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https://www.readbyqxmd.com/read/29161301/detection-and-quantification-of-dengue-virus-using-a-novel-biosensor-system-based-on-dengue-ns3-protease-activity
#1
Ming-Shu Hsieh, Mei-Yu Chen, Chun-Hsiang Hsieh, Chien-Hsiung Pan, Guann-Yi Yu, Hsin-Wei Chen
BACKGROUND: The traditional methods, plaque assays and immuno-focus assays, used to titrate infectious dengue virus (DENV) particles are time consuming and labor intensive. Here, we developed a DENV protease activity detection system (DENPADS) to visualize DENV infection in cells based on dengue protease activity. METHODOLOGY/PRINCIPAL FINDINGS: Dengue NS3 protease cleaves NS4B-NS5. BHK-21 cells stably expressing the sensor module comprising DENV-2 NS4 and the 10 amino-terminal amino acids of NS5 (N10NS5) fused with the SV40 nuclear localization signal (NLS) and Cre recombinase (Cre), were generated...
2017: PloS One
https://www.readbyqxmd.com/read/29152604/crosstalk-of-lkb1-and-pten-regulated-signals-in-liver-morphogenesis-and-tumor-development
#2
Chengyou Jia, Vivian Medina, Chenchang Liu, Lina He, Daohai Qian, Tu Taojian, Curtis T Okamoto, Bangyan L Stiles
Liver kinase B 1 (LKB1 or STK11) and PTEN (phosphatase and tensin homologue deleted on chromosome 10) are two tumor suppressors that regulate the mTOR signaling pathway. Deletion studies show that loss of either Lkb1 (Lkb(+/-) ) or Pten (Pten(loxP/loxP); Alb-Cre(+) ) leads to liver injury and development of hepatocarcinoma. In this study, we investigated the crosstalk of LKB1 and PTEN loss during tumorigenesis and liver development. We show here that haplo-insufficiency of Lkb1 in the liver leads to advanced tumor development in the Pten null mice (Pten(loxP/loxP); Lkb(loxP/+); Alb-Cre(+) )...
April 2017: Hepatol Commun
https://www.readbyqxmd.com/read/29149457/hmgb1-controls-liver-cancer-initiation-through-yap-dependent-aerobic-glycolysis
#3
Ruochan Chen, Shan Zhu, Xue-Gong Fan, Haichao Wang, Michael T Lotze, Herbert J Zeh, Timothy R Billiar, Rui Kang, Daolin Tang
Emerging studies have suggested that the Hippo pathway is involved in the tumorigenesis of hepatocellular carcinoma (HCC). However, the key regulator of the Hippo pathway in liver tumor metabolic reprogramming remains elusive. Here, we provide evidence to support that high mobility group box 1 (HMGB1), a chromosomal protein, plays a role in the regulation of the Hippo pathway during liver tumorigenesis. Cre/loxP recombination-mediated HMGB1 depletion in hepatocytes blocks diethylnitrosamine-induced liver cancer initiation in mice, whereas shRNA-mediated gene silencing of HMGB1 inhibits HCC cell proliferation...
November 17, 2017: Hepatology: Official Journal of the American Association for the Study of Liver Diseases
https://www.readbyqxmd.com/read/29134654/pten-loss-and-activation-of-k-ras-and-%C3%AE-catenin-cooperate-to-accelerate-prostate-tumourigenesis
#4
Matthew T Jefferies, Adam C Cox, Boris Y Shorning, Valerie Meniel, David Griffiths, Howard G Kynaston, Matthew J Smalley, Alan R Clarke
Aberrant phosphoinositide 3-kinase (PI3K), mitogen-activated protein kinase (MAPK) and WNT signalling are emerging as key events in the multistep nature of prostate tumourigenesis and progression. Here, we report a compound prostate cancer murine model in which these signalling pathways cooperate to produce a more aggressive prostate cancer phenotype. Using Cre-LoxP technology and the probasin promoter, we combined the loss of Pten (Pten(fl/fl) ), to activate the PI3K signalling pathway, with either dominant stabilized β-catenin [Catnb(+/lox(ex3)) ] or activated K-RAS (K-Ras(+/V12) ) to aberrantly activate WNT and MAPK signalling, respectively...
