G-CSFR Ubiquitination Critically Regulates Myeloid Cell Survival and Proliferation

The granulocyte colony-stimulating factor receptor (G-CSFR) is a critical regulator of granulopoiesis. Mutations in the G-CSFR in patients with severe congenital neutropenia (SCN) transforming to acute myelogenous leukemia (AML) have been shown to induce hypersensitivity and enhanced growth responses to G-CSF. Recent studies have demonstrated the importance of the ubiquitin/proteasome system in the initiation of negative signaling by the G-CSFR. To further investigate the role of ubiquitination in regulating G-CSFR signaling, we generated a mutant form of the G-CSFR (K762R/G-CSFR) which abrogates the attachment of ubiquitin to the lysine residue at position 762 of the G-CSFR that is deleted in the Δ716 G-CSFR form isolated from patients with SCN/AML. In response to G-CSF, mono-/polyubiquitination of the G-CSFR was impaired in cells expressing the mutant K762R/G-CSFR compared to cells transfected with the WT G-CSFR. Cells stably transfected with the K762R/G-CSFR displayed a higher proliferation rate, increased sensitivity to G-CSF, and enhanced survival following cytokine depletion, similar to previously published data with the Δ716 G-CSFR mutant. Activation of the signaling molecules Stat5 and Akt were also increased in K762R/G-CSFR transfected cells in response to G-CSF, and their activation remained prolonged after G-CSF withdrawal. These results indicate that ubiquitination is required for regulation of G-CSFR-mediated proliferation and cell survival. Mutations that disrupt G-CSFR ubiquitination at lysine 762 induce aberrant receptor signaling and hyperproliferative responses to G-CSF, which may contribute to leukemic transformation

Divergent Pathways in COS-7 Cells Mediate Defective Internalization and Intracellular Routing of Truncated G-CSFR Forms in SCN/AML

Expression of truncated G-CSFR forms in patients with SCN/AML induces hyperproliferation and prolonged cell survival. Previously, we showed that ligand internalization is delayed and degradation of truncated G-CSFR forms is defective in patients with SCN/AML.In this study, we investigated the potential roles of dileucine and tyrosine-based motifs within the cytoplasmic domain of the G-CSFR in modulating ligand/receptor internalization. Using standard binding assays with radiolabeled ligand and COS-7 cells, substitutions in the dileucine motif or deletion of tyrosine residues in the G-CSFR did not alter internalization. Attachment of the transferrin receptor YTRF internalization motif to a truncated G-CSFR form from a patient with SCN/AML corrected defective internalization, but not receptor degradation suggesting that receptor internalization and degradation occur independently via distinct domains and/or processes.Our data suggest that distinct domains within the G-CSFR mediate separate processes for receptor internalization and degradation. Our findings using standard binding assays differ from recently published data utilizing flow cytometry

Risk factor screening to identify women requiring oral glucose tolerance testing to diagnose gestational diabetes : a systematic review and meta-analysis and analysis of two pregnancy cohorts

BACKGROUND: Easily identifiable risk factors including: obesity and ethnicity at high risk of diabetes are commonly used to indicate which women should be offered the oral glucose tolerance test (OGTT) to diagnose gestational diabetes (GDM). Evidence regarding these risk factors is limited however. We conducted a systematic review (SR) and meta-analysis and individual participant data (IPD) analysis to evaluate the performance of risk factors in identifying women with GDM. METHODS: We searched MEDLINE, Medline in Process, Embase, Maternity and Infant Care and the Cochrane Central Register of Controlled Trials (CENTRAL) up to August 2016 and conducted additional reference checking. We included observational, cohort, case-control and cross-sectional studies reporting the performance characteristics of risk factors used to identify women at high risk of GDM. We had access to IPD from the Born in Bradford and Atlantic Diabetes in Pregnancy cohorts, all pregnant women in the two cohorts with data on risk factors and OGTT results were included. RESULTS: Twenty nine published studies with 211,698 women for the SR and a further 14,103 women from two birth cohorts (Born in Bradford and the Atlantic Diabetes in Pregnancy study) for the IPD analysis were included. Six studies assessed the screening performance of guidelines; six examined combinations of risk factors; eight evaluated the number of risk factors and nine examined prediction models or scores. Meta-analysis using data from published studies suggests that irrespective of the method used, risk factors do not identify women with GDM well. Using IPD and combining risk factors to produce the highest sensitivities, results in low specificities (and so higher false positives). Strategies that use the risk factors of age (>25 or >30) and BMI (>25 or 30) perform as well as other strategies with additional risk factors included. CONCLUSIONS: Risk factor screening methods are poor predictors of which pregnant women will be diagnosed with GDM. A simple approach of offering an OGTT to women 25 years or older and/or with a BMI of 25kg/m2 or more is as good as more complex risk prediction models. Research to identify more accurate (bio)markers is needed. Systematic Review Registration: PROSPERO CRD42013004608

Yeast : the soul of beer’s aroma—a review of flavour-active esters and higher alcohols produced by the brewing yeast

