Differential Role of Human Choline Kinase α and β Enzymes in Lipid Metabolism: Implications in Cancer Onset and Treatment

11 pages, 6 figures, 1 table.Background The Kennedy pathway generates phosphocoline and phosphoethanolamine through its two branches. Choline Kinase (ChoK) is the first enzyme of the Kennedy branch of synthesis of 1phosphocholine, the major component of the plasma membrane. ChoK family of proteins is composed by ChoKα and ChoKβ isoforms, the first one with two different variants of splicing. Recently ChoKα has been implicated in the carcinogenic process, since it is over-expressed in a variety of human cancers. However, no evidence for a role of ChoKβ in carcinogenesis has been reported. Methodology/Principal Findings Here we compare the in vitro and in vivo properties of ChoKα1 and ChoKβ in lipid metabolism, and their potential role in carcinogenesis. Both ChoKα1 and ChoKβ showed choline and ethanolamine kinase activities when assayed in cell extracts, though with different affinity for their substrates. However, they behave differentially when overexpressed in whole cells. Whereas ChoKβ display an ethanolamine kinase role, ChoKα1 present a dual choline/ethanolamine kinase role, suggesting the involvement of each ChoK isoform in distinct biochemical pathways under in vivo conditions. In addition, while overexpression of ChoKα1 is oncogenic when overexpressed in HEK293T or MDCK cells, ChoKβ overexpression is not sufficient to induce in vitro cell transformation nor in vivo tumor growth. Furthermore, a significant upregulation of ChoKα1 mRNA levels in a panel of breast and lung cancer cell lines was found, but no changes in ChoKβ mRNA levels were observed. Finally, MN58b, a previously described potent inhibitor of ChoK with in vivo antitumoral activity, shows more than 20-fold higher efficiency towards ChoKα1 than ChoKβ. Conclusion/Significance This study represents the first evidence of the distinct metabolic role of ChoKα and ChoKβ isoforms, suggesting different physiological roles and implications in human carcinogenesis. These findings constitute a step forward in the design of an antitumoral strategy based on ChoK inhibition.This work has been supported by grants to JCL from Comunidad de Madrid (GR-SAL-0821-2004), Ministerio de Ciencia e Innovación (SAF2008-03750, RD06/0020/0016), Fundación Mutua Madrileña, and by a grant to ARM from Fundación Mutua Madrileña.Peer reviewe

Spatiotemporal Distribution of Key Pelagic Microbes in a Seasonal Oxygen-Deficient Coastal Upwelling System of the Eastern South Pacific Ocean

Indexación: Scopus.The strong seasonal variability in physical-chemical conditions of the Eastern South Pacific Ocean creates an ideal setting to study spatiotemporal distribution of key marine microbial communities. We herein report a nearly 4-year-long time series of the variability in amoA gene counts of ammonia oxidizing archaea (AOA) and bacteria (betaproteobacteria, bAOB) by quantitative PCR, GI.1a Thaumarchaeota and MG-II Euryarchaeota by CARD-FISH, and the picoplanktonic community by flow cytometry for this area. During spring-summer, non-photosynthetic picoplankton such as MG-II Euryarchaeota and GI.1a Thaumarchaeota peaked at the surface and deeper waters, respectively. General AOA and bAOB achieved higher abundances at the oxycline mainly in summer (up to 105–104 amoA copies mL–1). Generalized additive models for location, scale, and shape (GAMLSS) indicated that season and depth account for 19–46% of variations in the abundance of the groups studied, particularly GI.1a Thaumarchaeota and AOA. The oxygen and nitrite concentration were statistically meaningful predictors for the studied groups. GAMLSS models indicate that ammonia oxidizing assemblage’s variability is coupled with ammonia, nitrite, and nitrate variations. Our results indicate that microbial abundances fluctuation is associated with upwelling variability and oxygen-deficient water conditions that shape the substrates availability and metabolic response of marine microbes, including keystone ammonia oxidizing assemblages and their ecological interactions. Overall, our results support planktonic nitrification activity and its contribution to nitrous oxide excess production in the time series off Concepción and the ecological dynamics regarding AOA and bAOB in coastal waters. © Copyright © 2020 Molina, Belmar, Levipan, Ramírez-Flandes, Anguita, Galán, Montes and Ulloa.https://www.frontiersin.org/articles/10.3389/fmars.2020.561597/ful

Contribution of common and rare variants to bipolar disorder susceptibility in extended pedigrees from population isolates.

