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https://www.readbyqxmd.com/read/28436588/reduced-erg-dosage-impairs-survival-of-hematopoietic-stem-and-progenitor-cells
#1
Ying Xie, Mia Lee Koch, Xin Zhang, Melanie J Hamblen, Frank J Godinho, Yuko Fujiwara, Huafeng Xie, Jan-Henning Klusmann, Stuart H Orkin, Zhe Li
ERG, an ETS family transcription factor frequently overexpressed in human leukemia, has been implicated as a key regulator of hematopoietic stem cells. However, how ERG controls normal hematopoiesis, particularly at the stem and progenitor cell level, and how it contributes to leukemogenesis remain incompletely understood. Using homologous recombination, we generated an Erg knockdown allele (Erg(kd) ) in which Erg expression can be conditionally restored by Cre recombinase. Erg(kd/kd) animals die at E10.5-E11...
April 24, 2017: Stem Cells
https://www.readbyqxmd.com/read/28424158/generation-of-a-multipurpose-prdm16-allele-by-targeted-trapping
#2
A Strassman, F Schnütgen, Q Dai, J C Jones, A C Gomez, L Pitstick, N E Holton, R Moskal, E R Leslie, H von Melchner, D R Beier, B C Bjork
Gene trap mutagenesis is a powerful tool to create loss-of-function mutations in mice and other model organisms. Modifications of traditional gene trap cassettes, including addition of conditional features in the form of Flip-excision (FlEx) arrays enabling directional gene trap cassette inversions by Cre and Flpe site-specific recombinases, greatly enhanced their experimental potential. By taking advantage of these conditional gene trap cassettes, we developed a generic strategy for generating conditional mutations and validated this strategy in mice carrying a multipurpose allele of the Prdm16 transcription factor gene...
April 19, 2017: Disease Models & Mechanisms
https://www.readbyqxmd.com/read/28423326/rapid-molecular-profiling-of-defined-cell-types-using-viral-trap
#3
Alexander R Nectow, Maria V Moya, Mats I Ekstrand, Awni Mousa, Kelly L McGuire, Caroline E Sferrazza, Bianca C Field, Gabrielle S Rabinowitz, Kirsty Sawicka, Yupu Liang, Jeffrey M Friedman, Nathaniel Heintz, Eric F Schmidt
Translational profiling methodologies enable the systematic characterization of cell types in complex tissues, such as the mammalian brain, where neuronal isolation is exceptionally difficult. Here, we report a versatile strategy for profiling CNS cell types in a spatiotemporally restricted fashion by engineering a Cre-dependent adeno-associated virus expressing an EGFP-tagged ribosomal protein (AAV-FLEX-EGFPL10a) to access translating mRNAs by translating ribosome affinity purification (TRAP). We demonstrate the utility of this AAV to target a variety of genetically and anatomically defined neural populations expressing Cre recombinase and illustrate the ability of this viral TRAP (vTRAP) approach to recapitulate the molecular profiles obtained by bacTRAP in corticothalamic neurons across multiple serotypes...
April 18, 2017: Cell Reports
https://www.readbyqxmd.com/read/28420716/a-men1-pancreatic-neuroendocrine-tumour-mouse-model-under-temporal-control
#4
Kate E Lines, Roeland P Vas Nunes, Morten Frost, Christopher J Yates, Mark Stevenson, Rajesh Thakker
Multiple endocrine neoplasia type 1 (MEN1) is an autosomal dominant disorder characterised by occurrence of parathyroid tumours, and neuroendocrine tumours (NETs) of the pancreatic islets and anterior pituitary. The MEN1 gene, encoding menin, is a tumour suppressor, but its precise role in initiating in vivo tumourigenesis remains to be elucidated. The availability of a temporally controlled conditional MEN1 mouse model would greatly facilitate the study of such early tumourigenic events, and overcome the limitations of other MEN1 knockout models, in which menin is lost from conception, or tumour development occurs asynchronously...
April 18, 2017: Endocrine Connections
https://www.readbyqxmd.com/read/28416649/optic-nerve-astrocyte-reactivity-protects-function-in-experimental-glaucoma-and-other-nerve-injuries
#5
Daniel Sun, Sara Moore, Tatjana C Jakobs
Reactive remodeling of optic nerve head astrocytes is consistently observed in glaucoma and other optic nerve injuries. However, it is unknown whether this reactivity is beneficial or harmful for visual function. In this study, we used the Cre recombinase (Cre)-loxP system under regulation of the mouse glial fibrillary acidic protein promoter to knock out the transcription factor signal transducer and activator of transcription 3 (STAT3) from astrocytes and test the effect this has on reactive remodeling, ganglion cell survival, and visual function after experimental glaucoma and nerve crush...
