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1.
J Biol Chem ; 299(3): 102890, 2023 03.
Article in English | MEDLINE | ID: mdl-36634851

ABSTRACT

Mutations in protein O-mannosyltransferases (POMTs) result in severe brain defects and congenital muscular dystrophies characterized by abnormal glycosylation of α-dystroglycan (α-Dg). However, neurological phenotypes of POMT mutants are not well understood, and the functional substrates of POMTs other than α-Dg remain unknown. Using a Drosophila model, here we reveal that Dg alone cannot account for the phenotypes of POMT mutants, and identify Protein tyrosine phosphatase 69D (PTP69D) as a gene interacting with POMTs in producing the abdomen rotation phenotype. Using RNAi-mediated knockdown, mutant alleles, and a dominant-negative form of PTP69D, we reveal that PTP69D is required for the wiring of larval sensory axons. We also found that PTP69D and POMT genes interact in this process, and that their interactions lead to complex synergistic or antagonistic effects on axon wiring phenotypes, depending on the mode of genetic manipulation. Using glycoproteomic approaches, we further characterized the glycosylation of the PTP69D transgenic construct expressed in genetic strains with different levels of POMT activity. We found that the PTP69D construct carries many O-linked mannose modifications when expressed in Drosophila with wild-type or ectopically upregulated expression of POMTs. These modifications were absent in POMT mutants, suggesting that PTP69D is a substrate of POMT-mediated O-mannosylation. Taken together, our results indicate that PTP69D is a novel functional substrate of POMTs that is required for axon connectivity. This mechanism of POMT-mediated regulation of receptor-type protein tyrosine phosphatase functions could potentially be conserved in mammals and may shed new light on the etiology of neurological defects in muscular dystrophies.


Subject(s)
Axons , Drosophila , Mannosyltransferases , Protein Tyrosine Phosphatases , Animals , Axons/metabolism , Drosophila/enzymology , Drosophila/metabolism , Drosophila Proteins/genetics , Dystroglycans/genetics , Dystroglycans/metabolism , Mammals/metabolism , Mannosyltransferases/metabolism , Protein Tyrosine Phosphatases/metabolism , Receptor-Like Protein Tyrosine Phosphatases/genetics
2.
Sci Data ; 9(1): 722, 2022 11 24.
Article in English | MEDLINE | ID: mdl-36433985

ABSTRACT

Plasmodium cynomolgi causes zoonotic malarial infections in Southeast Asia and this parasite species is important as a model for Plasmodium vivax and Plasmodium ovale. Each of these species produces hypnozoites in the liver, which can cause relapsing infections in the blood. Here we present methods and data generated from iterative longitudinal systems biology infection experiments designed and performed by the Malaria Host-Pathogen Interaction Center (MaHPIC) to delve deeper into the biology, pathogenesis, and immune responses of P. cynomolgi in the Macaca mulatta host. Infections were initiated by sporozoite inoculation. Blood and bone marrow samples were collected at defined timepoints for biological and computational experiments and integrative analyses revolving around primary illness, relapse illness, and subsequent disease and immune response patterns. Parasitological, clinical, haematological, immune response, and -omic datasets (transcriptomics, proteomics, metabolomics, and lipidomics) including metadata and computational results have been deposited in public repositories. The scope and depth of these datasets are unprecedented in studies of malaria, and they are projected to be a F.A.I.R., reliable data resource for decades.


