24 research outputs found
Two Cascade Reactions with Oleate Hydratases for the Sustainable Biosynthesis of Fatty Acid-Derived Fine Chemicals
Oleate hydratases (OHs) are of significant industrial interest for the sustainable generation of valuable fine chemicals. When combined with other enzymes in multi-step cascades, the direct formation of fatty acid congeners can be accomplished with minimal processing steps. In this study, two cascade reactions are presented, which can be applied in one-pot approaches. The first cascade was placed “upstream” of an OH derived from Rhodococcus erythropolis (OhyRe), where a lipase from Candida rugosa was applied to hydrolyze triglycerides into free fatty acids, a crucial step for OH conversion. Further, we tested the lipase–OhyRe cascade with various types of renewable triglycerides of plant and microbial origin. In this context, the most efficient conversion was observed for microbial oil from Cutaneotrichosporon oleaginosus leading the way toward its industrial application. In contrast, the second cascade was placed “downstream” of OhyRe, where a novel secondary alcohol dehydrogenase (secADH) was applied to oxidize the hydroxylated fatty acid into a fatty acid ketone. Optimal reaction parameters for the cascade with the secADH were established, which allows this to be applied to high-throughput screens. Moreover, we describe a light-dependent route, thereby extending the catalytic efficiency of the OH enzyme system
Neuroligin-mediated neurodevelopmental defects are induced by mitochondrial dysfunction and prevented by lutein in C. elegans
Complex-I-deficiency represents the most frequent pathogenetic cause of human mitochondriopathies. Therapeutic options for these neurodevelopmental life-threating disorders do not exist, partly due to the scarcity of appropriate model systems to study them. Caenorhabditis elegans is a genetically tractable model organism widely used to investigate neuronal pathologies. Here, we generate C. elegans models for mitochondriopathies and show that depletion of complex I subunits recapitulates biochemical, cellular and neurodevelopmental aspects of the human diseases. We exploit two models, nuo-5/NDUFS1- and lpd-5/NDUFS4-depleted animals, for a suppressor screening that identifies lutein for its ability to rescue animals' neurodevelopmental deficits. We uncover overexpression of synaptic neuroligin as an evolutionarily conserved consequence of mitochondrial dysfunction, which we find to mediate an early cholinergic defect in C. elegans. We show lutein exerts its beneficial effects by restoring neuroligin expression independently from its antioxidant activity, thus pointing to a possible novel pathogenetic target for the human disease.We would like to thank Professor Tom Johnson at the University of Colorado Boulder and Professor Shane Rea (now at University of Washington) for making possible theinitial customized HMAD RNAi screening. We also would like to thank Alison Kell, Jenny Cho, and Alessandro Torgovnick for technical help with RNAi screen and lifespan; Professor Proksch at the Heinrich Heine University of Duesseldorf for the compound library used in the suppressor screen; Núria Benseny-Cases at MIRAS beamline in ALBA Synchrotron Light Source, Cerdanyola del Vallès in Barcelona, and Genis Rabost for technical assistance with FTIR measurements and analysis; Anthony Luz and Ian Ryde at Duke University for assistance with Seahorse XFe24 analysis and DNA damage assays respectively; Dirk Schwitters for qPCR on brains from WT and NDUFS4−/− mice. Finally, we thank the Caenorhabditis Genetics Center (funded by the National Institutes of Health Office of Research Infrastructure Programs: P40OD010440) as well as the National Bioresource Project (NBRP) for C. elegans strains and Professors Sieburth, Kaplan and Calahorro for providing additional mutants used in this work. The Wood-Whelan fellowship covered Silvia Maglioni costs to visit Joel Meyer Laboratory. This work was possible thanks to financial support from the German Research Foundation (DFG grants VE663-3/1 and VE663/8-1), the Federal Ministry of Education and Research (JPI-HDHL, Grant no. 01EA1602), and the Heinrich Heine University of Duesseldorf (Strategic Research Funding 2014) to NV; the National Institutes of Health (P42ES010356) to J.N.M., a fellowship from the China Scholarship Council (CSC201607030005) to Z.L.; the Spanish Ministry of Science, Innovation and Universities (RTI2018-096273-B-I00) and the ‘Severo Ochoa’ Programme for Centers of Excellence in R&D (SEV-2015-0496) to A.L.; the ERC Stg 337327 MitoPexLyso to N.R.With funding from the Spanish government through the ‘Severo Ochoa Centre of Excellence’ accreditation (CEX2019-000917-S).Peer reviewe
COVID-19 among children seeking primary paediatric care with signs of an acute infection