April 17, 2017: Journal of Experimental Medicine
https://www.readbyqxmd.com/read/28416596/microrna-profiling-reveals-marker-of-motor-neuron-disease-in-als-models
#6
Mariah L Hoye, Erica D Koval, Amy J Wegener, Theodore S Hyman, Chengran Yang, David R O'Brien, Rebecca L Miller, Tracy Cole, Kathleen M Schoch, Tao Shen, Tomonori Kunikata, Jean-Philippe Richard, David H Gutmann, Nicholas J Maragakis, Holly B Kordasiewicz, Joseph D Dougherty, Timothy M Miller
Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder marked by the loss of motor neurons (MNs) in the brain and spinal cord, leading to fatally debilitating weakness. Because this disease predominantly affects MNs, we aimed to characterize the distinct expression profile of that cell type in order to elucidate underlying disease mechanisms and identify novel targets that inform on MN health during ALS disease timecourse. microRNAs (miRNAs) are short, non-coding RNAs that can shape the expression profile of a cell and, consequently, often exhibit cell type enriched expression...
April 17, 2017: Journal of Neuroscience: the Official Journal of the Society for Neuroscience
https://www.readbyqxmd.com/read/28412169/engineered-exosomes-as-vehicles-for-biologically-active-proteins
#7
Ulrich Sterzenbach, Ulrich Putz, Ley-Hian Low, John Silke, Seong-Seng Tan, Jason Howitt
Exosomes represent an attractive vehicle for the delivery of biomolecules. However, mechanisms for loading functional molecules into exosomes are relatively unexplored. Here we report the use of the evolutionarily conserved late-domain (L-domain) pathway as a mechanism for loading exogenous proteins into exosomes. We demonstrate that labeling of a target protein, Cre recombinase, with a WW tag leads to recognition by the L-domain-containing protein Ndfip1, resulting in ubiquitination and loading into exosomes...
April 13, 2017: Molecular Therapy: the Journal of the American Society of Gene Therapy
https://www.readbyqxmd.com/read/28409408/spontaneous-recombinase-activity-of-cre-ert2-in-vivo
#8
Jasmin Kristianto, Michael G Johnson, Ryley K Zastrow, Abigail B Radcliff, Robert D Blank
Inducible Cre-ERT recombinase technology is widely used for gene targeting studies. The second generation of inducible Cre-ERT recombinase, hemizygous B6.129S-Tg(UBC-cre/ERT2)1Ejb/J (hereafter abbreviated as Cre-ERT2), a fusion of a mutated estrogen receptor and Cre recombinase, was engineered to be more efficient and specific than the original Cre-ERT. The putative mechanism of selective Cre-mediated recombination is Cre sequestration in the cytoplasm in the basal state with translocation to the nucleus only in the presence of tamoxifen...
April 13, 2017: Transgenic Research
https://www.readbyqxmd.com/read/28409346/development-and-diseases-of-the-collecting-duct-system
#9
Lihe Chen, Paul J Higgins, Wenzheng Zhang
The collecting duct of the mammalian kidney is important for the regulation of extracellular volume, osmolarity, and pH. There are two major structurally and functionally distinct cell types: principal cells and intercalated cells. The former regulates Na(+) and water homeostasis, while the latter participates in acid-base homeostasis. In vivo lineage tracing using Cre recombinase or its derivatives such as CreGFP and CreER(T2) is a powerful new technique to identify stem/progenitor cells in their native environment and to decipher the origins of the tissue that they give rise to...
2017: Results and Problems in Cell Differentiation
https://www.readbyqxmd.com/read/28407687/cytokeratin-19-promoter-directs-the-expression-of-cre-recombinase-in-various-epithelia-of-transgenic-mice
#10
Gui-Feng Zhao, Shuang Zhao, Jia-Jie Liu, Ji-Cheng Wu, Hao-Yu He, Xiao-Qing Ding, Xue-Wen Yu, Ke-Qiang Huang, Zhi-Jie Li, Hua-Chuan Zheng
Cytokeratin 19 (K19) is expressed in various differentiated cells, including gastric, intestinal and bronchial epithelial cells, and liver duct cells. Here, we generated a transgenic mouse line, K19-Cre, in which the expression of Cre recombinase was controlled by the promoter of K19. To test the tissue distribution and excision activity of Cre recombinase, K19-Cre transgenic mice were bred with Rosa26 reporter strain and a mouse strain that carries PTEN conditional alleles (PTENLoxp/Loxp). At mRNA level, Cre was strongly expressed in the stomach, lung and intestine, while in stomach, lung, and liver at protein level...