Subject(s)
Malaria , Plasmodium cynomolgi , Animals , Host-Pathogen Interactions , Macaca mulatta , Plasmodium cynomolgi/physiology , Sporozoites , Systems Biology , Zoonoses
3.
Int J Radiat Biol ; 97(10): 1485-1501, 2021.
Article in English | MEDLINE | ID: mdl-34355643

ABSTRACT

PURPOSE: Chronic exposure to ionizing radiation (IR) at low doses (<100 mGy) has been insufficiently studied to understand fully the risk to health. Relatively little knowledge exists regarding how species and healthy tissues respond at the protein level to chronic exposure to low doses of IR, and mass spectrometric-based profiling of protein expression is a powerful tool for studying changes in protein abundance. MATERIALS AND METHODS: SDS gel electrophoresis, LC-MS/MS mass spectrometry-based approaches and bioinformatic data analytics were used to detect proteomic changes following chronic exposure to moderate/low doses of radiation in adults and normally developed Medaka fish (Oryzias latipes). RESULTS: Significant variations in the abundance of proteins involved in thyroid hormone signaling and lipid metabolism were detected, which could be related to the gonadal regression phenotype observed after 21.04 mGy and 204.3 mGy/day exposure. The global proteomic change was towards overexpression of proteins in muscle and skin, while the opposite effect was observed in internal organs. CONCLUSION: The present study provides information on the impacts of biologically relevant low doses of IR, which will be useful in future research for the identification of potential biomarkers of IR exposure and allow for a better assessment of radiation biosafety regulations.


Subject(s)
Oryzias , Animals , Chromatography, Liquid , Computational Biology , Proteomics , Radiation, Ionizing , Tandem Mass Spectrometry
4.
Cell Host Microbe ; 28(4): 586-601.e6, 2020 10 07.
Article in English | MEDLINE | ID: mdl-32841605

ABSTRACT

The SARS-CoV-2 betacoronavirus uses its highly glycosylated trimeric Spike protein to bind to the cell surface receptor angiotensin converting enzyme 2 (ACE2) glycoprotein and facilitate host cell entry. We utilized glycomics-informed glycoproteomics to characterize site-specific microheterogeneity of glycosylation for a recombinant trimer Spike mimetic immunogen and for a soluble version of human ACE2. We combined this information with bioinformatics analyses of natural variants and with existing 3D structures of both glycoproteins to generate molecular dynamics simulations of each glycoprotein both alone and interacting with one another. Our results highlight roles for glycans in sterically masking polypeptide epitopes and directly modulating Spike-ACE2 interactions. Furthermore, our results illustrate the impact of viral evolution and divergence on Spike glycosylation, as well as the influence of natural variants on ACE2 receptor glycosylation. Taken together, these data can facilitate immunogen design to achieve antibody neutralization and inform therapeutic strategies to inhibit viral infection.


Subject(s)
Betacoronavirus/metabolism , Coronavirus Infections/enzymology , Coronavirus Infections/virology , Peptidyl-Dipeptidase A/metabolism , Pneumonia, Viral/enzymology , Pneumonia, Viral/virology , Spike Glycoprotein, Coronavirus/metabolism , Angiotensin-Converting Enzyme 2 , COVID-19 , Glycosylation , HEK293 Cells , Humans , Molecular Dynamics Simulation , Pandemics , Peptidyl-Dipeptidase A/chemistry , Protein Domains , Protein Interaction Domains and Motifs , Receptors, Virus/chemistry , Receptors, Virus/metabolism , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/chemistry , Virus Internalization
5.
bioRxiv ; 2020 Jul 24.
Article in English | MEDLINE | ID: mdl-32743578

ABSTRACT

The current COVID-19 pandemic is caused by the SARS-CoV-2 betacoronavirus, which utilizes its highly glycosylated trimeric Spike protein to bind to the cell surface receptor ACE2 glycoprotein and facilitate host cell entry. We utilized glycomics-informed glycoproteomics to characterize site-specific microheterogeneity of glycosylation for a recombinant trimer Spike mimetic immunogen and for a soluble version of human ACE2. We combined this information with bioinformatic analyses of natural variants and with existing 3D-structures of both glycoproteins to generate molecular dynamics simulations of each glycoprotein alone and interacting with one another. Our results highlight roles for glycans in sterically masking polypeptide epitopes and directly modulating Spike-ACE2 interactions. Furthermore, our results illustrate the impact of viral evolution and divergence on Spike glycosylation, as well as the influence of natural variants on ACE2 receptor glycosylation that, taken together, can facilitate immunogen design to achieve antibody neutralization and inform therapeutic strategies to inhibit viral infection.

