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https://www.readbyqxmd.com/read/29649238/generation-of-multidrug-resistant-human-tissues-by-overexpression-of-the-abcg2-multidrug-transporter-in-embryonic-stem-cells
#1
Zsuzsa Erdei, Anita Schamberger, György Török, Kornélia Szebényi, György Várady, Tamás I Orbán, László Homolya, Balázs Sarkadi, Ágota Apáti
The ABCG2 multidrug transporter provides resistance against various endo- and xenobiotics, and protects the stem cells against toxins and stress conditions. We have shown earlier that a GFP-tagged version of ABCG2 is fully functional and may be used to follow the expression, localization and function of this transporter in living cells. In the present work we have overexpressed GFP-ABCG2, driven by a constitutive (CAG) promoter, in HUES9 human embryonic stem cells. Stem cell clones were generated to express the wild-type and a substrate-mutant (R482G) GFP-ABCG2 variant, by using the Sleeping Beauty transposon system...
2018: PloS One
https://www.readbyqxmd.com/read/29622795/antitumor-activity-of-cd56-chimeric-antigen-receptor-t-cells-in-neuroblastoma-and-sclc-models
#2
Denise L Crossland, Warren L Denning, Sonny Ang, Simon Olivares, Tiejuan Mi, Kirsten Switzer, Harjeet Singh, Helen Huls, Kate S Gold, Bonnie S Glisson, Laurence J Cooper, John V Heymach
The CD56 antigen (NCAM-1) is highly expressed on several malignancies with neuronal or neuroendocrine differentiation, including small-cell lung cancer and neuroblastoma, tumor types for which new therapeutic options are needed. We hypothesized that CD56-specific chimeric antigen receptor (CAR) T cells could target and eliminate CD56-positive malignancies. Sleeping Beauty transposon-generated CD56R-CAR T cells exhibited αβT-cell receptors, released antitumor cytokines upon co-culture with CD56+ tumor targets, demonstrated a lack of fratricide, and expression of cytolytic function in the presence of CD56+ stimulation...
April 6, 2018: Oncogene
https://www.readbyqxmd.com/read/29622087/generation-of-venus-fluorochrome-expressing-transgenic-handmade-cloned-buffalo-embryos-using-sleeping-beauty-transposon
#3
Dharmendra Kumar, Papori Sharma, Kennady Vijayalakshmy, Naresh L Selokar, Pradeep Kumar, Rasika Rajendran, P S Yadav
The objective of this study was to optimise the electroporation conditions for efficient integration of Venus construct in buffalo fetal fibroblasts using Sleeping Beauty (SB) based transposition and to produce Venus expressing transgenic cloned embryos through handmade cloning (HMC) approach. Primary culture of buffalo fetal fibroblast cells was established and subsequently cultured cells were co-transfected with Venus and helper plasmid at different combinations of electroporation condition. In different combinations of voltage, time and plasmid dose, we observed that 300 V, single pulse for 10 ms in 2 mm cuvette and 1...
April 2018: Tissue & Cell
https://www.readbyqxmd.com/read/29587177/advances-in-functional-genetic-screening-with-transposons-and-crispr-cas9-to-illuminate-cancer-biology
#4
REVIEW
Kathryn A O'Donnell
Large-scale genome sequencing studies have identified a wealth of mutations in human tumors and have dramatically advanced the field of cancer genetics. However, the functional consequences of an altered gene in tumor progression cannot always be inferred from mutation status alone. This underscores the critical need for complementary methods to assign functional significance to mutated genes in cancer. Transposons are mobile genetic elements that serve as powerful tools for insertional mutagenesis. Over the last decade, investigators have employed mouse models with on-demand transposon-mediated mutagenesis to perform unbiased genetic screens to identify clinically relevant genes that participate in the pathogenesis of human cancer...
March 24, 2018: Current Opinion in Genetics & Development
https://www.readbyqxmd.com/read/29575648/spontaneous-development-of-intratumoral-heterogeneity-in-a-transposon-induced-mouse-model-of-glioma
#5
Keisuke Sumiyoshi, Hideto Koso, Sumiko Watanabe
Glioma is the most common form of malignant brain cancer in adults. The Sleeping Beauty (SB) transposon-based glioma mouse model allows for effective in vivo analysis of candidate genes. In this study, we developed a transposon vector that encodes the triple combination of platelet-derived growth factor subunit A (PDGFA), and shRNAs against Nf1 and Trp53 (shNf1/shp53). Initiation and progression of glioma in the brain were monitored by expression of a fluorescent protein. Transduction of the vector into neural progenitor and stem cells (NPCs) in the subventricular zone (SVZ) of the neonatal brain induced proliferation of oligodendrocyte precursor cells, and promoted formation of highly penetrant malignant gliomas within 2 to 4 months...