March 14, 2017: Oncotarget
https://www.readbyqxmd.com/read/28401157/engineering-of-systematic-elimination-of-a-targeted-chromosome-in-human-cells
#11
Hiroshi Sato, Hiroki Kato, Haruyoshi Yamaza, Keiji Masuda, Huong Thi Nguyen Nguyen, Thanh Thi Mai Pham, Xu Han, Yuta Hirofuji, Kazuaki Nonaka
Embryonic trisomy leads to abortion or congenital genetic disorders in humans. The most common autosomal chromosome abnormalities are trisomy of chromosomes 13, 18, and 21. Although alteration of gene dosage is thought to contribute to disorders caused by extra copies of chromosomes, genes associated with specific disease phenotypes remain unclear. To generate a normal cell from a trisomic cell as a means of etiological analysis or candidate therapy for trisomy syndromes, we developed a system to eliminate a targeted chromosome from human cells...
2017: BioMed Research International
https://www.readbyqxmd.com/read/28377509/genetic-tracing-of-hepatocytes-in-liver-homeostasis-injury-and-regeneration
#12
Yue Wang, Xiuzhen Huang, Lingjuan He, Wenjuan Pu, Yan Li, Qiaozhen Liu, Yi Li, Libo Zhang, Wei Yu, Huan Zhao, Yingqun Zhou, Bin Zhou
The liver possesses a remarkable capacity to regenerate after damage. There is a heated debate on the origin of new hepatocytes following injuries in adult liver. Hepatic stem/progenitor cells have been proposed to produce functional hepatocytes after injury. Recent studies have argued against this model and suggested that pre-existing hepatocytes, rather than stem cells, contribute new hepatocytes. This hepatocyte-to-hepatocyte model is mainly based on labeling of hepatocytes with Cre-recombinase delivered by adeno-associated virus (AAV)...
April 4, 2017: Journal of Biological Chemistry
https://www.readbyqxmd.com/read/28377485/ischemic-cerebroprotection-conferred-by-myeloid-lineage-restricted-or-global-cd39-transgene-expression
#13
Amy E Baek, Nadia R Sutton, Danica Petrovic-Djergovic, Hui Liao, Jessica Ray, Joan Park, Yogendra Kanthi, David J Pinsky
Background -Cerebral tissue damage after an ischemic event can be exacerbated by inflammation and thrombosis. Elevated extracellular ATP and ADP levels are associated with cellular injury, inflammation and thrombosis. Ectonucleoside triphosphate diphosphohydrolase-1 (CD39), an enzyme expressed on the plasmalemma of leukocytes and endothelial cells, suppresses platelet activation and leukocyte infiltration by phosphohydrolyzing ATP/ADP. To investigate the effects of increased CD39 in an in vivo cerebral ischemia model, we developed a transgenic (TG) mouse expressing human CD39 (hCD39)...
April 4, 2017: Circulation
https://www.readbyqxmd.com/read/28368537/tissue-specific-ablation-of-the-lif-receptor-in-the-murine-uterine-epithelium-results-in-implantation-failure
#14
JrGang Cheng, Gracy Rosario, Tatiana V Cohen, Jianbo Hu, Colin L Stewart
The cytokine Leukemia inhibitory factor (LIF) is essential for rendering the uterus receptive for blastocyst implantation. In mice LIF receptor expression (LIFR) is largely restricted to the uterine luminal epithelium (LE). LIF, secreted from the endometrial glands (GE) binds to the LIFR, activating the Jak-Stat3 signaling pathway in the LE. JAK-STAT activation converts the LE to a receptive state so that juxtaposed blastocysts begin to implant. To specifically delete the LIFR in the LE we derived a line of mice in which Cre recombinase was inserted into the endogenous Lactoferrin gene (Ltf-Cre)...
March 22, 2017: Endocrinology
https://www.readbyqxmd.com/read/28364122/a-convenient-cas9-based-conditional-knockout-strategy-for-simultaneously-targeting-multiple-genes-in-mouse
#15
Jiang Chen, Yinan Du, Xueyan He, Xingxu Huang, Yun S Shi
The most powerful way to probe protein function is to characterize the consequence of its deletion. Compared to conventional gene knockout (KO), conditional knockout (cKO) provides an advanced gene targeting strategy with which gene deletion can be performed in a spatially and temporally restricted manner. However, for most species that are amphiploid, the widely used Cre-flox conditional KO (cKO) system would need targeting loci in both alleles to be loxP flanked, which in practice, requires time and labor consuming breeding...
