Signal peptide peptidase (SPP) dimer formation as assessed by fluorescence lifetime imaging microscopy (FLIM) in intact cells

BACKGROUND: Signal peptide peptidase (SPP) is an intramembrane cleaving protease identified by its cleavage of several type II membrane signal peptides. Conservation of intramembrane active site residues demonstrates that SPP, SPP family members, and presenilins (PSs) make up a family of intramembrane cleaving proteases. Because SPP appears to function without additional protein cofactors, the study of SPP may provide structural insights into the mechanism of intramembrane proteolysis by this biomedically important family of proteins. Previous studies have shown that SPP isolated from cells appears to be a homodimer, but some evidence exists that in vitro SPP may be active as a monomer. We have conducted additional experiments to determine if SPP exists as a monomer or dimer in vivo. RESULTS: Fluorescence lifetime imaging microscopy (FLIM) can be is used to determine intra- or intermolecular interactions by fluorescently labeling epitopes on one or two different molecules. If the donor and acceptor fluorophores are less than 10 nm apart, the donor fluorophore lifetime shortens proportionally to the distance between the fluorophores. In this study, we used two types of fluorescence energy transfer (FRET) pairs; cyan fluorescent protein (CFP) with yellow fluorescent protein (YFP) or Alexa 488 with Cy3 to differentially label the NH2- or COOH-termini of SPP molecules. A cell based SPP activity assay was used to show that all tagged SPP proteins are proteolytically active. Using FLIM we were able to show that the donor fluorophore lifetime of the CFP tagged SPP construct in living cells significantly decreases when either a NH2- or COOH-terminally YFP tagged SPP construct is co-transfected, indicating close proximity between two different SPP molecules. These data were then confirmed in cell lines stably co-expressing V5- and FLAG-tagged SPP constructs. CONCLUSION: Our FLIM data strongly suggest dimer formation between two separate SPP proteins. Although the tagged SPP constructs are expressed throughout the cell, SPP dimer detection by FLIM is seen predominantly at or near the plasma membrane

Scientists Want More Children

Scholars partly attribute the low number of women in academic science to the impact of the science career on family life. Yet, the picture of how men and women in science – at different points in the career trajectory – compare in their perceptions of this impact is incomplete. In particular, we know little about the perceptions and experiences of junior and senior scientists at top universities, institutions that have a disproportionate influence on science, science policy, and the next generation of scientists. Here we show that having fewer children than wished as a result of the science career affects the life satisfaction of science faculty and indirectly affects career satisfaction, and that young scientists (graduate students and postdoctoral fellows) who have had fewer children than wished are more likely to plan to exit science entirely. We also show that the impact of science on family life is not just a woman's problem; the effect on life satisfaction of having fewer children than desired is more pronounced for male than female faculty, with life satisfaction strongly related to career satisfaction. And, in contrast to other research, gender differences among graduate students and postdoctoral fellows disappear. Family factors impede talented young scientists of both sexes from persisting to research positions in academic science. In an era when the global competitiveness of US science is at risk, it is concerning that a significant proportion of men and women trained in the select few spots available at top US research universities are considering leaving science and that such desires to leave are related to the impact of the science career on family life. Results from our study may inform university family leave policies for science departments as well as mentoring programs in the sciences

A global perspective of advanced practice nursing research:a review of systematic reviews protocol

INTRODUCTION: In 2020, the World Health Organization called for the expansion and greater recognition of all nursing roles, including advanced practice nurses (APNs), to better meet patient care needs. As defined by the International Council of Nurses (ICN), the two most common APN roles include nurse practitioners (NPs) and clinical nurse specialists (CNSs). They help ensure care to communities as well as patients and families with acute, chronic or complex conditions. Moreover, APNs support providers to deliver high quality care and improve access to services. Currently, there is much variability in the use of advanced practice nursing roles globally. A clearer understanding of the roles that are in place across the globe, and how they are being used will support greater role harmonization, and inform global priorities for advanced practice nursing education, research, and policy reform. OBJECTIVE: To identify current gaps in advanced practice nursing research globally. MATERIALS AND METHODS: This review of systematic reviews will provide a description of the current state of the research, including gaps, on advanced practice nursing globally. We will include reviews that examine APNs, NPs or CNSs using recognized role definitions. We will search the CINAHL, EMBASE, Global Health, HealthStar, PubMed, Medline, Cochrane Library Database of Systematic Reviews and Controlled Trials Register, Database of Abstracts of Reviews of Effects, Joanna Briggs Institute, and Web of Science electronic databases for reviews published from January 2011 onwards, with no restrictions on jurisdiction or language. We will search the grey literature and hand search the reference lists of all relevant reviews to identify additional studies. We will extract country, patient, provider, health system, educational, and policy/scope of practice data. We will assess the quality of each included review using the CASP criteria, and summarize their findings. This review of systematic reviews protocol was developed following the PRISMA-P recommendations. PROSPERO REGISTRATION NUMBER: CRD42021278532