March 25, 2018: Cancer Science
https://www.readbyqxmd.com/read/29572319/ossabaw-pigs-with-a-pcsk9-gain-of-function-mutation-develop-accelerated-coronary-atherosclerotic-lesions-a-novel-model-for-preclinical-studies
#6
Fang Yuan, Liang Guo, Kyoung-Ha Park, John R Woollard, Kwon Taek-Geun, Kai Jiang, Tamene Melkamu, Bin Zang, Samantha L Smith, Scott C Fahrenkrug, Frank D Kolodgie, Amir Lerman, Renu Virmani, Lilach O Lerman, Daniel F Carlson
BACKGROUND: Ossabaw pigs are unique miniature swine with genetic predisposition to develop metabolic syndrome and coronary atherosclerosis after extended periods receiving atherogenic diets. We have hypothesized that transgenic Ossabaw swine expressing chimp PCSK9 (proprotein convertase subtilisin-like/kexin type 9) containing the D374Y gain of function would develop familial hypercholesterolemia and coronary artery plaques more rapidly than Landrace swine with the same transgene. METHODS AND RESULTS: Ossabaw and Landrace PCSK9 gain-of-function founders were generated by Sleeping Beauty transposition and cloning...
March 23, 2018: Journal of the American Heart Association
https://www.readbyqxmd.com/read/29562762/efficient-non-viral-t-cell-engineering-by-sleeping-beauty-minicircles-diminishing-dna-toxicity-and-mirnas-silencing-the-endogenous-tcr
#7
Julian Clauß, Matthias Obenaus, Csaba Miskey, Zoltán Ivics, Zsuzsanna Izsvák, Wolfgang Uckert, Mario Bunse
Transposon-based vectors have entered clinical trials as an alternative to viral vectors for genetic engineering of T cells. However, transposon vectors require DNA transfection into T cells which we found to cause adverse effects. T cell viability was decreased in a dose-dependent manner and DNA-transfected T cells showed a delayed response upon T cell receptor (TCR) stimulation with regard to blast formation, proliferation and surface expression of CD25 and CD28. Gene expression analysis demonstrated a DNA-dependent induction of a type I interferon response and IFN-β upregulation...
March 21, 2018: Human Gene Therapy
https://www.readbyqxmd.com/read/29542085/the-enhancer-trap-in-ciona
#8
Yasunori Sasakura
Enhancer trap is a famous application of transposons. This method is useful for the creation of marker transgenic lines that express a reporter gene in tissue- or organ-specific manner, characterization of enhancers in the genome, finding novel patterns of gene expression, and mutagenesis. In Ciona intestinalis, efficient enhancer traps with Minos and Sleeping beauty transposons have been reported. With the enhancer trap lines, the intronic enhancers regulating the expression of the Musashi gene, the compartment in the digestive tube, the presence of enhancers sensitive to the orientation of the gene that they regulate, and the functions of the Hox1 gene have been revealed...
2018: Advances in Experimental Medicine and Biology
https://www.readbyqxmd.com/read/29503198/efficient-non-viral-gene-delivery-into-human-hematopoietic-stem-cells-by-minicircle-sleeping-beauty-transposon-vectors
#9
Marta Holstein, Cristina Mesa-Nuñez, Csaba Miskey, Elena Almarza, Valentina Poletti, Marco Schmeer, Esther Grueso, Juan Carlos Ordóñez Flores, Dennis Kobelt, Wolfgang Walther, Manish K Aneja, Johannes Geiger, Halvard B Bonig, Zsuzsanna Izsvák, Martin Schleef, Carsten Rudolph, Fulvio Mavilio, Juan A Bueren, Guillermo Guenechea, Zoltán Ivics
The Sleeping Beauty (SB) transposon system is a non-viral gene delivery platform that combines simplicity, inexpensive manufacture, and favorable safety features in the context of human applications. However, efficient correction of hematopoietic stem and progenitor cells (HSPCs) with non-viral vector systems, including SB, demands further refinement of gene delivery techniques. We set out to improve SB gene transfer into hard-to-transfect human CD34+ cells by vectorizing the SB system components in the form of minicircles that are devoid of plasmid backbone sequences and are, therefore, significantly reduced in size...
January 31, 2018: Molecular Therapy: the Journal of the American Society of Gene Therapy
https://www.readbyqxmd.com/read/29470680/changes-in-skeletal-muscle-and-body-weight-on-sleeping-beauty-transposon-mediated-transgenic-mice-overexpressing-pig-migf-1
#10
Bo Gao, Wei Wang, Han Wu, Cai Chen, Dan Shen, Saisai Wang, Wei Chen, Li Zhang, Shuheng Chan, Chengyi Song
Insulin-like growth factor (IGF-I) is an important growth factor in mammals, but the functions of the local muscle-specific isoform of insulin-like growth factor 1 (mIGF-1) to skeletal muscle development have rarely been reported. To determine the effect of pig mIGF-1 on body development and muscle deposition in vivo and to investigate the molecular mechanisms, the transgenic mouse model was generated which can also provide experimental data for making transgenic pigs with pig endogenous IGF1 gene. We constructed a skeletal muscle-specific expression vector using 5'- and 3'-regulatory regions of porcine skeletal α-actin gene...
