In vivo and in vitro analysis of RNases in Bacillus subtilis

RNA degradation is a key process in the control of gene expression in bacteria and is essential for the cell’s homeostasis of nucleotide pools. A key player is the so-called RNA degradosome, proposed to be a membrane-associated complex containing endo- and exoribunucleases, as well as glycolytic enzymes and a DEAD-box RNA helicase. It is believed that endonuclease RNase Y is central to the formation of the RNA degradosome in Bacillus subtilis, leading to recruitment of RNases PnpA, RNase J1, and RNase J2, RNA helicase CshA, as well as glycolytic enzymes enolase and phosphofructokinase occurs. RNase Y has a transmembrane helix, and is "quasi“ essential; it is also required for mRNA processing following transcription. RNase Y also interacts with the so-called Y‑complex, consisting of YaaT, YlbF, and, YmcA (RicT, RicF, RicA), which is important for RNase Y-mediated processing of mRNA. How RNase Y can operate in two different protein complexes and acts in RNA decay as well as transcription-associated processes, is unclear. In this work, I show that the RNA degradosome is quite dynamic, having RNase Y, the glycolytic enzyme enolase, and the RNA helicase CshA and PnpA as central parts strongly reacting to a block in transcription, and thus to loss of mRNA substrate, while RNase J1 and J2 as well as the glycolytic enzyme phosphofructokinase show a much weaker response and are thus likely more peripheral components. The Y-complex clearly shows diffusion within the cytosol, but also the formation of membrane-associated accumulations, dissociating when transcription is blocked. Single molecule tracking (SMT) shows that the loss of one component of the Y-complex does not strongly affect the dynamics of the other proteins, suggesting that the complex forms a flexible association rather than a 1:1:1 stoichiometry. Biochemical analyses suggest that YaaT forms a membrane-anchor for the Y-complex, although it also has a cytosolic, freely diffusing fraction. A model will be presented that the Y-complex could function as an adaptor between nucleoid-associated mRNA synthesis and membrane-associated processing and degradation. II My thesis also presents a protocol for the successful purification of membrane-associated RNase Y, as a basis for further biochemical characterization of the protein, and interaction studies. Another essential process in which RNases play a crucial role is DNA replication. In addition to the RNA primers, which are necessary for the placement of Okazaki fragments, DNA/RNA hybrids must be processed and RNA must be removed to ensure the stability of the DNA. A part of the thesis work shows that B. subtilis replication forks intimately employ two RNases of the „H“ family, DNA polymerase A and exonuclease ExoR in vivo. Recruitment appears to be based on substrate availability rather than on specific protein/protein interactions, involving redundant enzymatic activities

Post monitoring of a cyclodextrin remediated chlorinated solvent contaminated aquifer

Hydroxypropyl-β-cyclodextrin (HPβCD) has been tested successfully in the laboratory and in the field for enhanced flushing of low-polarity contaminants from aquifers. The cyclodextrin molecule forms a toroidal structure, which has a hydrophobic cavity. Within this cavity, organic compounds of appropriate shape and size can form inclusion complexes, which is the basis for the use of cyclodextrin in groundwater remediation. The hydrophilic exterior of the molecule makes cyclodextrin highly water-soluble. The solubility of cyclodextrins can be further enhanced by adding functional groups, such as hydroxypropyl groups, to the cyclodextrin core. The aqueous solubility of HPβCD exceeds 950 g/L. These high solubilities are advantageous for field applications because they permit relatively high concentrations of the flushing agent. In order for cyclodextrin to become a feasible remediative alternative, it must be demonstrate a short term resistance to biodegradation during field application, but ultimately biodegrade so as not to pose a long term presence in the aquifer. The potential for degradation of cyclodextrin as well as changes in the chlorinated solvents and groundwater geochemistry were examined during the post monitoring of a field demonstration in a shallow aquifer at Little Creek Naval Amphibious Base in Virginia. It was found that a portion of the cyclodextrin remaining in the aquifer after the cessation of field activities biodegraded during the 425 days of post monitoring. This degradation also led to the degradation of the chlorinated solvents trichloroethylene and 1,1-trichloroethane through both biological and chemical processes. The aquifer remained anaerobic with average dissolved oxygen levels below 0.5 mg/L. Dissolved nitrate and sulfate concentrations within the cyclodextrin plume decreased due their being used as terminal electron acceptors during the degradation of the cyclodextrin. The concentrations of total iron at the field site showed no change over time. It can be concluded from this research that cyclodextrin remaining in the subsurface after cessation of active remediation will degrade due to microbial processes. The chlorinated solvents will also degrade through both chemical and biological processes to their daughter products. The terminal electron acceptors present within the cyclodextrin plume will also be used for energy during the degradation processes

Mobile assessment of heightened skin conductance in posttraumatic stress disorder

Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/137606/1/da22610.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/137606/2/da22610_am.pd