6.
PLoS Genet ; 16(2): e1008390, 2020 02.
Article in English | MEDLINE | ID: mdl-32084124

ABSTRACT

Base J, ß-D-glucosyl-hydroxymethyluracil, is a modification of thymine DNA base involved in RNA Polymerase (Pol) II transcription termination in kinetoplastid protozoa. Little is understood regarding how specific thymine residues are targeted for J-modification or the mechanism of J regulated transcription termination. To identify proteins involved in J-synthesis, we expressed a tagged version of the J-glucosyltransferase (JGT) in Leishmania tarentolae, and identified four co-purified proteins by mass spectrometry: protein phosphatase (PP1), a homolog of Wdr82, a potential PP1 regulatory protein (PNUTS) and a protein containing a J-DNA binding domain (named JBP3). Gel shift studies indicate JBP3 is a J-DNA binding protein. Reciprocal tagging, co-IP and sucrose gradient analyses indicate PP1, JGT, JBP3, Wdr82 and PNUTS form a multimeric complex in kinetoplastids, similar to the mammalian PTW/PP1 complex involved in transcription termination via PP1 mediated dephosphorylation of Pol II. Using RNAi and analysis of Pol II termination by RNA-seq and RT-PCR, we demonstrate that ablation of PNUTS, JBP3 and Wdr82 lead to defects in Pol II termination at the 3'-end of polycistronic gene arrays in Trypanosoma brucei. Mutants also contain increased antisense RNA levels upstream of transcription start sites, suggesting an additional role of the complex in regulating termination of bi-directional transcription. In addition, PNUTS loss causes derepression of silent Variant Surface Glycoprotein genes involved in host immune evasion. Our results suggest a novel mechanistic link between base J and Pol II polycistronic transcription termination in kinetoplastids.


Subject(s)
DNA, Kinetoplast/metabolism , Protozoan Proteins/metabolism , RNA Polymerase II/metabolism , Transcription Termination, Genetic , Trypanosoma brucei brucei/physiology , Animals , DNA, Kinetoplast/genetics , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Genes, Protozoan , Glucosides/metabolism , Glucosyltransferases/genetics , Glucosyltransferases/metabolism , Histones/genetics , Histones/metabolism , Leishmania/physiology , Mutation , Protozoan Proteins/genetics , RNA Interference , RNA Polymerase II/genetics , Thymine/metabolism , Uracil/analogs & derivatives , Uracil/metabolism
7.
Methods Protoc ; 2(3)2019 Jul 30.
Article in English | MEDLINE | ID: mdl-31366181

ABSTRACT

All terrestrial organisms are subject to evolutionary pressures associated with natural sources of ionizing radiation (IR). The legacy of human-induced IR associated with energy, weapons production, medicine, and research has changed the distribution and magnitude of these evolutionary pressures. To date, no study has systematically examined the effects of environmentally relevant doses of radiation exposure across an organismal proteome. This void in knowledge has been due, in part, to technological deficiencies that have hampered quantifiable environmentally relevant IR doses and sensitive detection of proteomic responses. Here, we describe a protocol that addresses both needs, combining quantifiable IR delivery with a reliable method to yield proteomic comparisons of control and irradiated Medaka fish. Exposures were conducted at the Savannah River Ecology Laboratory (SREL, in Aiken, SC), where fish were subsequently dissected into three tissue sets (carcasses, organs and intestines) and frozen until analysis. Tissue proteins were extracted, resolved by Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE), and each sample lane was divided into ten equal portions. Following in-gel tryptic digestion, peptides released from each gel portion were identified and quantified by Liquid Chromatography-Mass Spectrometry (LC-MS/MS) to obtain the most complete, comparative study to date of proteomic responses to environmentally relevant doses of IR. This method provides a simple approach for use in ongoing epidemiologic studies of chronic exposure to environmentally relevant levels of IR and should also serve well in physiological, developmental, and toxicological studies.