February 22, 2018: Biochemical Genetics
https://www.readbyqxmd.com/read/29449753/do-younger-sleeping-beauties-prefer-a-technological-prince
#11
Anthony F J van Raan, Jos J Winnink
In this paper we investigate recent Sleeping Beauties cited in patents (SB-SNPRs). We find that the increasing trend of the relative number of SBs stopped around 1998. Moreover, we find that the time lag between the publication year of the SB-SNPRs and their first citation in a patent is becoming shorter in recent years. Our observations also suggest that, on average, in the more recent years SBs are awakened increasingly earlier by a 'technological prince' rather than by a 'scientific prince'. These observations suggest that SBs with technological importance are 'discovered' earlier in an application-oriented context...
2018: Scientometrics
https://www.readbyqxmd.com/read/29445422/sleeping-beauty-transposon-integrates-into-non-ta-dinucleotides
#12
Yabin Guo, Yin Zhang, Kaishun Hu
Background: Sleeping Beauty transposon (SB) has become an increasingly important genetic tool for generating mutations in vertebrate cells. It is widely thought that SB exclusively integrates into TA dinucleotides. However, this strict TA-preference has not been rigorously tested in large numbers of insertion sites that now can be detected with next generation sequencing. Li et al. found 71 SB insertions in non-TA dinucleotides in 2013, suggesting that TA dinucleotides are not the only sites of SB integration, yet further studies on this topic have not been carried out...
2018: Mobile DNA
https://www.readbyqxmd.com/read/29444170/genotypic-and-phenotypic-characterization-of-the-sdccag8tn-sb-tyr-2161b-ca1c2ove-mouse-model
#13
Katie Weihbrecht, Wesley A Goar, Calvin S Carter, Val C Sheffield, Seongjin Seo
Nephronophthisis-related ciliopathies (NPHP-RC) are a group of disorders that present with end-stage renal failure in childhood/adolescence, kidney cysts, retinal degeneration, and cerebellar hypoplasia. One disorder that shares clinical features with NPHP-RC is Bardet-Biedl Syndrome (BBS). Serologically defined colon cancer antigen 8 (SDCCAG8; also known as NPHP10 and BBS16) is an NPHP gene that is also associated with BBS. To better understand the patho-mechanisms of NPHP and BBS caused by loss of SDCCAG8 function, we characterized an SDCCAG8 mouse model (Sdccag8Tn(sb-Tyr)2161B...
2018: PloS One
https://www.readbyqxmd.com/read/29443066/constitutive-and-inducible-systems-for-genetic-in-vivo-modification-of-mouse-hepatocytes-using-hydrodynamic-tail-vein-injection
#14
Eric K Hubner, Christian Lechler, Thomas N Rösner, Birgit Kohnke-Ertel, Roland M Schmid, Ursula Ehmer
In research models of liver cancer, regeneration, inflammation, and fibrosis, flexible systems for in vivo gene expression and silencing are highly useful. Hydrodynamic tail vein injection of transposon-based constructs is an efficient method for genetic manipulation of hepatocytes in adult mice. In addition to constitutive transgene expression, this system can be used for more advanced applications, such as shRNA-mediated gene knock-down, implication of the CRISPR/Cas9 system to induce gene mutations, or inducible systems...
February 2, 2018: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/29432776/establishment-of-cre-mediated-hbv-recombinant-cccdna-rcccdna-cell-line-for-cccdna-biology-and-antiviral-screening-assays
#15
Min Wu, Jin Li, Lei Yue, Lu Bai, Yaming Li, Jieliang Chen, Xiaonan Zhang, Zhenghong Yuan
Hepatitis B virus (HBV) covalently closed circular DNA (cccDNA), existing in hepatocyte nuclei as a stable minichromosome, plays a central role in the life cycle of the virus and permits the persistence of infection. Despite being essential for HBV infection, little is known about the molecular mechanisms of cccDNA formation, regulation and degradation, and there is no therapeutic agents directly targeting cccDNA, fore mostly due to the lack of robust, reliable and quantifiable HBV cccDNA models. In this study, combined the Cre/loxP and sleeping beauty transposons system, we established HepG2-derived cell lines integrated with 2-60 copies of monomeric HBV genome flanked by loxP sites (HepG2-HBV/loxP)...
