Supporting data – Floating Gardens, Chicago River, April 29, 2018 to November 19, 2019

Between April 29, 2018 to November 19, 2019, water samples were collected upstream and downstream of a floating garden installed along the Chicago River. At each location, samples were collected at the surface and at 0.3 m below the surface. In-situ measurements of Dissolved Oxygen, Specific Conductance, and Temperature were recorded with a YSI 85. Anion samples were analyzed using a Ion Chromatograph for fluoride (F-), chloride (Cl-), nitrate as nitrogen (NO3-N), phosphate (PO4-3), and sulfate (SO4-2). The available dataset provides the recorded field parameters and the analyzed ion concentrations

The Effectiveness of an Artificial Floating Wetland to Remove Nutrients in an Urban Stream: A Pilot-Study in the Chicago River, Chicago, IL USA

Ever expanding urbanized landscapes are increasingly impacting streams that run through them. Among other stressors, urban streams often are host to elevated concentrations of nutrients, salts, and heavy metals. The pollutants, coupled with high temperatures, are drivers of ecosystem degradation in urban streams. The installation of artificial floating wetlands (AFWs) has been successful in mitigating the effects of urbanization in lakes and wastewater treatment ponds, but rarely have they been tested in streams. This pilot-study examined the ability of an AFW to improve water quality in an urban stream. The small, 90 m2 AFW was installed to improve the aquatic habitat and aesthetics of a small section of the Chicago River, Chicago, IL USA.Water samples and in-situ measurements were collected from the surface and at 0.3 m depth of upstream and downstream of the AFW. Samples were analyzed for nitrate-as-nitrogen, phosphate, chloride, and heavy metals. Comparison of upstream and downstream waters showed that the AFW lowered the concentrations of nitrate-as-nitrogen and phosphate during the growing season by 6.9% and 6.0%, respectively. Nitrate was also removed during the dormant season; however, phosphate was not removed during that time. Plant or microbial uptake of the nutrients are believed to be the dominant mechanisms in the growing season with denitrification serving as the primary pathway in the dormant season. Despite not having a measurable effect on the water temperature, the AFW was an effective means to reduce concentrations of nitrate and phosphorus, decreasing the potential for eutrophication

The interaction between the ER membrane protein UNC93B and TLR3, 7, and 9 is crucial for TLR signaling

Toll-like receptors (TLRs) sense the presence of microbial and viral pathogens by signal transduction mechanisms that remain to be fully elucidated. A single point mutation (H412R) in the polytopic endoplasmic reticulum (ER)–resident membrane protein UNC93B abolishes signaling via TLR3, 7, and 9. We show that UNC93B specifically interacts with TLR3, 7, 9, and 13, whereas introduction of the point mutation H412R in UNC93B abolishes their interactions. We establish the physical interaction of the intracellular TLRs with UNC93B in splenocytes and bone marrow–derived dendritic cells. Further, by expressing chimeric TLRs, we show that TLR3 and 9 bind to UNC93B via their transmembrane domains. We propose that a physical association between UNC93B and TLRs in the ER is essential for proper TLR signaling

The Folliculin Tumor Suppressor Is a GAP for the RagC/D GTPases That Signal Amino Acid Levels to mTORC1

The mTORC1 kinase is a master growth regulator that senses numerous environmental cues, including amino acids. The Rag GTPases interact with mTORC1 and signal amino acid sufficiency by promoting the translocation of mTORC1 to the lysosomal surface, its site of activation. The Rags are unusual GTPases in that they function as obligate heterodimers, which consist of RagA or B bound to RagC or D. While the loading of RagA/B with GTP initiates amino acid signaling to mTORC1, the role of RagC/D is unknown. Here, we show that RagC/D is a key regulator of the interaction of mTORC1 with the Rag heterodimer and that, unexpectedly, RagC/D must be GDP bound for the interaction to occur. We identify FLCN and its binding partners, FNIP1/2, as Rag-interacting proteins with GAP activity for RagC/D, but not RagA/B. Thus, we reveal a role for RagC/D in mTORC1 activation and a molecular function for the FLCN tumor suppressor.United States. National Institutes of Health (CA103866)United States. National Institutes of Health (AI47389)United States. Department of Defense (W81XWH-07-0448)National Cancer Institute (U.S.) (F30CA180754