8.
Glycobiology ; 28(2): 90-99, 2018 02 01.
Article in English | MEDLINE | ID: mdl-29190349

ABSTRACT

Bacillus circulans Jordan 32352 was isolated from decaying organic matter in the New Jersey soil in the early 1930s. This soil-dwelling bacterium produced an enzyme capable of degrading the type 3 capsular polysaccharide (Pn3P) of Streptococcus pneumoniae (Spn). Early reports of this enzyme, Pn3Pase, demonstrated its inducibility by, and specificity for Pn3P. We set out to identify and clone this enzyme for its recombinant expression and characterization. We first sequenced the genome of this bacterial species, and reclassified the Pn3Pase producing bacterium as Paenibacillus species 32352. We identified the putative protein of Pn3Pase through mass spectrometry-based proteomics and cloned the gene for recombinant expression. We then characterized the oligosaccharide products generated upon the enzymatic depolymerization of Pn3P. Sequence analysis suggests that this glycoside hydrolase belongs to a new carbohydrate-active enzyme GH family. To our knowledge, this is the only enzyme to demonstrate Pn3P depolymerization activity.


Subject(s)
Bacterial Capsules/metabolism , Bacterial Proteins/metabolism , Glycoside Hydrolases/metabolism , Paenibacillus/enzymology , Polysaccharides, Bacterial/metabolism , Streptococcus pneumoniae/metabolism , Glycoside Hydrolases/chemistry , Glycoside Hydrolases/genetics
9.
Article in English | MEDLINE | ID: mdl-28174667

ABSTRACT

BACKGROUND: Overweight and obesity are a major concern that may influence the operational capacity of the UK Naval Service (NS). This study was conducted to evaluate the feasibility of trialling and implementing a modified web-based weight loss programme for overweight and obese NS personnel. METHODS: The feasibility of a web-based weight loss programme with minimal face to face support was evaluated using a non-randomised design, based on the Reach, Efficacy, Adoption, and Implementation (RE-AIM) dimensions of a framework designed for analysing implementation of interventions in practice. RESULTS: It was estimated that 6% (n = 58) of eligible NS personnel at recruitment sites were reached, based on personnel's expressions of interest to take part in the study. The potential efficacy of the intervention was evaluated by analysing participants' change in weight (kg) in the two groups. Forty-three participants were allocated to the intervention (n = 21) or control group (n = 22). Website usage was low, with 1.5 sessions accessed on average, over a 12-week follow-up. Changes in body weight over 12 weeks appeared to be observed for participants in the intervention group but not in the control group. The average weight loss observed in the intervention group (mean = -1.9 kg, SD = 2.1) appeared to reach significance, 95% CI [-2.8, -1.0], whereas no significant weight loss was apparent among control group participants (mean = -0.8 kg, SD = 3.8), 95% CI [-2.4, 0.8]. However, this feasibility study was not powered to test for within or group differences. Recruitment rates varied across five NS establishments invited to take part in the study, suggesting that the web-based weight loss programme was not adopted to the same extent across all bases. The online programme was not implemented as intended in terms of regular usage by participants and support provision by physical training instructors. CONCLUSION: The results suggest that the intervention may warrant further investigation provided that engagement with the intervention by both staff and participants can be improved.

10.
Ergonomics ; 59(2): 325-326, 2016 Feb.
Article in English | MEDLINE | ID: mdl-28185480
11.
Ergonomics ; 59(2): 195-206, 2016.
Article in English | MEDLINE | ID: mdl-26225885