Current evidence from case/control studies indicates that genetic risk for psychiatric disorders derives primarily from numerous common variants, each with a small phenotypic impact. The literature describing apparent segregation of bipolar disorder (BP) in numerous multigenerational pedigrees suggests that, in such families, large-effect inherited variants might play a greater role. To identify roles of rare and common variants on BP, we conducted genetic analyses in 26 Colombia and Costa Rica pedigrees ascertained for bipolar disorder 1 (BP1), the most severe and heritable form of BP. In these pedigrees, we performed microarray SNP genotyping of 838 individuals and high-coverage whole-genome sequencing of 449 individuals. We compared polygenic risk scores (PRS), estimated using the latest BP1 genome-wide association study (GWAS) summary statistics, between BP1 individuals and related controls. We also evaluated whether BP1 individuals had a higher burden of rare deleterious single-nucleotide variants (SNVs) and rare copy number variants (CNVs) in a set of genes related to BP1. We found that compared with unaffected relatives, BP1 individuals had higher PRS estimated from BP1 GWAS statistics (P = 0.001 ~ 0.007) and displayed modest increase in burdens of rare deleterious SNVs (P = 0.047) and rare CNVs (P = 0.002 ~ 0.033) in genes related to BP1. We did not observe rare variants segregating in the pedigrees. These results suggest that small-to-moderate effect rare and common variants are more likely to contribute to BP1 risk in these extended pedigrees than a few large-effect rare variants

Astrocyte-to-neuron communication through integrin-engaged Thy-1/CBP/Csk/Src complex triggers neurite retraction via the RhoA/ROCK pathway.

Two key proteins for cellular communication between astrocytes and neurons are αvβ3 integrin and the receptor Thy-1. Binding of these molecules in the same (cis) or on adjacent (trans) cellular membranes induces Thy-1 clustering, triggering actin cytoskeleton remodeling. Molecular events that could explain how the Thy-1-αvβ3 integrin interaction signals have only been studied separately in different cell types, and the detailed transcellular communication and signal transduction pathways involved in neuronal cytoskeleton remodeling remain unresolved. Using biochemical and genetic approaches, single-molecule tracking, and high-resolution nanoscopy, we provide evidence that upon binding to αvβ3 integrin, Thy-1 mobility decreased while Thy-1 nanocluster size increased. This occurred concomitantly with inactivation and exclusion of the non-receptor tyrosine kinase Src from the Thy-1/C-terminal Src kinase (Csk)-binding protein (CBP)/Csk complex. The Src inactivation decreased the p190Rho GTPase activating protein phosphorylation, promoting RhoA activation, cofilin, and myosin light chain II phosphorylation and, consequently, neurite shortening. Finally, silencing the adaptor CBP demonstrated that this protein was a key transducer in the Thy-1 signaling cascade. In conclusion, these data support the hypothesis that the Thy-1-CBP-Csk-Src-RhoA-ROCK axis transmitted signals from astrocytic integrin-engaged Thy-1 (trans) to the neuronal actin cytoskeleton. Importantly, the β3 integrin in neurons (cis) was not found to be crucial for neurite shortening. This is the first study to detail the signaling pathway triggered by αvβ3, the endogenous Thy-1 ligand, highlighting the role of membrane-bound integrins as trans acting ligands in astrocyte-neuron communication

3D printing technologies in personalized medicine, nanomedicines, and biopharmaceuticals

3D printing technologies enable medicine customization adapted to patients' needs. There are several 3D printing techniques available, but majority of dosage forms and medical devices are printed using nozzle-based extrusion, laser-writing systems, and powder binder jetting. 3D printing has been demonstrated for a broad range of applications in development and targeting solid, semi-solid and locally applied or implanted medicines. 3D printed solid dosage forms allow the combination of one or more drugs within the same solid dosage form to improve patient compliance, facilitate deglutition, tailor the release profile, or fabricate new medicines for which no dosage form is available. Sustained release 3D-printed implants, stents and medical devices have been used mainly for joint replacement therapies, medical prostheses, and cardiovascular applications. Locally applied medicines such as wound dressing, microneedles, and medicated contact lenses have also been manufactured using 3D printing techniques. The challenge is to select the 3D printing tech-nique most suitable for each application and the type of pharmaceutical ink that should be devel-oped that possesses the required physicochemical and biological performance. The integration of biopharmaceuticals and nanotechnology-based drugs along with 3D printing ("Nanoprinting") brings printed personalized nanomedicines within the most innovative perspectives for the coming years. Continuous manufacturing through the use of 3D-printed microfluidic chips facilitates their translation into clinical practice

Estrategias de guía e ingredientes dietéticos de precisión para enfermedades crónicas en población pre-sénior y sénior

