Selective targeting of activating and inhibitory Smads by distinct WWP2 ubiquitin ligase isoforms differentially modulates TGFβ signalling and EMT

Ubiquitin-dependent mechanisms have emerged as essential regulatory elements controlling cellular levels of Smads and TGFß-dependent biological outputs such as epithelial–mesenchymal transition (EMT). In this study, we identify a HECT E3 ubiquitin ligase known as WWP2 (Full-length WWP2-FL), together with two WWP2 isoforms (N-terminal, WWP2-N; C-terminal WWP2-C), as novel Smad-binding partners. We show that WWP2-FL interacts exclusively with Smad2, Smad3 and Smad7 in the TGFß pathway. Interestingly, the WWP2-N isoform interacts with Smad2 and Smad3, whereas WWP2-C interacts only with Smad7. In addition, WWP2-FL and WWP2-C have a preference for Smad7 based on protein turnover and ubiquitination studies. Unexpectedly, we also find that WWP2-N, which lacks the HECT ubiquitin ligase domain, can also interact with WWP2-FL in a TGFß-regulated manner and activate endogenous WWP2 ubiquitin ligase activity causing degradation of unstimulated Smad2 and Smad3. Consistent with our protein interaction data, overexpression and knockdown approaches reveal that WWP2 isoforms differentially modulate TGFß-dependent transcription and EMT. Finally, we show that selective disruption of WWP2 interactions with inhibitory Smad7 can stabilise Smad7 protein levels and prevent TGFß-induced EMT. Collectively, our data suggest that WWP2-N can stimulate WWP2-FL leading to increased activity against unstimulated Smad2 and Smad3, and that Smad7 is a preferred substrate for WWP2-FL and WWP2-C following prolonged TGFß stimulation. Significantly, this is the first report of an interdependent biological role for distinct HECT E3 ubiquitin ligase isoforms, and highlights an entirely novel regulatory paradigm that selectively limits the level of inhibitory and activating Smads

Fosmidomycin Uptake into Plasmodium and Babesia-Infected Erythrocytes Is Facilitated by Parasite-Induced New Permeability Pathways

., a mouse malaria parasite. and related parasites. Our data provide further evidence that parasite-induced new permeability pathways may be exploited as routes for drug delivery

HIV incidence and prevalence among adults aged 15-64 years in Rwanda: Results from the Rwanda Population-based HIV Impact Assessment (RPHIA) and District-level Modeling, 2019

Objectives The 2018–19 Rwanda Population-based HIV Impact Assessment (RPHIA) was conducted to measure national HIV incidence and prevalence. District-level estimates were modeled to inform resources allocation. Methods RPHIA was a nationally representative cross-sectional household survey. Consenting adults were interviewed and tested for HIV using the national diagnostic algorithm followed by laboratory-based confirmation of HIV status, and testing for viral load (VL), limiting antigen (LAg) avidity and presence of antiretrovirals. Incidence was calculated using normalized optical density ≤ 1•5, VL ≥ 1,000 copies/mL, and undetectable antiretrovirals. Survey and programmatic data were used to model district-level HIV incidence and prevalence. Results Of 31,028 eligible adults, 98•7% participated in RPHIA and 934 tested HIV positive. HIV prevalence among adults in Rwanda was 3•0% (95% CI:2•7–3•3). National HIV incidence was 0•08% (95% CI:0•02–0•14) and 0•11% (95% CI:0•00–0•26) in the City of Kigali (CoK). Based on district-level modeling, HIV incidence was greatest in the three CoK districts (0•11% to 0•15%) and varied across other districts (0•03% to 0•10%). Conclusions HIV prevalence among adults in Rwanda is 3.0%; HIV incidence is low at 0.08%. District-level modeling has identified disproportionately affected urban hotspots: areas to focus resources

Primary Hyperparathyroidism Influences the Expression of Inflammatory and Metabolic Genes in Adipose Tissue

Background: Primary hyperparathyroidism (PHPT) is characterised by increased production of parathyroid hormone (PTH) resulting in elevated serum calcium levels. The influence on bone metabolism with altered bone resorption is the most studied clinical condition in PHPT. In addition to this, patients with PHPT are at increased risk of non-skeletal diseases, such as impaired insulin sensitivity, arterial hypertension and increased risk of death by cardiovascular diseases (CVD), possibly mediated by a chronic low-grade inflammation. The aim of this study was to investigate whether adipose tissue reflects the low-grade inflammation observed in PHPT patients. Methodology/Principal Findings: Subcutaneous fat tissue from the neck was sampled from 16 non-obese patients with PHPT and from 16 patients operated for benign thyroid diseases, serving as weight-matched controls. RNA was extracted and global gene expression was analysed with Illumina BeadArray Technology. We found 608 differentially expressed genes (q-value,0.05), of which 347 were up-regulated and 261 were down-regulated. Gene ontology analysis showed that PHPT patients expressed increased levels of genes involved in immunity and defense (e.g. matrix metallopeptidase 9, S100 calcium binding protein A8 and A9, CD14, folate receptor 2), and reduced levels of genes involved in metabolic processes. Analysis of transcription factor binding sites present in the differentially expressed genes corroborated the up-regulation of inflammatory processes. Conclusions/Significance: Our findings demonstrate that PHPT strongly influences gene regulation in fat tissue, which may result in altered adipose tissue function and release of pathogenic factors that increase the risk of CVD

Multilocus Sequence Typing as a Replacement for Serotyping in Salmonella enterica

Salmonella enterica subspecies enterica is traditionally subdivided into serovars by serological and nutritional characteristics. We used Multilocus Sequence Typing (MLST) to assign 4,257 isolates from 554 serovars to 1092 sequence types (STs). The majority of the isolates and many STs were grouped into 138 genetically closely related clusters called eBurstGroups (eBGs). Many eBGs correspond to a serovar, for example most Typhimurium are in eBG1 and most Enteritidis are in eBG4, but many eBGs contained more than one serovar. Furthermore, most serovars were polyphyletic and are distributed across multiple unrelated eBGs. Thus, serovar designations confounded genetically unrelated isolates and failed to recognize natural evolutionary groupings. An inability of serotyping to correctly group isolates was most apparent for Paratyphi B and its variant Java. Most Paratyphi B were included within a sub-cluster of STs belonging to eBG5, which also encompasses a separate sub-cluster of Java STs. However, diphasic Java variants were also found in two other eBGs and monophasic Java variants were in four other eBGs or STs, one of which is in subspecies salamae and a second of which includes isolates assigned to Enteritidis, Dublin and monophasic Paratyphi B. Similarly, Choleraesuis was found in eBG6 and is closely related to Paratyphi C, which is in eBG20. However, Choleraesuis var. Decatur consists of isolates from seven other, unrelated eBGs or STs. The serological assignment of these Decatur isolates to Choleraesuis likely reflects lateral gene transfer of flagellar genes between unrelated bacteria plus purifying selection. By confounding multiple evolutionary groups, serotyping can be misleading about the disease potential of S. enterica. Unlike serotyping, MLST recognizes evolutionary groupings and we recommend that Salmonella classification by serotyping should be replaced by MLST or its equivalents

A922 Sequential measurement of 1 hour creatinine clearance (1-CRCL) in critically ill patients at risk of acute kidney injury (AKI)

Meeting abstrac