Design and Evaluation of a New National Pharmacy Internship Programme in Ireland

Objective. To design, deliver and evaluate a National Pharmacy Internship Programme to meet the educational needs of pharmacy graduates for registration as competent pharmacists, recognising the learning by way of an educational award. Methods. The programme was designed as a twelve month, full-time, blended-learning, competency-based programme leading to an educational award at Master’s Level. Intern performance was assessed academically and by pharmacy tutor (preceptor) appraisals. Interns who demonstrated competency were invited to sit the Professional Registration Examination (PRE). Feasibility and performance were evaluated and a longitudinal approach allowed intern and tutor views to be compared to the former pre-registration year. Results. Overall performance in the PRE was good and relatively consistent with almost all interns proceeding to register as pharmacists. Interns expressed a view that the programme had enabled them to develop the knowledge, skills and overall competencies required for future independent practice as a pharmacist. Tutors considered the programme to have built on prior learning and provided a sufficiently rounded experience for professional practice. Tutors also expressed the view that the programme was an improved educational experience over the former, less structured, pre-registration training. Conclusion. The National Pharmacy Internship Programme was a structured, competency-based programme at Master’s Level. The education and training was perceived to be an improvement on the previous pre-registration year. The programme quality assured pharmacy education outcomes at entry-to-practice on a national basis, and uniquely recognised the learning by way of an educational award

Site-Specific Chemoenzymatic Labeling of Aerolysin Enables the Identification of New Aerolysin Receptors

Aerolysin is a secreted bacterial toxin that perforates the plasma membrane of a target cell with lethal consequences. Previously explored native and epitope-tagged forms of the toxin do not allow site-specific modification of the mature toxin with a probe of choice. We explore sortase-mediated transpeptidation reactions (sortagging) to install fluorophores and biotin at three distinct sites in aerolysin, without impairing binding of the toxin to the cell membrane and with minimal impact on toxicity. Using a version of aerolysin labeled with different fluorophores at two distinct sites we followed the fate of the C-terminal peptide independently from the N-terminal part of the toxin, and show its loss in the course of intoxication. Making use of the biotinylated version of aerolysin, we identify mesothelin, urokinase plasminogen activator surface receptor (uPAR, CD87), glypican-1, and CD59 glycoprotein as aerolysin receptors, all predicted or known to be modified with a glycosylphosphatidylinositol anchor. The sortase-mediated reactions reported here can be readily extended to other pore forming proteins.National Institutes of Health (U.S.) (grant R01 AI087879

Autoantibodies Produced at the Site of Tissue Damage Provide Evidence of Humoral Autoimmunity in Inclusion Body Myositis

Inclusion body myositis (IBM) belongs to a group of muscle diseases known as the inflammatory myopathies. The presence of antibody-secreting plasma cells in IBM muscle implicates the humoral immune response in this disease. However, whether the humoral immune response actively contributes to IBM pathology has not been established. We sought to investigate whether the humoral immune response in IBM both in the periphery and at the site of tissue damage was directed towards self-antigens. Peripheral autoantibodies present in IBM serum but not control serum recognized self-antigens in both muscle tissue and human-derived cell lines. To study the humoral immune response at the site of tissue damage in IBM patients, we isolated single plasma cells directly from IBM-derived muscle tissue sections and from these cells, reconstructed a series of recombinant immunoglobulins (rIgG). These rIgG, each representing a single muscle-associated plasma cell, were examined for reactivity to self-antigens. Both, flow cytometry and immunoblotting revealed that these rIgG recognized antigens expressed by cell lines and in muscle tissue homogenates. Using a mass spectrometry-based approach, Desmin, a major intermediate filament protein, expressed abundantly in muscle tissue, was identified as the target of one IBM muscle-derived rIgG. Collectively, these data support the view that IBM includes a humoral immune response in both the periphery and at the site of tissue damage that is directed towards self-antigens

Reviewer agreement trends from four years of electronic submissions of conference abstract