March 31, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28363983/synaptotagmin2-syt2-drives-fast-release-redundantly-with-syt1-at-the-output-synapses-of-parvalbumin-expressing-inhibitory-neurons
#16
Brice Bouhours, Enida Gjoni, Olexiy Kochubey, Ralf Schneggenburger
Parvalbumin-expressing inhibitory neurons in the mammalian CNS are specialized for fast transmitter release at their output synapses. However, the Ca(2+) sensor(s) employed by identified inhibitory synapses, including the output synapses of PV-expressing inhibitory neurons, have only recently started to be addressed. Here, we investigated the roles of Syt1 and Syt2 at two types of fast-releasing inhibitory connections in the mammalian CNS, the MNTB to LSO glycinergic synapse, and the basket/stellate cell - Purkinje GABAergic synapse in the cerebellum...
March 31, 2017: Journal of Neuroscience: the Official Journal of the Society for Neuroscience
https://www.readbyqxmd.com/read/28363520/sequence-segregation-improves-non-covalent-protein-delivery
#17
Federica Sgolastra, Coralie M Backlund, E Ilker Ozay, Brittany M deRonde, Lisa M Minter, Gregory N Tew
The impermeability of the plasma membrane towards large, hydrophilic biomolecules is a major obstacle in their use and development against intracellular targets. To overcome such limitations, protein transduction domains (PTDs) have been used as protein carriers, however they often require covalent fusion to the protein for efficient delivery. In an effort to develop more efficient and versatile biological vehicles, a series of PTD-inspired polyoxanorbornene-based synthetic mimics with identical chemical compositions but different hydrophobic/hydrophilic segregation were used to investigate the role of sequence segregation on protein binding and uptake into Jurkat T cells and HEK293Ts...
March 28, 2017: Journal of Controlled Release: Official Journal of the Controlled Release Society
https://www.readbyqxmd.com/read/28361238/a-novel-genetic-tool-for-metabolic-optimization-of-corynebacterium-glutamicum-efficient-and-repetitive-chromosomal-integration-of-synthetic-promoter-driven-expression-libraries
#18
Jing Shen, Jun Chen, Peter Ruhdal Jensen, Christian Solem
Fine-tuning the expression level of multiple genes is usually pivotal for metabolic optimization. We have developed a tool for this purpose for the important industrial workhorse Corynebacterium glutamicum that allows for the introduction of synthetic promoter-driven expression libraries of arbitrary genes. We first devised a method for introducing genetic elements into the chromosome repeatedly, relying on site-specific recombinases and the vector pJS31 serving as the carrier. The pJS31 vector contains a synthetic cassette including a phage attachment site attP for integration, a bacterial attachment site attB for subsequent integration, a multiple cloning site, and two modified loxP sites to facilitate easy removal of undesirable vector elements...
March 30, 2017: Applied Microbiology and Biotechnology
https://www.readbyqxmd.com/read/28360836/genetic-tracing-of-cav3-2-t-type-calcium-channel-expression-in-the-peripheral-nervous-system
#19
Yinth A Bernal Sierra, Julia Haseleu, Alexey Kozlenkov, Valérie Bégay, Gary R Lewin
Characterizing the distinct functions of the T-type ion channel subunits Cav3.1, 3.2 or 3.3 has proven difficult due to their highly conserved amino-acid sequences and the lack of pharmacological blockers specific for each subunit. To precisely determine the expression pattern of the Cav3.2 channel in the nervous system we generated two knock-in mouse strains that express EGFP or Cre recombinase under the control of the Cav3.2 gene promoter. We show that in the brains of these animals, the Cav3.2 channel is predominantly expressed in the dentate gyrus of the hippocampus...
2017: Frontiers in Molecular Neuroscience
https://www.readbyqxmd.com/read/28358866/phenotypically-silent-cre-recombination-within-the-postnatal-ventricular-conduction-system
#20
Samadrita Bhattacharyya, Minoti Bhakta, Nikhil Vilas Munshi
The cardiac conduction system (CCS) is composed of specialized cardiomyocytes that initiate and maintain cardiac rhythm. Any perturbation to the normal sequence of electrical events within the heart can result in cardiac arrhythmias. To understand how cardiac rhythm is established at the molecular level, several genetically modified mouse lines expressing Cre recombinase within specific CCS compartments have been created. In general, Cre driver lines have been generated either by homologous recombination of Cre into an endogenous locus or Cre expression driven by a randomly inserted transgene...
2017: PloS One
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