ABSTRACT

An assessment of the Health and Safety Executive's (HSE) stress indicator tool was conducted to determine whether it was suitable for use with Ministry of Defence (MoD) personnel. A total of 1031 respondents from the Army, Navy, Air Force and MoD civilians completed a questionnaire containing the HSE tool and supplementary measures of work-life balance, engagement, deployment, leave taken and hours of work. Six measures of adverse reaction to the stressors were also reported: perceptions of job stress, job stress and health, psychological strain, fatigue after work, work ability and quality of working life. The stressor scales, particularly the 'demands' and 'relationships' scales, were associated with adverse outcomes as was the work-life balance scale. The HSE tool had some validity when used with MoD personnel, but its content was too narrow. The content validity of the tool can be improved for use in a military context with the addition of a 'work-life balance' scale'. PRACTITIONER SUMMARY: The HSE stress tool was tested with a mixed sample of MoD employees. The 'Demands and "Relationships" scales were associated with adverse outcomes. An additional Work­Life Balance' scale improved the content validity, demonstrating the importance of assessing the psychometric qualities of scales when used within particular contexts to ensure validity.


Subject(s)
Military Personnel/psychology , Occupational Diseases/diagnosis , Stress, Psychological/diagnosis , Surveys and Questionnaires/standards , Workplace/psychology , Adult , Fatigue/psychology , Female , Humans , Job Satisfaction , Male , Middle Aged , Occupational Diseases/psychology , Psychometrics , Quality of Life , Stress, Psychological/psychology , United Kingdom , Work-Life Balance
12.
J Back Musculoskelet Rehabil ; 28(1): 119-28, 2015.
Article in English | MEDLINE | ID: mdl-25061033

ABSTRACT

BACKGROUND: Psychosocial factors are known to play a key role in determining the progress of back pain patients. However, it is not known whether these factors are applicable to military personnel, who tend to be fitter than the general population. OBJECTIVE: The aim was to identify physical and psychological predictors in a prospective study of the outcome of back pain rehabilitation over 6 months and a longer follow-up time of between 15 and 32 months. METHODS: Two hundred and fifty military personnel reporting for a residential rehabilitation programme completed a battery of physical and psychological tests. The physical tests included 800 m run time and the Biering-Sorensen test. The psychological/psychosocial measures included items on fear avoidance, self efficacy, anxiety and depression and occupational psychosocial factors such as job satisfaction. RESULTS: Self efficacy and 800 m run time predicted self-reported functional ability at 6 months and medical discharge/return to full fitness at 15­32 months. Patients with 800 m run times of more than 3 minutes 31 seconds had a four times greater chance of medical discharge from the Armed forces. CONCLUSIONS: Eight hundred metre run time and self-efficacy were independent predictors of both self-reported functional ability at 6 months and return to full fitness/medical discharge at 15­32 months. Self-efficacy also predicted 40% of the variance in the intensity of back pain and 10% of other non-back pain. Rehabilitation should include greater emphasis on physical fitness and on improving self-efficacy.


Subject(s)
Activities of Daily Living/psychology , Back/physiopathology , Low Back Pain/rehabilitation , Adult , Anxiety/psychology , Depression/psychology , Fear/psychology , Female , Humans , Low Back Pain/physiopathology , Low Back Pain/psychology , Male , Military Personnel , Physical Examination , Physical Fitness , Predictive Value of Tests , Prospective Studies , Treatment Outcome
13.
Bioinformatics ; 31(2): 242-5, 2015 Jan 15.
Article in English | MEDLINE | ID: mdl-25217575

ABSTRACT

MOTIVATION: In the field of glycomics research, several different techniques are used for structure elucidation. Although multiple techniques are often used to increase confidence in structure assignments, most glycomics databases allow storing of only a single type of experimental data. In addition, the methods used to prepare a sample for analysis is seldom recorded making it harder to reproduce the analytical data and results. RESULTS: We have extended the freely available EUROCarbDB framework to allow the submission of experimental data and the reporting of several orthogonal experimental datasets. The features aim to increase the understandability and reproducibility of the reported data. AVAILABILITY AND IMPLEMENTATION: The installation with the glycan standards is available at http://glycomics.ccrc.uga.edu/eurocarb/. The source code of the project is available at https://code.google.com/p/ucdb/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.