Among the most important factors influencing beer quality is the presence of well-adjusted amounts of higher alcohols and esters. Thus, a heavy body of literature focuses on these substances and on the parameters influencing their production by the brewing yeast. Additionally, the complex metabolic pathways involved in their synthesis require special attention. More than a century of data, mainly in genetic and proteomic fields, has built up enough information to describe in detail each step in the pathway for the synthesis of higher alcohols and their esters, but there is still place for more. Higher alcohols are formed either by anabolism or catabolism (Ehrlich pathway) of amino acids. Esters are formed by enzymatic condensation of organic acids and alcohols. The current paper reviews the up-to-date knowledge in the pathways involving the synthesis of higher alcohols and esters by brewing yeasts. Fermentation parameters affecting yeast response during biosynthesis of these aromatic substances are also fully reviewed.Eduardo Pires gratefully acknowledges the Fundacao para a Ciencia e a Tecnologia (FCT, Portugal) for the PhD fellowship support (SFRH/BD/61777/2009). The financial contributions of the EU FP7 project Ecoefficient Biodegradable Composite Advanced Packaging (EcoBioCAP, grant agreement no. 265669) as well as of the Grant Agency of the Czech Republic (project GACR P503/12/1424) are also gratefully acknowledged. The authors thank the Ministry of Education, Youth and Sports of the Czech Republic (MSM 6046137305) for their financial support

Neutrophil Extracellular Traps in Breast Cancer and Beyond: Current Perspectives on NET Stimuli, Thrombosis and Metastasis, and Clinical Utility for Diagnosis and Treatment

Abstract The formation of neutrophil extracellular traps (NETs), known as NETosis, was first observed as a novel immune response to bacterial infection, but has since been found to occur abnormally in a variety of other inflammatory disease states including cancer. Breast cancer is the most commonly diagnosed malignancy in women. In breast cancer, NETosis has been linked to increased disease progression, metastasis, and complications such as venous thromboembolism. NET-targeted therapies have shown success in preclinical cancer models and may prove valuable clinical targets in slowing or halting tumor progression in breast cancer patients. We will briefly outline the mechanisms by which NETs may form in the tumor microenvironment and circulation, including the crosstalk between neutrophils, tumor cells, endothelial cells, and platelets as well as the role of cancer-associated extracellular vesicles in modulating neutrophil behavior and NET extrusion. The prognostic implications of cancer-associated NETosis will be explored in addition to development of novel therapeutics aimed at targeting NET interactions to improve outcomes in patients with breast cancer

Perinatal asphyxia: current status and approaches towards neuroprotective strategies, with focus on sentinel proteins

Delivery is a stressful and risky event menacing the newborn. The mother-dependent respiration has to be replaced by autonomous pulmonary breathing immediately after delivery. If delayed, it may lead to deficient oxygen supply compromising survival and development of the central nervous system. Lack of oxygen availability gives rise to depletion of NAD+ tissue stores, decrease of ATP formation, weakening of the electron transport pump and anaerobic metabolism and acidosis, leading necessarily to death if oxygenation is not promptly re-established. Re-oxygenation triggers a cascade of compensatory biochemical events to restore function, which may be accompanied by improper homeostasis and oxidative stress. Consequences may be incomplete recovery, or excess reactions that worsen the biological outcome by disturbed metabolism and/or imbalance produced by over-expression of alternative metabolic pathways. Perinatal asphyxia has been associated with severe neurological and psychiatric sequelae with delayed clinical onset. No specific treatments have yet been established. In the clinical setting, after resuscitation of an infant with birth asphyxia, the emphasis is on supportive therapy. Several interventions have been proposed to attenuate secondary neuronal injuries elicited by asphyxia, including hypothermia. Although promising, the clinical efficacy of hypothermia has not been fully demonstrated. It is evident that new approaches are warranted. The purpose of this review is to discuss the concept of sentinel proteins as targets for neuroprotection. Several sentinel proteins have been described to protect the integrity of the genome (e.g. PARP-1; XRCC1; DNA ligase IIIα; DNA polymerase β, ERCC2, DNA-dependent protein kinases). They act by eliciting metabolic cascades leading to (i) activation of cell survival and neurotrophic pathways; (ii) early and delayed programmed cell death, and (iii) promotion of cell proliferation, differentiation, neuritogenesis and synaptogenesis. It is proposed that sentinel proteins can be used as markers for characterising long-term effects of perinatal asphyxia, and as targets for novel therapeutic development and innovative strategies for neonatal care

Ca isotope fingerprints of early crust-mantle evolution

Among the most important factors influencing beer quality is the presence of well-adjusted amounts of higher alcohols and esters; as well as the successful reduction of undesirable by-products such as diacetyl. While higher alcohols and esters contribute rather positively to the beer aroma, diacetyl is mostly unwelcome for beer types with lighter taste. Thus, the complex metabolic pathways in yeast responsible for the synthesis of both pleasant and unpleasant by-products of fermentation were given special attention in this last chapter