The Roles of eIF2 Kinases PKR and GCN2 during Mouse Adenovirus Type 1 Infection

During viral infection, a major innate host defense mechanism is to reduce global protein synthesis by phosphorylation of eukaryotic translation inhibition factor 2a (eIF2a). eIF2a is phosphorylated by four different cellular kinases, protein kinase R (PKR), general control nonderepressible 2 (GCN2), PKR-like endoplasmic reticulum kinase, and heme-regulated inhibitor, each responding to a different cellular stress. GCN2, which phosphorylates eIF2a in response to amino acid starvation, UV irradiation, and oxidative stress, can also phosphorylate eIF2a in response to viral infection. PKR senses double-stranded RNA produced by virus infection. Many viruses have methods of inhibiting PKR activation or its downstream effects, thus circumventing protein synthesis shutdown. These methods include sequestering double-stranded RNA or producing proteins that bind to and inhibit PKR activation. Here we describe our finding that PKR was antiviral in mouse adenovirus type 1 (MAV 1) infection. We also showed that in multiple cell types, PKR was depleted during MAV 1 infection. Inhibiting the proteasome reduced the PKR depletion seen in MAV-1-infected cells, indicating that proteasomal degradation is responsible for PKR degradation during MAV-1 infection. Time course experiments showed that the degradation occurred early after infection. Infecting cells with UV-inactivated virus prevented PKR degradation, whereas inhibiting viral DNA replication did not. Together these results suggest that an early viral gene is responsible for the degradation. Degradation of PKR is a rare mechanism to oppose PKR activity, and it has only been described in six RNA viruses. To our knowledge, this is the first example of a DNA virus counteracting PKR by degrading it. In addition to PKR being degraded by MAV-1 during infection, we tested whether GCN2 plays an antiviral role, as seen for several other viruses. Here we describe that GCN2-/- deficient (atc) mice and peritoneal macrophages were significantly more susceptible to infection by MAV 1. However, atc mouse embryonic fibroblasts had similar viral yields compared to wild type mouse embryonic fibroblasts, suggesting that there is a cell-type specific antiviral effect of GCN2. There were no differences in viral yields in infected organs of atc mice at 8 days post infection compared to wild type organs, except for the cecum, indicating that the difference in survival between atc and wild type mice is not likely a result of increased viremia in atc mice. There was also no significant difference in histology of organs from MAV-1-infected atc and wild type mice. However, cytokine analysis showed that MAV-1-infected atc mice had significantly higher levels of interleukin 1a, interleukin 1b, and interferon y in the brain compared to infected wild type mouse brains at 7 days post infection, suggesting that a difference in inflammatory response could be responsible for the decreased survival of atc mice in response to MAV-1 infection. Determining how GCN2 affects the immune response to MAV 1 infection could provide insight into how GCN2 is playing an antiviral role in DNA virus infection.PHDCellular & Molecular BiologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttps://deepblue.lib.umich.edu/bitstream/2027.42/150028/1/danigood_1.pd

Meiotic cellular rejuvenation is coupled to nuclear remodeling in budding yeast.

Production of healthy gametes in meiosis relies on the quality control and proper distribution of both nuclear and cytoplasmic contents. Meiotic differentiation naturally eliminates age-induced cellular damage by an unknown mechanism. Using time-lapse fluorescence microscopy in budding yeast, we found that nuclear senescence factors - including protein aggregates, extrachromosomal ribosomal DNA circles, and abnormal nucleolar material - are sequestered away from chromosomes during meiosis II and subsequently eliminated. A similar sequestration and elimination process occurs for the core subunits of the nuclear pore complex in both young and aged cells. Nuclear envelope remodeling drives the formation of a membranous compartment containing the sequestered material. Importantly, de novo generation of plasma membrane is required for the sequestration event, preventing the inheritance of long-lived nucleoporins and senescence factors into the newly formed gametes. Our study uncovers a new mechanism of nuclear quality control and provides insight into its function in meiotic cellular rejuvenation

Children's biobehavioral reactivity to challenge predicts DNA methylation in adolescence and emerging adulthood.

A growing body of research has documented associations between adverse childhood environments and DNA methylation, highlighting epigenetic processes as potential mechanisms through which early external contexts influence health across the life course. The present study tested a complementary hypothesis: indicators of children's early internal, biological, and behavioral responses to stressful challenges may also be linked to stable patterns of DNA methylation later in life. Children's autonomic nervous system reactivity, temperament, and mental health symptoms were prospectively assessed from infancy through early childhood, and principal components analysis (PCA) was applied to derive composites of biological and behavioral reactivity. Buccal epithelial cells were collected from participants at 15 and 18 years of age. Findings revealed an association between early life biobehavioral inhibition/disinhibition and DNA methylation across many genes. Notably, reactive, inhibited children were found to have decreased DNA methylation of the DLX5 and IGF2 genes at both time points, as compared to non-reactive, disinhibited children. Results of the present study are provisional but suggest that the gene's profile of DNA methylation may constitute a biomarker of normative or potentially pathological differences in reactivity. Overall, findings provide a foundation for future research to explore relations among epigenetic processes and differences in both individual-level biobehavioral risk and qualities of the early, external childhood environment