Subject(s)
Carbohydrates/chemistry , Computational Biology/methods , Databases as Topic , Glycomics/methods , Software , Humans
14.
J Proteome Res ; 13(7): 3314-29, 2014 Jul 03.
Article in English | MEDLINE | ID: mdl-24884609

ABSTRACT

Inter-alpha-trypsin inhibitor heavy chain H4 (ITIH4) is a 120 kDa acute-phase glycoprotein produced primarily in the liver, secreted into the blood, and identified in serum. ITIH4 is involved in liver development and stabilization of the extracellular matrix (ECM), and its expression is altered in liver disease. In this study, we aimed to characterize glycosylation of recombinant and serum-derived ITIH4 using analytical mass spectrometry. Recombinant ITIH4 was analyzed to optimize glycopeptide analyses, followed by serum-derived ITIH4. First, we confirmed that the four ITIH4 N-X-S/T sequons (N81, N207, N517, and N577) were glycosylated by treating ITIH4 tryptic/GluC glycopeptides with PNGaseF in the presence of (18)O water. Next, we performed glycosidase-assisted LC-MS/MS analysis of ITIH4 trypsin-GluC glycopeptides enriched via hydrophilic interaction liquid chromatography to characterize ITIH4 N-glycoforms. While microheterogeneity of N-glycoforms differed between ITIH4 protein expressed in HEK293 cells and protein isolated from serum, occupancy of N-glycosylation sites did not differ. A fifth N-glycosylation site was discovered at N274 with the rare nonconsensus NVV motif. Site N274 contained high-mannose N-linked glycans in both serum and recombinant ITIH4. We also identified isoform-specific ITIH4 O-glycoforms and documented that utilization of O-glycosylation sites on ITIH4 differed between the cell line and serum.


Subject(s)
Glycoproteins/blood , Protein Processing, Post-Translational , Proteinase Inhibitory Proteins, Secretory/blood , Amino Acid Sequence , Blood Proteins/chemistry , Carbohydrate Conformation , Carbohydrate Sequence , Glycoproteins/chemistry , Glycosylation , HEK293 Cells , Humans , Molecular Sequence Data , Protein Isoforms , Proteinase Inhibitory Proteins, Secretory/chemistry
15.
Article in English | MEDLINE | ID: mdl-25657638

ABSTRACT

Increased flux through the hexosamine biosynthetic pathway and the corresponding increase in intracellular glycosylation of proteins via O-linked ß-N-acetylglucosamine (O-GlcNAc) is sufficient to induce insulin resistance (IR) in multiple systems. Previously, our group used shotgun proteomics to identify multiple rodent adipocytokines and secreted proteins whose levels are modulated upon the induction of IR by indirectly and directly modulating O-GlcNAc levels. We have validated the relative levels of several of these factors using immunoblotting. Since adipocytokines levels are regulated primarily at the level of transcription and O-GlcNAc alters the function of many transcription factors, we hypothesized that elevated O-GlcNAc levels on key transcription factors are modulating secreted protein expression. Here, we show that upon the elevation of O-GlcNAc levels and the induction of IR in mature 3T3-F442a adipocytes, the transcript levels of multiple secreted proteins reflect the modulation observed at the protein level. We validate the transcript levels in male mouse models of diabetes. Using inguinal fat pads from the severely IR db/db mouse model and the mildly IR diet-induced mouse model, we have confirmed that the secreted proteins regulated by O-GlcNAc modulation in cell culture are likewise modulated in the whole animal upon a shift to IR. By comparing the promoters of similarly regulated genes, we determine that Sp1 is a common cis-acting element. Furthermore, we show that the LPL and SPARC promoters are enriched for Sp1 and O-GlcNAc modified proteins during insulin resistance in adipocytes. Thus, the O-GlcNAc modification of proteins bound to promoters, including Sp1, is linked to adipocytokine transcription during insulin resistance.