Aim: It can be challenging to distinguish COVID-19 in children from other common infections. We set out to determine the rate at which children consulting a primary care paediatrician with an acute infection are infected with SARS-CoV-2 and to compare distinct findings. Method: In seven out-patient clinics, children aged 0–13 years with any new respiratory or gastrointestinal symptoms and presumed infection were invited to be tested for SARS-CoV-2. Factors that were correlated with testing positive were determined. Samples were collected from 25 January 2021 to 01 April 2021. Results: Seven hundred and eighty-three children participated in the study (median age 3 years and 0 months, range 1 month to 12 years and 11 months). Three hundred and fifty-eight were female (45.7%). SARS-CoV-2 RNA was detected in 19 (2.4%). The most common symptoms in children with as well as without detectable SARS-CoV-2 RNA were rhinitis, fever and cough. Known recent exposure to a case of COVID-19 was significantly correlated with testing positive, but symptoms or clinical findings were not. Conclusion: COVID-19 among the children with symptoms of an acute infection was uncommon, and the clinical presentation did not differ significantly between children with and without evidence of an infection with SARS-CoV-2
2-d stability of the Neel wall
We are interested in thin-film samples in micromagnetism, where the magnetization m is a 2-d unit-length vector field. More precisely we are interested in transition layers which connect two opposite magnetizations, so called N\ue9el walls. We prove stability of the 1-d transition layer under 2-d perturbations. This amounts to the investigation of the following singularly perturbed energy functional: E2d(m)= \u3b5 2b | 07m| 2dx + 1/2 2b | 07-1/2 07\u307m|2dx. The topological structure of this two-dimensional problem allows us to use a duality argument to infer the optimal lower bound. The lower bound relies on an \u3b5-perturbation of the following logarithmically failing interpolation inequality 2b | 071/2/\u3c6|2dx \uac sup|\u3c6| 2b | 07\u3c6|dx
Modulation of oxidative phosphorylation and redox homeostasis in mitochondrial NDUFS4 deficiency via mesenchymal stem cells
Abstract Background Disorders of the oxidative phosphorylation (OXPHOS) system represent a large group among the inborn errors of metabolism. The most frequently observed biochemical defect is isolated deficiency of mitochondrial complex I (CI). No effective treatment strategies for CI deficiency are so far available. The purpose of this study was to investigate whether and how mesenchymal stem cells (MSCs) are able to modulate metabolic function in fibroblast cell models of CI deficiency. Methods We used human and murine fibroblasts with a defect in the nuclear DNA encoded NDUFS4 subunit of CI. Fibroblasts were co-cultured with MSCs under different stress conditions and intercellular mitochondrial transfer was assessed by flow cytometry and fluorescence microscopy. Reactive oxygen species (ROS) levels were measured using MitoSOX-Red. Protein levels of CI were analysed by blue native polyacrylamide gel electrophoresis (BN-PAGE). Results Direct cellular interactions and mitochondrial transfer between MSCs and human as well as mouse fibroblast cell lines were demonstrated. Mitochondrial transfer was visible in 13.2% and 6% of fibroblasts (e.g. fibroblasts containing MSC mitochondria) for human and mouse cell lines, respectively. The transfer rate could be further stimulated via treatment of cells with TNF-α. MSCs effectively lowered cellular ROS production in NDUFS4-deficient fibroblast cell lines (either directly via co-culture or indirectly via incubation of cell lines with cell-free MSC supernatant). However, CI protein expression and activity were not rescued by MSC treatment. Conclusion This study demonstrates the interplay between MSCs and fibroblast cell models of isolated CI deficiency including transfer of mitochondria as well as modulation of cellular ROS levels. Further exploration of these cellular interactions might help to develop MSC-based treatment strategies for human CI deficiency
The long non-coding RNA HOTAIRM1 promotes tumor aggressiveness and radiotherapy resistance in glioblastoma
Glioblastoma is the most common malignant primary brain tumor. To date, clinically relevant biomarkers are restricted to isocitrate dehydrogenase (IDH) gene 1 or 2 mutations and O6-methylguanine DNA methyltransferase (MGMT) promoter methylation. Long non-coding RNAs (lncRNAs) have been shown to contribute to glioblastoma pathogenesis and could potentially serve as novel biomarkers. The clinical significance of HOXA Transcript Antisense RNA, Myeloid-Specific 1 (HOTAIRM1) was determined by analyzing HOTAIRM1 in multiple glioblastoma gene expression data sets for associations with prognosis, as well as, IDH mutation and MGMT promoter methylation status. Finally, the role of HOTAIRM1 in glioblastoma biology and radiotherapy resistance was characterized in vitro and in vivo. We identified HOTAIRM1 as a candidate lncRNA whose up-regulation is significantly associated with shorter survival of glioblastoma patients, independent from IDH mutation and MGMT promoter methylation. Glioblastoma cell line models uniformly showed reduced cell viability, decreased invasive growth and diminished colony formation capacity upon HOTAIRM1 down-regulation. Integrated proteogenomic analyses revealed impaired mitochondrial function and determination of reactive oxygen species (ROS) levels confirmed increased ROS levels upon HOTAIRM1 knock-down. HOTAIRM1 knock-down decreased expression of transglutaminase 2 (TGM2), a candidate protein implicated in mitochondrial function, and knock-down of TGM2 mimicked the phenotype of HOTAIRM1 down-regulation in glioblastoma cells. Moreover, HOTAIRM1 modulates radiosensitivity of glioblastoma cells both in vitro and in vivo. Our data support a role for HOTAIRM1 as a driver of biological aggressiveness, radioresistance and poor outcome in glioblastoma. Targeting HOTAIRM1 may be a promising new therapeutic approach