Exercise May Ameliorate the Detrimental Side Effects of High Vitamin D Supplementation on Muscle Function in Mice

© 2020 The Authors. Journal of Bone and Mineral Research published by American Society for Bone and Mineral Research. Vitamin D is commonly prescribed to normalize deficiencies and to treat osteoporosis. However, the effect vitamin D supplements have on skeletal muscle health is equivocal. Although vitamin D is known to play a role in the various processes that maintain muscle integrity and function, recent studies utilizing high bolus dose vitamin D supplementation has demonstrated an increased risk of falls. Thus, the aim of this study was to investigate the effects of high vitamin D supplementation on skeletal muscle function with and without exercise enrichment. Four-week old C57BL/10 mice (n = 48) were separated into either normal vitamin D (1500 IU/kg diet; unsupplemented) or high vitamin D (20,000 IU/kg diet; supplemented) treatment groups. Each dietary group was further separated into interventional subgroups where mice either remained sedentary or received exercise-enrichment for 8 weeks in the form of voluntary running. Following the intervention period, whole body in vivo and ex vivo contractile analysis were performed. High vitamin D supplementation decreased force production in the slow-twitch soleus muscles of sedentary mice (p <.01); however, exercise normalized this effect. Eight weeks of exercise did not improve fatigue resistance of the extensor digitorum longus (EDL) or soleus muscles in unsupplemented mice, likely due to low levels of activation in these muscles. In contrast, fatigability was improved in the EDL (p <.01) and even more so in the soleus (p <.001) in the supplemented exercise-enriched group. Our data highlights that increasing vitamin D levels above normal reduces postural muscle force as seen in the soleus. Thus, unnecessary vitamin D supplementation may contribute to the increased risk of falls observed in some studies. Interestingly, when vitamin D supplementation was combined with exercise, force production was effectively restored, and fatigue resistance improved, even in muscles lowly activated. Regular exercise may modulate the effects of vitamin D on skeletal muscle, and be recommended for individuals receiving vitamin D supplements. © 2020 The Authors. Journal of Bone and Mineral Research published by American Society for Bone and Mineral Research

Identifying the Cytotoxic Effects of Mycobacteriophage Genes

A bacteriophage is a virus that infects and reproduces in bacteria. During productive infections—those that result in construction and release of infectious phage particles—key host cell metabolic processes are modified by the infecting phage and redirected toward making new phage particles. Protein-protein interactions are likely involved in this process. In this work, gene 80 of mycobacteriophage Vix, a gene cytotoxic to host strain Mycobacterium smegmatis, was studied. Our hypothesis was that an interaction between the Vix80 gene product and a host cell protein caused growth inhibition. Comparative analysis of the Vix80 protein sequence shows a conserved domain of unknown function 2786 (DUF2786) near the N-terminus. The Vix80 gene was dissected, and the N-terminal 66 residues, encompassing the entire DUF2786 domain, was found to be cytotoxic to M. smegmatis. DUF2786 was found to be homologous to a region of three M. smegmatis ORFs, two of which are related by alternate initiation points of the same sequence. Using in vitro protein pull-down and in vivo two-hybrid analyses, efforts are underway to look for possible interactions of Vix80 with itself and the three host proteins. As part of this process, different constructs of the Vix80 protein were expressed in Escherichia coli. We have determined that certain Vix80 constructs are also lethal to the E.coli host cell, while others permit growth, suggesting a conservation of cytotoxic function. Identifying the relevant phage and host gene products and understanding how phage exploit their host’s weaknesses could lead to new therapeutic options for many bacterial illnesses

“Lessons that simply cannot be lectured”: Highlighting the experiences and benefits of undergraduates engaged in community psychology research