December 2017: Journal of Pathology
https://www.readbyqxmd.com/read/29133437/abnormal-microglia-and-enhanced-inflammation-related-gene-transcription-in-mice-with-conditional-deletion-of-ctcf-in-camk2a-cre-expressing-neurons
#5
Bryan E McGill, Ruteja A Barve, Susan E Maloney, Amy Strickland, Nicholas Rensing, Peter Wang, Michael Wong, Richard Head, David F Wozniak, Jeffrey Milbrandt
CCCTC-binding factor (CTCF) is an 11 zinc finger DNA-binding domain protein that regulates gene expression by modifying three dimensional chromatin structure. Human mutations in CTCF cause intellectual disability and autistic features. Knocking out Ctcf in mouse embryonic neurons is lethal by neonatal age, but the effects of CTCF deficiency in postnatal neurons are less well studied. We knocked out Ctcf postnatally in glutamatergic forebrain neurons under the control of Camk2a-CreCtcf(loxP/loxP) ;Camk2a-Cre+ (Ctcf CKO) mice of both sexes were viable and exhibited profound deficits in spatial learning/memory, impaired motor co-ordination, and decreased sociability by 4 months of age...
November 13, 2017: Journal of Neuroscience: the Official Journal of the Society for Neuroscience
https://www.readbyqxmd.com/read/29131159/enhancing-the-precision-of-genetic-lineage-tracing-using-dual-recombinases
#6
Lingjuan He, Yan Li, Yi Li, Wenjuan Pu, Xiuzhen Huang, Xueying Tian, Yue Wang, Hui Zhang, Qiaozhen Liu, Libo Zhang, Huan Zhao, Juan Tang, Hongbin Ji, Dongqing Cai, Zhibo Han, Zhongchao Han, Yu Nie, Shengshou Hu, Qing-Dong Wang, Ruilin Sun, Jian Fei, Fengchao Wang, Ting Chen, Yan Yan, Hefeng Huang, William T Pu, Bin Zhou
The Cre-loxP recombination system is the most widely used technology for in vivo tracing of stem or progenitor cell lineages. The precision of this genetic system largely depends on the specificity of Cre recombinase expression in targeted stem or progenitor cells. However, Cre expression in nontargeted cell types can complicate the interpretation of lineage-tracing studies and has caused controversy in many previous studies. Here we describe a new genetic lineage tracing system that incorporates the Dre-rox recombination system to enhance the precision of conventional Cre-loxP-mediated lineage tracing...
November 13, 2017: Nature Medicine
https://www.readbyqxmd.com/read/29129659/generation-of-conditional-acvrl1-knockout-mice-by-crispr-cas9-mediated-gene-targeting
#7
Ming Xu, Hongzhi Xu, Jian Chen, Chunjui Chen, Feng Xu, Zhiyong Qin
OBJECTIVES: This study aimed to generate mutant mice containing the Acvrl1 gene flanked with LoxP sequences to allow conditional deletion ofAcvrl1 by the LoxP/Cre system. Such mice may facilitate the development of brain arteriovenous malformation (BAVM) models. METHODS: The CRISPR/Cas9 technique was used to edit Acvrl1. Two single guide RNAs (sgRNAs) with recognition sites on intron 3 and 8 and a donor vector that was homologous with the targeted gene and contained two LoxP sequences were designed and constructed...
November 9, 2017: Molecular and Cellular Probes
https://www.readbyqxmd.com/read/29111103/muscle-specific-knockout-of-general-control-of-amino-acid-synthesis-5-gcn5-does-not-enhance-basal-or-endurance-exercise-induced-mitochondrial-adaptation
#8
Jessica R Dent, Vitor F Martins, Kristoffer Svensson, Samuel A LaBarge, Noah C Schlenk, Mary C Esparza, Elisa H Buckner, Gretchen A Meyer, D Lee Hamilton, Simon Schenk, Andrew Philp
OBJECTIVE: Lysine acetylation is an important post-translational modification that regulates metabolic function in skeletal muscle. The acetyltransferase, general control of amino acid synthesis 5 (GCN5), has been proposed as a regulator of mitochondrial biogenesis via its inhibitory action on peroxisome proliferator activated receptor-γ coactivator-1α (PGC-1α). However, the specific contribution of GCN5 to skeletal muscle metabolism and mitochondrial adaptations to endurance exercise in vivo remain to be defined...