BACKGROUND: The purpose of this study was to determine the inter-rater agreement between reviewers on the quality of abstract submissions to an annual national scientific meeting (Canadian Association of Emergency Physicians; CAEP) to identify factors associated with low agreement. METHODS: All abstracts were submitted using an on-line system and assessed by three volunteer CAEP reviewers blinded to the abstracts' source. Reviewers used an on-line form specific for each type of study design to score abstracts based on nine criteria, each contributing from two to six points toward the total (maximum 24). The final score was determined to be the mean of the three reviewers' scores using Intraclass Correlation Coefficient (ICC). RESULTS: 495 Abstracts were received electronically during the four-year period, 2001 – 2004, increasing from 94 abstracts in 2001 to 165 in 2004. The mean score for submitted abstracts over the four years was 14.4 (95% CI: 14.1–14.6). While there was no significant difference between mean total scores over the four years (p = 0.23), the ICC increased from fair (0.36; 95% CI: 0.24–0.49) to moderate (0.59; 95% CI: 0.50–0.68). Reviewers agreed less on individual criteria than on the total score in general, and less on subjective than objective criteria. CONCLUSION: The correlation between reviewers' total scores suggests general recognition of "high quality" and "low quality" abstracts. Criteria based on the presence/absence of objective methodological parameters (i.e., blinding in a controlled clinical trial) resulted in higher inter-rater agreement than the more subjective and opinion-based criteria. In future abstract competitions, defining criteria more objectively so that reviewers can base their responses on empirical evidence may lead to increased consistency of scoring and, presumably, increased fairness to submitters

Formation of mammalian preribosomes proceeds from intermediate to composed state during ribosome maturation

http://www.jbc.org/content/early/2019/05/10/jbc.AC119.008378.full.pdfIn eukaryotes, ribosome assembly is a rate-limiting step in ribosomal biogenesis that takes place in a distinctive subnuclear organelle, the nucleolus. How ribosomes get assembled at the nucleolar site by forming initial preribosomal complexes remains poorly characterized. In this study, using several human and murine cell lines, we developed a method for isolation of native mammalian preribosomal complexes by lysing cell nuclei through mild sonication. A sucrose gradient fractionation of the nuclear lysate resolved several ribonucleoprotein (RNP) complexes containing rRNAs and ribosomal proteins. Characterization of the RNP complexes with MS-based protein identification and Northern blotting-based rRNA detection approaches identified two types of preribosomes we named here as intermediate preribosomes (IPRibs) and composed preribosome (CPRib). IPRib complexes comprised large preribosomes (105S to 125S in size) containing the rRNA modification factors and premature rRNAs. We further observed that a distinctive CPRib complex consists of an 85S preribosome assembled with mature rRNAs and a ribosomal biogenesis factor, Ly1 antibody-reactive (LYAR), that does not associate with premature rRNAs and rRNA modification factors. rRNA-labeling experiments uncovered that IPRib assembly precedes CPRib complex formation. We also found that formation of the preribosomal complexes is nutrient-dependent because the abundances of IPRib and CPRib decreased substantially when cells were either deprived of amino acids or exposed to an mTOR kinase inhibitor. These findings indicate that preribosomes form via dynamic and nutrient-dependent processing events and progress from an intermediate to a composed state during ribosome maturatio

Genome-Wide Association Study Identifies GPC5 as a Novel Genetic Locus Protective against Sudden Cardiac Arrest

BACKGROUND:Existing studies indicate a significant genetic component for sudden cardiac arrest (SCA) and genome-wide association studies (GWAS) provide an unbiased approach for identification of novel genes. We performed a GWAS to identify genetic determinants of SCA. METHODOLOGY/PRINCIPAL FINDINGS:We used a case-control design within the ongoing Oregon Sudden Unexpected Death Study (Oregon-SUDS). Cases (n = 424) were SCAs with coronary artery disease (CAD) among residents of Portland, OR (2002-07, population approximately 1,000,000) and controls (n = 226) were residents with CAD, but no history of SCA. All subjects were of White-European ancestry and GWAS was performed using Affymetrix 500K/5.0 and 6.0 arrays. High signal markers were genotyped in SCA cases (n = 521) identified from the Atherosclerosis Risk in Communities Study (ARIC) and the Cardiovascular Health Study (CHS) (combined n = 19,611). No SNPs reached genome-wide significance (p<5x10(-8)). SNPs at 6 loci were prioritized for follow-up primarily based on significance of p<10(-4) and proximity to a known gene (CSMD2, GPR37L1, LIN9, B4GALNT3, GPC5, and ZNF592). The minor allele of GPC5 (GLYPICAN 5, rs3864180) was associated with a lower risk of SCA in Oregon-SUDS, an effect that was also observed in ARIC/CHS whites (p<0.05) and blacks (p<0.04). In a combined Cox proportional hazards model analysis that adjusted for race, the minor allele exhibited a hazard ratio of 0.85 (95% CI 0.74 to 0.98; p<0.01). CONCLUSIONS/SIGNIFICANCE:A novel genetic locus for SCA, GPC5, was identified from Oregon-SUDS and successfully validated in the ARIC and CHS cohorts. Three other members of the Glypican family have been previously implicated in human disease, including cardiac conditions. The mechanism of this specific association requires further study