, the goal of the present study was to investigate the health of mothers and infants in a suburban municipality in Luanda (Cacuaco), in order to provide a baseline for future comparisons. This was a prevalence study investigating infants younger than 2 years of age and their mothers. Mothers were interviewed, and children’s height and weight were measured. Of 749 mothers interviewed, 98.5 % (95 % CI 98.2–99.1 %) had at least one prenatal visit and 51.7 % (95 % CI 47.4–56.3 %) had a health card. Most mothers with a health card had their first prenatal visit before the 20th week of pregnancy, and had at least four prenatal visits; 81.1 % (95 % CI 78.3–84.1 %) of mothers also had their child’s health card. Prevalence of exclusive breastfeeding at 6 months was 19 % (95 % CI 16.2–23.1 %). Prevalence of low height-for-age and low BMI-for-age were 32 and 6 %, respectively. Mothers with higher education levels were more likely to have had their first prenatal visit earlier, to have had more prenatal visits, to have given birth at a health facility, and to have her own and her child’s health cards. Results showed a high prevalence of prenatal care and a low frequency of acute malnutrition. Maternal education level, among factors studied, was the predominant correlate of more positive health behaviors. These findings suggest important progress of mother and child health in Cacuaco, and may serve as a baseline for the planning of health interventions

Morphological Instabilities in a growing Yeast Colony: Experiment and Theory

We study the growth of colonies of the yeast Pichia membranaefaciens on agarose film. The growth conditions are controlled in a setup where nutrients are supplied through an agarose film suspended over a solution of nutrients. As the thickness of the agarose film is varied, the morphology of the front of the colony changes. The growth of the front is modeled by coupling it to a diffusive field of inhibitory metabolites. Qualitative agreement with experiments suggests that such a coupling is responsible for the observed instability of the front.Comment: RevTex, 4 pages and 3 figure

Rebaling of silage and haylage and its effects on forage microbial and chemical composition - A pilot study

Use of big bale silage and haylage can be difficult on farms where daily forage consumption is comparatively low as speed of deterioration of forage after bale opening may be faster than feed-out rate. Production of smaller bales at harvest is possible, but expensive and work-intensive. Therefore, a pilot study of rebaling forage stored in big bales to smaller bales was conducted. Three separate experiments were included, where microbial and chemical composition of silage and haylage was studied before and after rebaling. In Experiment III, residual big bale forage stored and opened together with rebaled forage was included. Results showed that rebaled haylage and silage had higher yeast counts compared to initial forage; however, residual bales in Experiment III had yeast counts similar to rebaled forage, indicating an effect of storage time rather than of rebaling. In Experiment II, mould counts were higher in rebaled compared to initial silage, but not in haylage. Chemical composition was similar in initial and rebaled forage except for ammonia-N. In Experiment III, ammonia-N was higher in rebaled compared to initial and residual forage and was the only chemical variable affected by rebaling. Bale temperature during aerobic storage followed ambient temperature until day 6-8 in Experiment I and until day 14 in Experiment III where ambient temperature was lower. In conclusion, rebaling can be done without large changes in chemical composition of the forage, but yeast and mould counts may be higher in rebaled forage, and this risk should be considered when using this procedure

Management by boundaries : Insights into the role of boundary objects in a community-based tourism development project

Community-based tourism development typically assumes co-operation between different stakeholder groups at the local level, and thus combines different types of knowledge. However, this does not imply that a consensus exists between the stakeholders in the first place. In this article, we present a potential conceptual tool, namely boundary objects that could support stakeholders from different knowledge communities in working jointly towards a common goal and generate commitment towards it. The literature concerning knowledge communities and boundary objects is used as a theoretical framework. A three-year community-based tourism development project comprises the data of the article, and is used as a case study to illustrate the role of different knowledge communities, and to analyse the selected boundary objects. The results illustrate the importance of proper design of boundary objects in community-based tourism development processes, and highlight the features of a successful boundary object in generating ownership feelings towards development activities. (C) 2018 Elsevier Ltd. All rights reserved.Peer reviewe

The NSAID glafenine rescues class 2 CFTR mutants via cyclooxygenase 2 inhibition of the arachidonic acid pathway