October 16, 2017: Molecular Metabolism
https://www.readbyqxmd.com/read/29109405/application-of-cre-lox-gene-switch-to-limit-the-cry-expression-in-rice-green-tissues
#9
Hao Chen, Ju Luo, Peng Zheng, Xiaobo Zhang, Cuicui Zhang, Xinyuan Li, Mugui Wang, Yuqing Huang, Xuejiao Liu, Mehmood Jan, Yujun Liu, Peisong Hu, Jumin Tu
The presence of genetically modified (GM) protein in the endosperm is important information for the public when considering the biological safety of transgenic rice. To limit the expression of GM proteins to rice green tissues, we developed a modified Cre-lox gene switch using two cassettes named KEY and LOCK. KEY contains a nuclear-localized Cre recombinase driven by the green-tissue-specific promoter rbcS. LOCK contains a Nos terminator (NosT), which is used to block the expression of the gene of interest (GOI), bounded by two loxP sites...
November 6, 2017: Scientific Reports
https://www.readbyqxmd.com/read/29089561/on-demand-optogenetic-activation-of-human-stem-cell-derived-neurons
#10
Simon D Klapper, Evelyn J Sauter, Anka Swiersy, Max A E Hyman, Christian Bamann, Ernst Bamberg, Volker Busskamp
The widespread application of human stem-cell-derived neurons for functional studies is impeded by complicated differentiation protocols, immaturity, and deficient optogene expression as stem cells frequently lose transgene expression over time. Here we report a simple but precise Cre-loxP-based strategy for generating conditional, and thereby stable, optogenetic human stem-cell lines. These cells can be easily and efficiently differentiated into functional neurons, and optogene expression can be triggered by administering Cre protein to the cultures...
October 31, 2017: Scientific Reports
https://www.readbyqxmd.com/read/29089384/the-guanine-nucleotide-exchange-factor-arf-like-protein-13b-is-essential-for-assembly-of-the-mouse-photoreceptor-transition-zone-and-outer-segment
#11
Christin Hanke-Gogokhia, Zhijian Wu, Ali Sharif, Hussein Yazigi, Jeanne M Frederick, Wolfgang Baehr
ARL13b is a small GTPase that functions as a guanosine nucleotide exchange factor (GEF) for ARL3-GDP. ARL13b is located exclusively in photoreceptor outer segments (OS) presumably anchored to discs by palmitoylation, whereas ARL3 is an inner segment cytoplasmic protein. Hypomorphic mutations affecting the ARL13b G‑domain inactivate GEF activity and lead to Joubert Syndrome (JS) in human. However, the molecular mechanisms in ARL13b mutation-induced JS, particularly the function of primary cilia, are still incompletely understood...
October 31, 2017: Journal of Biological Chemistry
https://www.readbyqxmd.com/read/29082358/conditional-disruption-of-mir17-92-in-osteoclasts-led-to-activation-of-osteoclasts-and-loss-of-trabecular-bone-in-part-through-suppression-of-the-mir17-mediated-downregulation-of-protein-tyrosine-phosphatase-oc-in-mice
#12
Kin-Hing William Lau, Virginia M Stiffel, Charles H Rundle, Mehran Amoui, Jordan Tapia, Tyler D White, Matilda H-C Sheng
This study sought to understand the regulation of an osteoclastic protein-tyrosine phosphatase (PTP-oc), a positive regulator of osteoclast activaty. Our past studies suggested that PTP-oc is regulated post-transcriptionally. The 3'-UTR of PTP-oc mRNA contains a target site for miR17. During osteoclastic differentiation, there was an inverse relationship between the cellular levels of miR17 (expressed as one of the six cluster genes of miR17~92) and PTP-oc mRNA. Overexpression of pre-miR17~92 in mouse osteoclast precursors reduced PTP-oc mRNA level and the size of the derived osteoclasts; whereas deletion of miR17~92 or inhibition of miR17 resulted in the formation of larger osteoclasts containing more nuclei that expressed higher PTP-oc mRNA levels and created larger resorption pits...
October 5, 2017: JBMR Plus
https://www.readbyqxmd.com/read/29070461/-rictor-mtorc2-regulates-blood-testis-barrier-and-spermatogenesis-in-mice
#13
He-Ling Dong, Hong-Yuan Wu, You Fu, Meng Dai, Xiao-Chun Bai, Hong Wang
OBJECTIVE: To investigate the role of Rictor/mTORC2 in the formation of blood testis barrier (BTB), testicular development, and spermatogenesis. METHODS: Amh Cre positive mice homozygous for rictor loxP with Sertoli cell specific deletion of rictor were obtained by cross breeding Amh Cre mice with rictor loxP mice. The histology of the reproductive organs, seminiferous tubules and epididymis of the transgenic mice was observed with HE staining. The cell subgroups of the germ cells in the seminiferous tubule were detected by flow cytometry with propidium iodide labeling...