Closeup: Hawaii\u27s Women\u27s Studies Program

As the University of Hawaii seeks to cut its budget and its position count, Women\u27s Studies at the Manoa campus can only hope that its modest requirements, low profile, absence of an administrative head, and experience in developing within the most precarious of situations may allow it to survive. None of the faculty have tenure; only two out of eight have career ladder positions; three of our faculty will not know until late summer whether they have even part-time jobs; one has been fired; and the rest have job security for one more year

Cadmium, Smoking, and Human Blood DNA Methylation Profiles in Adults from the Strong Heart Study.

The epigenetic effects of individual environmental toxicants in tobacco remain largely unexplored. Cadmium (Cd) has been associated with smoking-related health effects, and its concentration in tobacco smoke is higher in comparison with other metals. We studied the association of Cd and smoking exposures with human blood DNA methylation (DNAm) profiles. We also evaluated the implication of findings to relevant methylation pathways and the potential contribution of Cd exposure from smoking to explain the association between smoking and site-specific DNAm. We conducted an epigenome-wide association study of urine Cd and self-reported smoking (current and former vs. never, and cumulative smoking dose) with blood DNAm in 790,026 CpGs (methylation sites) measured with the Illumina Infinium Human MethylationEPIC (Illumina Inc.) platform in 2,325 adults 45-74 years of age who participated in the Strong Heart Study in 1989-1991. In a mediation analysis, we estimated the amount of change in DNAm associated with smoking that can be independently attributed to increases in urine Cd concentrations from smoking. We also conducted enrichment analyses and in silico protein-protein interaction networks to explore the biological relevance of the findings. At a false discovery rate (FDR)-corrected level of 0.05, we found 6 differentially methylated positions (DMPs) for Cd; 288 and 17, respectively, for current and former smoking status; and 77 for cigarette pack-years. Enrichment analyses of these DMPs displayed enrichment of 58 and 6 Gene Ontology and Kyoto Encyclopedia of Genes and Genomes gene sets, respectively, including biological pathways for cancer and cardiovascular disease. In in silico protein-to-protein networks, we observed key proteins in DNAm pathways directly and indirectly connected to Cd- and smoking-DMPs. Among DMPs that were significant for both Cd and current smoking (annotated to PRSS23, AHRR, F2RL3, RARA, and 2q37.1), we found statistically significant contributions of Cd to smoking-related DNAm. Beyond replicating well-known smoking epigenetic signatures, we found novel DMPs related to smoking. Moreover, increases in smoking-related Cd exposure were associated with differential DNAm. Our integrative analysis supports a biological link for Cd and smoking-associated health effects, including the possibility that Cd is partly responsible for smoking toxicity through epigenetic changes. https://doi.org/10.1289/EHP6345.This work was supported by grants by the National Heart, Lung, and Blood Institute (NHLBI) (under contract numbers 75N92019D00027, 75N92019D00028, 75N92019D00029, & 75N92019D00030) and previous grants (R01HL090863, R01HL109315, R01HL109301, R01HL109284, R01HL109282, and R01HL109319 and cooperative agreements U01HL41642, U01HL41652, U01HL41654, U01HL65520, and U01HL65521), by the National Institute of Health Sciences (R01ES021367, R01ES025216, P42ES010349, P30ES009089), by the Spanish Funds for Research In Health Sciences, Carlos III Health Institute, co-funded by European Regional Development Fund (CP12/03080 and PI15/00071), by Chilean CONICYT/FONDECYT-POSTDOCTORADO Nº3180486 (A.L.R.-C) and a fellowship from “La Caixa” Foundation (ID 100010434). The fellowship code is “LCF/BQ/DR19/11740016.”S

Kelp carbon sink potential decreases with warming due to accelerating decomposition