Globally, Community Psychology (CP) currently struggles with a lack of visibility and a shortage of platforms from which to engage future researchers and practitioners. Many of these future researchers and practitioners study psychology as part of their undergraduate education, making psychology departments in undergraduate institutions an ideal point of exposure. Discouragingly, however, undergraduate students are unlikely to be exposed to CP in their course curriculums forcing students to seek out opportunities for exposure elsewhere (Jimenez, 2016; Glantsman, McMahon &amp; Njoku et al., 2015). This is especially true of Community Psychology within the United States. This study aimed to explore how students got involved in CP research and the benefits of that involvement. A total of 34 former undergraduate research assistants involved in CP research teams at Michigan State University completed a 20-item online questionnaire comprised of open- and closed-ended questions. Quantitative data were analyzed descriptively and qualitative data were analyzed using an iterative coding process. Results showed that the majority of students had no experience in CP and merely stumbled upon the research. Despite this, respondents reported gaining many skills, competencies, and knowledge during their time on the research teams. Respondents particularly benefited from: 1) gaining hands-on and community-based research skills; 2) attaining transferable skills they could carry with them in future positions; 3) acquiring CP-specific competencies; and 4) cultivating greater awareness and passion for social issues. Finally, the majority of respondents discussed how their involvement on CP research teams honed in their interest and passions and/or shaped their professional trajectory.&nbsp; These findings provide preliminary evidence of the benefits of CP research involvement at an undergraduate level and has important implications for how CP as a field may choose to invest in undergraduate research opportunities.&nbsp

“Lessons that simply cannot be lectured”: Highlighting the experiences and benefits of undergraduates engaged in community psychology research

Globally, Community Psychology (CP) currently struggles with a lack of visibility and a shortage of platforms from which to engage future researchers and practitioners. Many of these future researchers and practitioners study psychology as part of their undergraduate education, making psychology departments in undergraduate institutions an ideal point of exposure. Discouragingly, however, undergraduate students are unlikely to be exposed to CP in their course curriculums forcing students to seek out opportunities for exposure elsewhere (Jimenez, 2016; Glantsman, McMahon &amp; Njoku et al., 2015). This is especially true of Community Psychology within the United States. This study aimed to explore how students got involved in CP research and the benefits of that involvement. A total of 34 former undergraduate research assistants involved in CP research teams at Michigan State University completed a 20-item online questionnaire comprised of open- and closed-ended questions. Quantitative data were analyzed descriptively and qualitative data were analyzed using an iterative coding process. Results showed that the majority of students had no experience in CP and merely stumbled upon the research. Despite this, respondents reported gaining many skills, competencies, and knowledge during their time on the research teams. Respondents particularly benefited from: 1) gaining hands-on and community-based research skills; 2) attaining transferable skills they could carry with them in future positions; 3) acquiring CP-specific competencies; and 4) cultivating greater awareness and passion for social issues. Finally, the majority of respondents discussed how their involvement on CP research teams honed in their interest and passions and/or shaped their professional trajectory.&nbsp; These findings provide preliminary evidence of the benefits of CP research involvement at an undergraduate level and has important implications for how CP as a field may choose to invest in undergraduate research opportunities.&nbsp

Evaluation and establishment of a ward-based geriatric liaison service for older urological surgical patients: Proactive care of Older People undergoing Surgery (POPS)-Urology.

OBJECTIVE: To assess the impact of introducing and embedding a structured geriatric liaison service, Proactive care of Older People undergoing Surgery (POPS)-Urology, using comprehensive geriatric assessment methodology, on an inpatient urology ward. PATIENTS AND METHODS: A phased quality improvement project was undertaken using stepwise interventions. Phase 1 was a before-and-after study with initiation of a daily board round, weekly multidisciplinary meeting, and targeted geriatrician-led ward rounds for elective and emergency urology patients aged ≥65 years admitted over two 1-month periods. Outcomes were recorded from medical records and discharge documentation, including length of inpatient stay, medical and surgical complications, and 30-day readmission and mortality rates. Phase 2 was a quality improvement project involving Plan-Do-Study-Act cycles and qualitative staff surveys in order to create a Geriatric Surgical Checklist (GSCL) to standardize the intervention in Phase 1, improve equity of care by extending it to all ages, improve team-working and streamline handovers for multidisciplinary staff. RESULTS: Phase 1 included 112 patients in the control month and 130 in the intervention month. The length of inpatient stay was reduced by 19% (mean 4.9 vs 4.0 days; P = 0.01), total postoperative complications were lower (risk ratio 0.24 [95% confidence interval 0.10, 0.54]; P = 0.001). A non-significant trend was seen towards fewer cancellations of surgery (10 vs 5%; P = 0.12) and 30-day readmissions (8 vs 3%; P = 0.07). In Phase 2, the GSCL was created and incrementally improved. Questionnaires repeated at intervals showed that the GSCL helped staff to understand their role better in multidisciplinary meetings, improved their confidence to raise issues, reduced duplication of handovers and standardized identification of geriatric issues. Equity of care was improved by providing the intervention to patients of all ages, despite which the time taken for the daily board round did not lengthen. CONCLUSION: This is the first known paper describing the benefits of daily proactive geriatric intervention in elective and emergency urological surgery. The results suggest that using a multidisciplinary team board round helps to facilitate collaborative working between surgical and geriatric medicine teams. The GSCL enables systematic identification of patients who require a focused comprehensive geriatric assessment. There is potential to transfer the GSCL package to other surgical specialties and hospitals to improve postoperative outcomes