April 2018: Antiviral Research
https://www.readbyqxmd.com/read/29425968/characterization-of-calcium-signals-in-human-induced-pluripotent-stem-cell-derived-dentate-gyrus-neuronal-progenitors-and-mature-neurons-stably-expressing-an-advanced-calcium-indicator-protein
#16
Gergő Vőfély, Tünde Berecz, Eszter Szabó, Kornélia Szebényi, Edit Hathy, Tamás I Orbán, Balázs Sarkadi, László Homolya, Maria C Marchetto, János M Réthelyi, Ágota Apáti
Pluripotent stem cell derived human neuronal progenitor cells (hPSC-NPCs) and their mature neuronal cell culture derivatives may efficiently be used for central nervous system (CNS) drug screening, including the investigation of ligand-induced calcium signalization. We have established hippocampal NPC cultures derived from human induced PSCs, which were previously generated by non-integrating Sendai virus reprogramming. Using established protocols these NPCs were differentiated into hippocampal dentate gyrus neurons...
February 6, 2018: Molecular and Cellular Neurosciences
https://www.readbyqxmd.com/read/29409568/the-effect-of-selective-c-met-inhibitor-on-hcc-in-the-met-active-%C3%AE-catenin-mutated-mouse-model
#17
Na Zhan, Adeola Adebayo Michael, Kaiyuan Wu, Gang Zeng, Aaron Bell, Junyan Tao, Satdarshan P Monga
Simultaneous mutations in CTNNB1 and activation of c-MET occurs in 9-12.5% of patients with hepatocellular cancer (HCC). Co-expression of c-MET-V5 and mutant-β-catenin-Myc in mouse liver by sleeping beauty transposon/transposase and hydrodynamic tail vein injection (SB-HTVI) led to development of HCC with 70% molecular identity to the clinical subset. Using this model, we investigated the effect of EMD1214063, a highly selective c-MET inhibitor. Five weeks after SB-HTVI when tumors were established, EMD1214063 (10mg/kg) was administered by gastric lavage as a single-agent on 5-days-on/3-days-off schedule, compared to vehicle only control...
February 6, 2018: Gene Expression
https://www.readbyqxmd.com/read/29405533/the-role-of-sleep-in-aesthetic-perception-and-empathy-a-mediation-analysis
#18
Sara Peretti, Daniela Tempesta, Valentina Socci, Maria C Pino, Monica Mazza, Marco Valenti, Luigi De Gennaro, Cinzia Di Dio, Antonella Marchetti, Michele Ferrara
The ability to experience aesthetics plays a fundamental role in human social interactions, as well as the capacity to feel empathy. Some studies have shown that beauty perception shares part of the neural network underlying emotional and empathic abilities, which are also known to affect sleep quality and duration. In this study, we evaluated for the first time the effects of sleep on the relation between aesthetic perception and empathic abilities in healthy subjects using a mediation analysis approach. One-hundred and twenty-six subjects participated in this study...
February 6, 2018: Journal of Sleep Research
https://www.readbyqxmd.com/read/29364270/an-efficient-in-vitro-transposition-method-by-a-transcriptionally-regulated-sleeping-beauty-system-packaged-into-an-integration-defective-lentiviral-vector
#19
Daniela Benati, Fabienne Cocchiarella, Alessandra Recchia
The Sleeping Beauty (SB) transposon is a non-viral integrating system with proven efficacy for gene transfer and functional genomics. To optimize the SB transposon machinery, a transcriptionally regulated hyperactive transposase (SB100X) and T2-based transposon are employed. Typically, the transposase and transposon are provided transiently by plasmid transfection and SB100X expression is driven by a constitutive promoter. Here, we describe an efficient method to deliver the SB components to human cells that are resistant to several physical and chemical transfection methods, to control SB100X expression and stably integrate a gene of interest (GOI) through a "cut and paste" SB mechanism...
January 12, 2018: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/29362220/integration-of-oncogenes-via-sleeping-beauty-as-a-mouse-model-of-hpv16-oral-tumors-and-immunologic-control
#20
Yi-Hsin Lin, Ming-Chieh Yang, Ssu-Hsueh Tseng, Rosie Jiang, Andrew Yang, Emily Farmer, Shiwen Peng, Talia Henkle, Yung-Nien Chang, Chien-Fu Hung, T-C Wu
Human papillomavirus type 16 (HPV16) is the etiologic factor for cervical cancer and a subset of oropharyngeal cancers. Although several prophylactic HPV vaccines are available, no effective therapeutic strategies to control active HPV diseases exist. Tumor implantation models are traditionally used to study HPV-associated buccal tumors. However, they fail to address precancerous phases of disease progression and display tumor microenvironments distinct from those observed in patients. Previously, K14-E6/E7 transgenic mouse models have been used to generate spontaneous tumors...
January 23, 2018: Cancer Immunology Research
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