Influence of cognitive reserve on neuropsychological functioning in bipolar disorder: Findings from a 5‐year longitudinal study

Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/136365/1/bdi12470_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/136365/2/bdi12470.pd

Equivalent linear change in cognition between individuals with bipolar disorder and healthy controls over 5 years

ObjectivesCognitive dysfunction is a key feature of bipolar disorder (BD). However, not much is known about its temporal stability, as some studies have demonstrated a neurodegenerative model in BD while others have shown no change in cognitive functioning over time. Building upon our prior work, which examined the natural course of executive functioning, the current study aimed to investigate the natural course of memory, emotion processing, and fine motor dexterity over a 5‐year period in BD and healthy control (HC) samples.MethodsUsing a 5‐year longitudinal cohort, 90 individuals with BD and 17 HCs were administered a battery of neuropsychological tests at study baseline and at 1 and 5 years after study entry that captured four areas of cognitive performance: visual memory, auditory memory, emotion processing, and fine motor dexterity.ResultsLatent growth curve modeling showed no group differences in the slopes of any of the cognitive factors between the BD and HC groups. Age at baseline was negatively associated with visual memory, emotion processing, and fine motor dexterity. Education level was positively associated with auditory and visual memory and fine motor. Female gender was negatively associated with emotion processing.ConclusionsExtending our prior work on longitudinal evaluation of executive functioning, individuals with BD show similar linear change in other areas of cognitive functioning including memory, emotion processing, and fine motor dexterity as compared to unaffected HCs. Age, education, and gender may have some differential effects on cognitive changes.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/142144/1/bdi12532.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/142144/2/bdi12532_am.pd

Reflections and Considerations on Running Creative Visualization Learning Activities

This paper draws together nine strategies for creative visualization activities. Teaching visualization often involves running learning activities where students perform tasks that directly support one or more topics that the teacher wishes to address in the lesson. As a group of educators and researchers in visualization, we reflect on our learning experiences. Our activities and experiences range from dividing the tasks into smaller parts, considering different learning materials, to encouraging debate. With this paper, our hope is that we can encourage, inspire, and guide other educators with visualization activities. Our reflections provide an initial starting point of methods and strategies to craft creative visualisation learning activities, and provide a foundation for developing best practices in visualization education

Reflections and Considerations on Running Creative Visualization Learning Activities

This paper draws together nine strategies for creative visualization activities. Teaching visualization often involves running learning activities where students perform tasks that directly support one or more topics that the teacher wishes to address in the lesson. As a group of educators and researchers in visualization, we reflect on our learning experiences. Our activities and experiences range from dividing the tasks into smaller parts, considering different learning materials, to encouraging debate. With this paper, our hope is that we can encourage, inspire, and guide other educators with visualization activities. Our reflections provide an initial starting point of methods and strategies to craft creative visualisation learning activities, and provide a foundation for developing best practices in visualization education.Comment: 8 pages, 3 figures. Accepted at 4th IEEE Workshop on Visualization Guidelines in Research, Design, and Education (VisGuides 2022), at IEEE VIS 202

The patient reporting and action for a safe environment (PRASE) intervention: a feasibility study

Background: There is growing interest in the role of patients in improving patient safety. One such role is providing feedback on the safety of their care. Here we describe the development and feasibility testing of an intervention that collects patient feedback on patient safety, brings together staff to consider this feedback and to plan improvement strategies. We address two research questions: i) to explore the feasibility of the process of systematically collecting feedback from patients about the safety of care as part of the PRASE intervention; and, ii) to explore the feasibility and acceptability of the PRASE intervention for staff, and to understand more about how staff use the patient feedback for service improvement. Method: We conducted a feasibility study using a wait-list controlled design across six wards within an acute teaching hospital. Intervention wards were asked to participate in two cycles of the PRASE (Patient Reporting & Action for a Safe Environment) intervention across a six-month period. Participants were patients on participating wards. To explore the acceptability of the intervention for staff, observations of action planning meetings, interviews with a lead person for the intervention on each ward and recorded researcher reflections were analysed thematically and synthesised. Results: Recruitment of patients using computer tablets at their bedside was straightforward, with the majority of patients willing and able to provide feedback. Randomisation of the intervention was acceptable to staff, with no evidence of differential response rates between intervention and control groups. In general, ward staff were positive about the use of patient feedback for service improvement and were able to use the feedback as a basis for action planning, although engagement with the process was variable. Gathering a multidisciplinary team together for action planning was found to be challenging, and implementing action plans was sometimes hindered by the need to co-ordinate action across multiple services. Discussion: The PRASE intervention was found to be acceptable to staff and patients. However, before proceeding to a full cluster randomised controlled trial, the intervention requires adaptation to account for the difficulties in implementing action plans within three months, the need for a facilitator to support the action planning meetings, and the provision of training and senior management support for participating ward teams. Conclusions: The PRASE intervention represents a promising method for the systematic collection of patient feedback about the safety of hospital care