16.
J Biol Chem ; 288(48): 34680-98, 2013 Nov 29.
Article in English | MEDLINE | ID: mdl-24155237

ABSTRACT

Glycan structures on glycoproteins and glycolipids play critical roles in biological recognition, targeting, and modulation of functions in animal systems. Many classes of glycan structures are capped with terminal sialic acid residues, which contribute to biological functions by either forming or masking glycan recognition sites on the cell surface or secreted glycoconjugates. Sialylated glycans are synthesized in mammals by a single conserved family of sialyltransferases that have diverse linkage and acceptor specificities. We examined the enzymatic basis for glycan sialylation in animal systems by determining the crystal structures of rat ST6GAL1, an enzyme that creates terminal α2,6-sialic acid linkages on complex-type N-glycans, at 2.4 Å resolution. Crystals were obtained from enzyme preparations generated in mammalian cells. The resulting structure revealed an overall protein fold broadly resembling the previously determined structure of pig ST3GAL1, including a CMP-sialic acid-binding site assembled from conserved sialylmotif sequence elements. Significant differences in structure and disulfide bonding patterns were found outside the sialylmotif sequences, including differences in residues predicted to interact with the glycan acceptor. Computational substrate docking and molecular dynamics simulations were performed to predict and evaluate the CMP-sialic acid donor and glycan acceptor interactions, and the results were compared with kinetic analysis of active site mutants. Comparisons of the structure with pig ST3GAL1 and a bacterial sialyltransferase revealed a similar positioning of donor, acceptor, and catalytic residues that provide a common structural framework for catalysis by the mammalian and bacterial sialyltransferases.


Subject(s)
Crystallography, X-Ray , Polysaccharides/chemistry , Sialic Acids/metabolism , Sialyltransferases/chemistry , Animals , Bacteria/enzymology , Bacteria/genetics , Binding Sites , Molecular Docking Simulation , Molecular Dynamics Simulation , Polysaccharides/biosynthesis , Protein Conformation , Rats , Sialic Acids/chemistry , Sialyltransferases/metabolism , Structure-Activity Relationship , Swine/genetics , beta-D-Galactoside alpha 2-6-Sialyltransferase
17.
Ergonomics ; 56(11): 1629-39, 2013.
Article in English | MEDLINE | ID: mdl-24047248

ABSTRACT

Questionnaire data captured in January-March 2007 were examined in relation to turnover in males and females during the next five years. In general, most of the workplace stressors (such as role conflict or peer support) were not antecedents of turnover in any group. Junior personnel with psychological strain in 2007 had an increased risk of turnover in the next five years. Low commitment to the service in 2007 increased the odds of turnover in male and female juniors and in female officers. Female juniors with less effective skills for coping with stress and who exercised less frequently on a weekly basis were more likely to leave. An incidental finding was that the odds of turnover were three times greater in female officers with children than in female officers with no children. Stress management interventions focusing on effective coping and sports and exercise participation which are targeted appropriately may improve retention.


Subject(s)
Military Personnel/psychology , Occupational Health , Personnel Turnover , Stress, Psychological , Adaptation, Psychological , Adult , Female , Humans , Male , Middle Aged , Naval Medicine , Stress, Psychological/psychology , United Kingdom , Workplace/psychology
18.
Ergonomics ; 56(10): 1515-24, 2013.
Article in English | MEDLINE | ID: mdl-23879800

ABSTRACT

The Cognitive Failures Questionnaire (CFQ) is used in ergonomics research to measure behavioural problems associated with attentiveness and memory in everyday life. CFQ scores have been related to constructs such as accident proneness and outcomes such as human error and psychological strain. The two-year test-retest reliability of the CFQ is reported together with the findings of factor analyses of CFQ data from 535 respondents. Evidence for the predictive and criterion validity and internal reliability of the CFQ is provided. Psychological strain was measured concurrently with CFQ on both testing occasions, two years apart. The test-retest reliability of the summated CFQ score was found to be 0.71, while for the General Health Questionnaire (GHQ-12) strain measure it was 0.32.The relative variance stability was five times greater for the CFQ than the GHQ, indicating that scores on these questionnaires are not covariates. The use of the CFQ as a measure of cognitive control capacity is also discussed. PRACTITIONER SUMMARY: Ergonomists have long been interested in human error and the role of high work demands due to poor equipment design and excessive workload. The CFQ measures attentiveness in daily life and is shown to have excellent psychometric properties that make it suitable for use in both laboratory and field studies as a trait measure of attentiveness in daily life.