El envejecimiento de la población supone un importante reto, económico y cualitativo, para el sistema de salud orientándolo hacia una atención de tipo preventivo, en la que la nutrición de precisión (NP) y la prescripción de hábitos saludables adquieren relevancia capital. El fin de la NP es procurar una nutrición adaptada a cada individuo, entendiendo que la prevención o el tratamiento de trastornos crónicos (obesidad, diabetes, enfermedad cardiovascular, etc.) deben abordarse de un modo integral, considerando información personal y clínica relevante, edad y características feno- y genotípicas. La elaboración de la presente guía surge de la necesidad de desarrollar modelos nutricionales de precisión que permitan la individualización del tratamiento nutricional, con énfasis en el adulto mayor. Las necesidades nutricionales, las recomendaciones dietéticas y los ingredientes para una NP en las personas pre-sénior y sénior quedan resumidas en realizar al menos 3 comidas diarias, reducir las calorías totales, optar por una alimentación variada y equilibrada con alimentos frescos y de alta densidad nutricional, incorporar verduras, legumbres y pescado, consumir productos lácteos y fibra, preferir carnes blancas en lugar de rojas, evitar frituras, embutidos y alimentos procesados, moderar el consumo de sal, café y alcohol, e hidratarse adecuadamente.The aging of the population underlines an important challenge for the health system not only from sanitary and economic reasons but also by quality perspectives concerning preventive care, where precision nutrition (PN) and the prescription or advice on healthy habits becomes relevant. PN focuses on provide nutrition adapted to each individual, understanding that the prevention or treatment of chronic disorders (obesity, diabetes, cardiovascular disease, etc.) must be addressed in a comprehensive way, considering not only relevant personal and clinical information, but also healthy aging and phenotypical and genotypical features. This guide was prepared due to the need to develop precision nutritional models that allow individualized nutritional treatment for each subject and physiopathological particularities with emphasis on the elderly. Therefore, the requirements of the Spanish pre-senior and senior populations, dietary recommendations and precision foods are reviewed in this document: have at least three daily meals, reduce total calories, choose a varied and balanced diet with fresh foods and high nutritional density, add vegetables, legumes and fish, consume dairy products and fiber, prefer white meat instead of red, avoid fried foods, sausages and processed foods, moderate the consumption of salt, coffee and alcohol, and get hydrated

Morphological changes in diabetic kidney are associated with increased O-GlcNAcylation of cytoskeletal proteins including α-actinin 4

Abstract Purpose The objective of the present study is to identify proteins that change in the extent of the modification with O-linked N-acetylglucosamine (O-GlcNAcylation) in the kidney from diabetic model Goto-Kakizaki (GK) rats, and to discuss the relation between O-GlcNAcylation and the pathological condition in diabetes. Methods O-GlcNAcylated proteins were identified by two-dimensional gel electrophoresis, immunoblotting and peptide mass fingerprinting. The level of O-GlcNAcylation of these proteins was examined by immunoprecipitation, immunoblotting and in situ Proximity Ligation Assay (PLA). Results O-GlcNAcylated proteins that changed significantly in the degree of O-GlcNAcylation were identified as cytoskeletal proteins (α-actin, α-tubulin, α-actinin 4, myosin) and mitochondrial proteins (ATP synthase β, pyruvate carboxylase). The extent of O-GlcNAcylation of the above proteins increased in the diabetic kidney. Immunofluorescence and in situ PLA studies revealed that the levels of O-GlcNAcylation of actin, α-actinin 4 and myosin were significantly increased in the glomerulus and the proximal tubule of the diabetic kidney. Immunoelectron microscopy revealed that immunolabeling of α-actinin 4 is disturbed and increased in the foot process of podocytes of glomerulus and in the microvilli of proximal tubules. Conclusion These results suggest that changes in the O-GlcNAcylation of cytoskeletal proteins are closely associated with the morphological changes in the podocyte foot processes in the glomerulus and in microvilli of proximal tubules in the diabetic kidney. This is the first report to show that α-actinin 4 is O-GlcNAcylated. α-Actinin 4 will be a good marker protein to examine the relation between O-GlcNAcylation and diabetic nephropathy.</p

Activating Transcription Factor 4 Modulates TGFβ-Induced Aggressiveness in Triple-Negative Breast Cancer via SMAD2/3/4 and mTORC2 Signaling

Purpose: On the basis of the identified stress-independent cellular functions of activating transcription factor 4 (ATF4), we reported enhanced ATF4 levels in MCF10A cells treated with TGFβ1. ATF4 is overexpressed in patients with triple-negative breast cancer (TNBC), but its impact on patient survival and the underlying mechanisms remain unknown. We aimed to determine ATF4 effects on patients with breast cancer survival and TNBC aggressiveness, and the relationships between TGFβ and ATF4. Defining the signaling pathways may help us identify a cell signaling-tailored gene signature.Experimental Design: Patient survival data were determined by Kaplan-Meier analysis. Relationship between TGFβ and ATF4, their effects on aggressiveness (tumor proliferation, metastasis, and stemness), and the underlying pathways were analyzed in three TNBC cell lines and in vivo using patient-derived xenografts (PDX).Results: ATF4 overexpression correlated with TNBC patient survival decrease and a SMAD-dependent crosstalk between ATF4 and TGFβ was identified. ATF4 expression inhibition reduced migration, invasiveness, mammosphere-forming efficiency, proliferation, epithelial-mesenchymal transition, and antiapoptotic and stemness marker levels. In PDX models, ATF4 silencing decreased metastases, tumor growth, and relapse after chemotherapy. ATF4 was shown to be active downstream of SMAD2/3/4 and mTORC2, regulating TGFβ/SMAD and mTOR/RAC1-RHOA pathways independently of stress. We defined an eight-gene signature with prognostic potential, altered in 45% of 2,509 patients with breast cancer.Conclusions: ATF4 may represent a valuable prognostic biomarker and therapeutic target in patients with TNBC, and we identified a cell signaling pathway-based gene signature that may contribute to the development of combinatorial targeted therapies for breast cancer