Most cases of cystic fibrosis (CF) are caused by class 2 mutations in the cystic fibrosis transmembrane regulator (CFTR). These proteins preserve some channel function but are retained in the endoplasmic reticulum (ER). Partial rescue of the most common CFTR class 2 mutant, F508del-CFTR, has been achieved through the development of pharmacological chaperones (Tezacaftor and Elexacaftor) that bind CFTR directly. However, it is not clear whether these drugs will rescue all class 2 CFTR mutants to a medically relevant level. We have previously shown that the nonsteroidal anti-inflammatory drug (NSAID) ibuprofen can correct F508del-CFTR trafficking. Here, we utilized RNAi and pharmacological inhibitors to determine the mechanism of action of the NSAID glafenine. Using cellular thermal stability assays (CETSAs), we show that it is a proteostasis modulator. Using medicinal chemistry, we identified a derivative with a fourfold increase in CFTR corrector potency. Furthermore, we show that these novel arachidonic acid pathway inhibitors can rescue difficult-to-correct class 2 mutants, such as G85E-CFTR > 13%, that of non-CF cells in well-differentiated HBE cells. Thus, the results suggest that targeting the arachidonic acid pathway may be a profitable way of developing correctors of certain previously hard-to-correct class 2 CFTR mutations

A framework for real-time monitoring, analysis and adaptive sampling of viral amplicon nanopore sequencing

The ongoing SARS-CoV-2 pandemic demonstrates the utility of real-time sequence analysis in monitoring and surveillance of pathogens. However, cost-effective sequencing requires that samples be PCR amplified and multiplexed via barcoding onto a single flow cell, resulting in challenges with maximising and balancing coverage for each sample. To address this, we developed a real-time analysis pipeline to maximise flow cell performance and optimise sequencing time and costs for any amplicon based sequencing. We extended our nanopore analysis platform MinoTour to incorporate ARTIC network bioinformatics analysis pipelines. MinoTour predicts which samples will reach sufficient coverage for downstream analysis and runs the ARTIC networks Medaka pipeline once sufficient coverage has been reached. We show that stopping a viral sequencing run earlier, at the point that sufficient data has become available, has no negative effect on subsequent down-stream analysis. A separate tool, SwordFish, is used to automate adaptive sampling on Nanopore sequencers during the sequencing run. This enables normalisation of coverage both within (amplicons) and between samples (barcodes) on barcoded sequencing runs. We show that this process enriches under-represented samples and amplicons in a library as well as reducing the time taken to obtain complete genomes without affecting the consensus sequence

Sticky knowledge: A possible model for investigating implementation in healthcare contexts

<p>Abstract</p> <p>Background</p> <p>In health care, a well recognized gap exists between what we know should be done based on accumulated evidence and what we actually do in practice. A body of empirical literature shows organizations, like individuals, are difficult to change. In the business literature, knowledge management and transfer has become an established area of theory and practice, whilst in healthcare it is only starting to establish a firm footing. Knowledge has become a business resource, and knowledge management theorists and practitioners have examined how knowledge moves in organisations, how it is shared, and how the return on knowledge capital can be maximised to create competitive advantage. New models are being considered, and we wanted to explore the applicability of one of these conceptual models to the implementation of evidence-based practice in healthcare systems.</p> <p>Methods</p> <p>The application of a conceptual model called sticky knowledge, based on an integration of communication theory and knowledge transfer milestones, into a scenario of attempting knowledge transfer in primary care.</p> <p>Results</p> <p>We describe Szulanski's model, the empirical work he conducted, and illustrate its potential applicability with a hypothetical healthcare example based on improving palliative care services. We follow a doctor through two different posts and analyse aspects of knowledge transfer in different primary care settings. The factors included in the sticky knowledge model include: causal ambiguity, unproven knowledge, motivation of source, credibility of source, recipient motivation, recipient absorptive capacity, recipient retentive capacity, barren organisational context, and arduous relationship between source and recipient. We found that we could apply all these factors to the difficulty of implementing new knowledge into practice in primary care settings.</p> <p>Discussion</p> <p>Szulanski argues that knowledge factors play a greater role in the success or failure of a knowledge transfer than has been suspected, and we consider that this conjecture requires further empirical work in healthcare settings.</p

THE ROLE OF INTERDEPENDENCE IN THE MICRO-FOUNDATIONS OF ORGANIZATION DESIGN: TASK, GOAL, AND KNOWLEDGE INTERDEPENDENCE

Interdependence is a core concept in organization design, yet one that has remained consistently understudied. Current notions of interdependence remain rooted in seminal works, produced at a time when managers’ near-perfect understanding of the task at hand drove the organization design process. In this context, task interdependence was rightly assumed to be exogenously determined by characteristics of the work and the technology. We no longer live in that world, yet our view of interdependence has remained exceedingly task-centric and our treatment of interdependence overly deterministic. As organizations face increasingly unpredictable workstreams and workers co-design the organization alongside managers, our field requires a more comprehensive toolbox that incorporates aspects of agent-based interdependence. In this paper, we synthesize research in organization design, organizational behavior, and other related literatures to examine three types of interdependence that characterize organizations’ workflows: task, goal, and knowledge interdependence. We offer clear definitions for each construct, analyze how each arises endogenously in the design process, explore their interrelations, and pose questions to guide future research