Subject(s)
Arousal , Memory , Stress, Psychological/psychology , Surveys and Questionnaires , Adult , Female , Humans , Male , Middle Aged , Psychometrics , Reproducibility of Results , Task Performance and Analysis
19.
Accid Anal Prev ; 49: 532-5, 2012 Nov.
Article in English | MEDLINE | ID: mdl-23036431

ABSTRACT

Understanding why accidents occur in the work place has a long and convoluted history. This paper adds to this corpus of research by investigating the relationship between an individual's level of cognitive failure, psychological stress, and work place accident occurrence. Retrospective analysis of accident-case individuals vs. control-match individuals on the General Health Questionnaire (GHQ) and Cognitive Failures Questionnaire (CFQ) was undertaken from amalgamated data of two Royal Navy databases. Individuals in the accident-case sub-sample had higher GHQ and CFQ scores when compared to matched-controls. Mediated regression analysis revealed high GHQ score predicted accidents but was transmitted through high CFQ scores. Individuals who are stressed are more likely to have an accident in the workplace because of a propensity for cognitive failures. A specific recommendation to reduce accident risk in the work-place is discussed.


Subject(s)
Accident Proneness , Accidents, Occupational/psychology , Cognition , Military Personnel/psychology , Stress, Psychological , Case-Control Studies , Cohort Studies , Health Surveys , Humans , Logistic Models , Naval Medicine , Psychological Tests , Retrospective Studies , Surveys and Questionnaires , United Kingdom
20.
J Biol Chem ; 287(15): 12195-203, 2012 Apr 06.
Article in English | MEDLINE | ID: mdl-22371497

ABSTRACT

O-Linked ß-N-acetylglucosamine, or O-GlcNAc, is a dynamic post-translational modification that cycles on and off serine and threonine residues of nucleocytoplasmic proteins. The O-GlcNAc modification shares a complex relationship with phosphorylation, as both modifications are capable of mutually inhibiting the occupation of each other on the same or nearby amino acid residue. In addition to diabetes, cancer, and neurodegenerative diseases, O-GlcNAc appears to play a significant role in cell growth and cell cycle progression, although the precise mechanisms are still not well understood. A recent study also found that all four core nucleosomal histones (H2A, H2B, H3, and H4) are modified with O-GlcNAc, although no specific sites on H3 were reported. Here, we describe that histone H3, a protein highly phosphorylated during mitosis, is modified with O-GlcNAc. Several biochemical assays were used to validate that H3 is modified with O-GlcNAc. Mass spectrometry analysis identified threonine 32 as a novel O-GlcNAc site. O-GlcNAc was detected at higher levels on H3 during interphase than mitosis, which inversely correlated with phosphorylation. Furthermore, increased O-GlcNAcylation was observed to reduce mitosis-specific phosphorylation at serine 10, serine 28, and threonine 32. Finally, inhibiting OGA, the enzyme responsible for removing O-GlcNAc, hindered the transition from G2 to M phase of the cell cycle, displaying a phenotype similar to preventing mitosis-specific phosphorylation on H3. Taken together, these data indicate that O-GlcNAcylation regulates mitosis-specific phosphorylations on H3, providing a mechanistic switch that orchestrates the G2-M transition of the cell cycle.


Subject(s)
Acetylglucosamine/physiology , Histones/metabolism , Mitosis , Protein Processing, Post-Translational , Acetylglucosamine/metabolism , Amino Acid Sequence , G2 Phase , Glycosylation , HeLa Cells , Histones/chemistry , Humans , Immunoprecipitation , Molecular Sequence Data , Peptide Fragments/chemistry , Phosphorylation , Threonine/metabolism
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