A review of COFRADIC techniques targeting protein N-terminal acetylation

Acetylation of nascent protein Nalpha-termini is a common modification among archae and eukaryotes and can influence the structure and function of target proteins. This modification has been studied on an individual protein or (synthetic) peptide level or on a proteome scale using two-dimensional polyacrylamide gel electrophoresis. We recently developed mass spectrometry driven proteome analytical approaches specifically targeting the amino (N) terminus of proteins based on the concept of diagonal reverse-phase chromatography. We here review how this so-called combined fractional diagonal chromatography (COFRADIC) technique can be used in combination with differential mass-tagging strategies as to both qualitatively and quantitatively assess protein Nalpha-acetylation in whole proteomes

N-glycosylation site analysis reveals sex-related differences in protein N-glycosylation in the rice brown planthopper (Nilaparvata lugens)

Glycosylation is a common modification of proteins and critical for a wide range of biological processes. Differences in protein glycosylation between sexes have already been observed in humans, nematodes and trematodes, and have recently also been reported in the rice pest insect Nilaparvata lugens. Although protein N-glycosylation in insects is nowadays of high interest because of its potential for exploitation in pest control strategies, the functionality of differential N-glycosylation between sexes is yet unknown. In this study, therefore, the occurrence and role of sex-related protein N-glycosylation in insects were examined. A comprehensive investigation of the N-glycosylation sites from the adult stages of N. lugens was conducted, allowing a qualitative and quantitative comparison between sexes at the glycopeptide level. N-glycopeptide enrichment via lectin capturing using the high mannose/paucimannose-binding lectin Concanavalin A, or the Rhizoctonia solani agglutinin which interacts with complex N-glycans, resulted in the identification of over 1300 N-glycosylation sites derived from over 600 glycoproteins. Comparison of these N-glycopeptides revealed striking differences in protein N-glycosylation between sexes. Male- and female-specific N-glycosylation sites were identified, and some of these sex-specific N-glycosylation sites were shown to be derived from proteins with a putative role in insect reproduction. In addition, differential glycan composition between males and females was observed for proteins shared across sexes. Both lectin blotting experiments as well as transcript expression analyses with complete insects and insect tissues confirmed the observed differences in N-glycosylation of proteins between sexes. In conclusion, this study provides further evidence for protein N-glycosylation to be sex-related in insects. Furthermore, original data on N-glycosylation sites of N. lugens adults are presented, providing novel insights into planthopper's biology and information for future biological pest control strategies

Extracellular vesicles from follicular and ampullary fluid isolated by density gradient ultracentrifugation improve bovine embryo development and quality

Extracellular vesicles (EVs) have been isolated from follicular (FF) and ampullary oviduct fluid (AOF), using different isolation methods. However, it is not clear whether different purification methods can affect the functionality of resulting EVs. Here, we compared two methods (OptiPrep (TM) density gradient ultracentrifugation (ODG UC) and single-step size exclusion chromatography (SEC) (qEV IZON (TM) single column)) for the isolation of EVs from bovine FF and AOF. Additionally, we evaluated whether the addition of EVs derived either by ODG UC or SEC from FF or AOF during oocyte maturation would yield extra benefits for embryo developmental competence. The characterization of EVs isolated using ODG UC or SEC from FF and AOF did not show any differences in terms of EV sizes (40-400 nm) and concentrations (2.4 +/- 0.2 x 10(12)-1.8 +/- 0.2 x 10(13) particles/mL). Blastocyst yield and quality was higher in groups supplemented with EVs isolated from FF and AOF by ODG UC, with higher total cell numbers and a lower apoptotic cell ratio compared with the other groups (p < 0.05). Supplementing in vitro maturation media with EVs derived by ODG UC from AOF was beneficial for bovine embryo development and quality

The Rwandan agrarian and land sector modernisation:confronting macro performance with lived experiences on the ground

Rwanda has embarked on an ambitious policy package to modernise and professionalise the agrarian and land sector. Its reform fits into a broader call – supported by major international donors – to implement a Green Revolution in Sub-Saharan Africa. After 10 years of implementation, there is increased production output and value-addition in commercialised commodity chains. These are promising results. However, poverty reduction, particularly in more recent years, seems limited. Moreover, micro-level evidence from the field calls into question the long-term sustainability of the agricultural and land sector reform. In this article, a group of researchers, having engaged in in-depth qualitative research in a variety of settings and over an extended period, bring together their main research results and combine their key findings to challenge the dominant discourse on Rwanda as a model for development

Whole-genome sequencing reveals a coding non-pathogenic variant tagging a non-coding pathogenic hexanucleotide repeat expansion in C9orf72 as cause of amyotrophic lateral sclerosis

Motor neuron degeneration in amyotrophic lateral sclerosis (ALS) has a familial cause in 10% of patients. Despite significant advances in the genetics of the disease, many families remain unexplained. We performed whole-genome sequencing in five family members from a pedigree with autosomal-dominant classical ALS. A family-based elimination approach was used to identify novel coding variants segregating with the disease. This list of variants was effectively shortened by genotyping these variants in 2 additional unaffected family members and 1500 unrelated population-specific controls. A novel rare coding variant in SPAG8 on chromosome 9p13.3 segregated with the disease and was not observed in controls. Mutations in SPAG8 were not encountered in 34 other unexplained ALS pedigrees, including 1 with linkage to chromosome 9p13.2–23.3. The shared haplotype containing the SPAG8 variant in this small pedigree was 22.7 Mb and overlapped with the core 9p21 linkage locus for ALS and frontotemporal dementia. Based on differences in coverage depth of known variable tandem repeat regions between affected and non-affected family members, the shared haplotype was found to contain an expanded hexanucleotide (GGGGCC)n repeat in C9orf72 in the affected members. Our results demonstrate that rare coding variants identified by whole-genome sequencing can tag a shared haplotype containing a non-coding pathogenic mutation and that changes in coverage depth can be used to reveal tandem repeat expansions. It also confirms (GGGGCC)n repeat expansions in C9orf72 as a cause of familial ALS

Family physician perceptions of climate change, migration, health, and healthcare in Sub-Saharan Africa : an exploratory study

Although family physicians (FPs) are community-oriented primary care generalists and should be the entry point for the populationâs interaction with the health system, they are underrepresented in research on the climate change, migration, and health(care) nexus (hereafter referred to as the nexus). Similarly, FPs can provide valuable insights into building capacity through integrating health-determining sectors for climate-resilient and migration-inclusive health systems, especially in Sub-Saharan Africa (SSA). Here, we explore FPsâ perceptions on the nexus in SSA and on intersectoral capacity building. Three focus groups conducted during the 2019 WONCA-Africa conference in Uganda were transcribed verbatim and analyzed using an inductive thematic approach. Participantsâ perceived interactions related to (1) migration and climate change, (2) migration for better health and healthcare, (3) health impacts of climate change and the role of healthcare, and (4) health impacts of migration and the role of healthcare were studied. We coined these complex and reinforcing interactions as continuous feedback loops intertwined with socio-economic, institutional, and demographic context. Participants identified five intersectoral capacity-building opportunities on micro, meso, macro, and supra (international) levels: multi-dimensional and multi-layered governance structures; improving FP training and primary healthcare working conditions; health advocacy in primary healthcare; collaboration between the health sector and civil society; and more responsibilities for high-income countries. This exploratory study presents a unique and novel perspective on the nexus in SSA which contributes to interdisciplinary research agendas and FP policy responses on national, regional, and global levels