Cycling of organic carbon in the ocean has the potential to mitigate or exacerbate global climate change, but major questions remain about the environmental controls on organic carbon flux in the coastal zone. Here, we used a field experiment distributed across 28° of latitude, and the entire range of 2 dominant kelp species in the northern hemisphere, to measure decomposition rates of kelp detritus on the seafloor in relation to local environmental factors. Detritus decomposition in both species were strongly related to ocean temperature and initial carbon content, with higher rates of biomass loss at lower latitudes with warmer temperatures. Our experiment showed slow overall decomposition and turnover of kelp detritus and modeling of coastal residence times at our study sites revealed that a significant portion of this production can remain intact long enough to reach deep marine sinks. The results suggest that decomposition of these kelp species could accelerate with ocean warming and that low-latitude kelp forests could experience the greatest increase in remineralization with a 9% to 42% reduced potential for transport to long-term ocean sinks under short-term (RCP4.5) and long-term (RCP8.5) warming scenarios. However, slow decomposition at high latitudes, where kelp abundance is predicted to expand, indicates potential for increasing kelp-carbon sinks in cooler (northern) regions. Our findings reveal an important latitudinal gradient in coastal ecosystem function that provides an improved capacity to predict the implications of ocean warming on carbon cycling. Broad-scale patterns in organic carbon decomposition revealed here can be used to identify hotspots of carbon sequestration potential and resolve relationships between carbon cycling processes and ocean climate at a global scale.publishedVersio

Genome-wide association study identifies peanut allergy-specific loci and evidence of epigenetic mediation in US children

Food allergy (FA) affects 2%-10% of US children and is a growing clinical and public health problem. Here we conduct the first genome-wide association study of well-defined FA, including specific subtypes (peanut, milk and egg) in 2,759 US participants (1,315 children and 1,444 parents) from the Chicago Food Allergy Study, and identify peanut allergy (PA)-specific loci in the HLA-DR and -DQ gene region at 6p21.32, tagged by rs7192 (P=5.5 × 10 -8) and rs9275596 (P=6.8 × 10 -10), in 2,197 participants of European ancestry. We replicate these associations in an independent sample of European ancestry. These associations are further supported by meta-analyses across the discovery and replication samples. Both single-nucleotide polymorphisms (SNPs) are associated with differential DNA methylation levels at multiple CpG sites (

A case of autism with an interstitial deletion on 4q leading to hemizygosity for genes encoding for glutamine and glycine neurotransmitter receptor sub-units (AMPA 2, GLRA3, GLRB) and neuropeptide receptors NPY1R, NPY5R

BACKGROUND: Autism is a pervasive developmental disorder characterized by a triad of deficits: qualitative impairments in social interactions, communication deficits, and repetitive and stereotyped patterns of behavior. Although autism is etiologically heterogeneous, family and twin studies have established a definite genetic basis. The inheritance of idiopathic autism is presumed to be complex, with many genes involved; environmental factors are also possibly contributory. The analysis of chromosome abnormalities associated with autism contributes greatly to the identification of autism candidate genes. CASE PRESENTATION: We describe a child with autistic disorder and an interstitial deletion on chromosome 4q. This child first presented at 12 months of age with developmental delay and minor dysmorphic features. At 4 years of age a diagnosis of Pervasive Developmental Disorder was made. At 11 years of age he met diagnostic criteria for autism. Cytogenetic studies revealed a chromosome 4q deletion. The karyotype was 46, XY del 4 (q31.3-q33). Here we report the clinical phenotype of the child and the molecular characterization of the deletion using molecular cytogenetic techniques and analysis of polymorphic markers. These studies revealed a 19 megabase deletion spanning 4q32 to 4q34. Analysis of existing polymorphic markers and new markers developed in this study revealed that the deletion arose on a paternally derived chromosome. To date 33 genes of known or inferred function are deleted as a consequence of the deletion. Among these are the AMPA 2 gene that encodes the glutamate receptor GluR2 sub-unit, GLRA3 and GLRB genes that encode glycine receptor subunits and neuropeptide Y receptor genes NPY1R and NPY5R. CONCLUSIONS: The deletion in this autistic subject serves to highlight specific autism candidate genes. He is hemizygous for AMPA 2, GLRA3, GLRB, NPY1R and NPY5R. GluR2 is the major determinant of AMPA receptor structure. Glutamate receptors maintain structural and functional plasticity of synapses. Neuropeptide Y and its receptors NPY1R and NPY5R play a role in hippocampal learning and memory. Glycine receptors are expressed in very early cortical development. Molecular cytogenetic studies and DNA sequence analysis in other patients with autism will be necessary to confirm that these genes are involved in autism