October 20, 2017: Nan Fang Yi Ke da Xue Xue Bao, Journal of Southern Medical University
https://www.readbyqxmd.com/read/29069450/improvement-of-a-yeast-self-excising-integrative-vector-by-prevention-of-expression-leakage-of-the-intronated-cre-recombinase-gene-during-plasmid-maintenance-in-escherichia-coli
#14
Michael O Agaphonov
The use of plasmids possessing a regulatable gene coding for a site-specific recombinase together with its recognition sequences significantly facilitates genome manipulations since it allows self-excision of the portion of the genetic construct integrated into the host genome. Stable maintenance of such plasmids in Escherichia coli, which is used for plasmid preparation, requires prevention of recombinase synthesis in this host, which can be achieved by interrupting the recombinase gene with an intron. Based on this approach, Saccharomyces cerevisiae and Hansenula polymorpha self-excising vectors possessing intronated gene for Cre recombinase and its recognition sites (LoxP) were previously constructed...
December 1, 2017: FEMS Microbiology Letters
https://www.readbyqxmd.com/read/29052235/establishment-of-a-markerless-multiple-gene-deletion-method-based-on-cre-loxp-mutant-system-for-bacillus-pumilus
#15
Zheng-Bing Guan, Kai-Qiang Wang, Yan Shui, Xiang-Ru Liao
In this study, we established a Cre/loxP mutant recombination system (Cre/lox71-66 system) for markerless gene deletion to facilitate our follow-up rational genetic engineering to the strain Bacillus pumilus W3. This modified method uses two mutant loxP sites, which after recombination creates a double-mutant loxP site that is poorly recognized by Cre recombinase, facilitating multiple gene deletions in a single genetic background. Two selected genes, cotA and sigF, were continuously knocked out and verified at different levels using this method...
October 20, 2017: Journal of Basic Microbiology
https://www.readbyqxmd.com/read/29051279/expansion-of-epor-negative-macrophages-besides-erythroblasts-by-elevated-epor-signaling-in-erythrocytosis-mouse-models
#16
Jieyu Wang, Yoshihiro Hayashi, Asumi Yokota, Zefeng Xu, Yue Zhang, Rui Huang, Xiaomei Yan, Hongyun Liu, Liping Ma, Mohammad Azam, James P Bridges, Jose A Cancelas, Theodosia A Kalfa, Xiuli An, Zhijian Xiao, Gang Huang
Activated EPO receptor (EPOR) signaling causes erythrocytosis. The important role of macrophages for the erythroid expansion and differentiation process has been reported, both in baseline and stress erythropoiesis. However, the significance of EPOR signaling for regulation of macrophages contributing to erythropoiesis has not been fully understood. Here we show that EPOR signaling activation quickly expands both erythrocytes and macrophages in vivo in mouse models of primary and secondary erythrocytosis. To mimic the chimeric condition and expansion of the disease clone in the polycythemia vera patients, we combined Cre-inducible Jak2V617F/+ allele with LysM-Cre allele which expresses in mature myeloid cells and some of the HSC/Ps (LysM-Cre;Jak2V617F/+ mice)...
October 19, 2017: Haematologica
https://www.readbyqxmd.com/read/29034260/clodronate-improves-survival-of-transplanted-hoxb8-myeloid-progenitors-with-constitutively-active-gmcsfr-in-immunocompetent-mice
#17
Simon Lee, Saul Kivimäe, Francis C Szoka
New methods to produce large numbers of myeloid progenitor cells, precursors to macrophages (MΦs), by maintaining Hoxb8 transcription factor activity(1) has reinvigorated interest in MΦ cell therapies. We generated Hoxb8-dependent myeloid progenitors (HDPs) by transducing lineage-negative bone marrow cells with a constitutively expressed Hoxb8 flanked by loxP. HDPs proliferate indefinitely and differentiate into MΦ when Hoxb8 is removed by a tamoxifen-inducible Cre. We genetically modified HDPs with a constitutively active GMCSF receptor and the tamoxifen-induced transcription factor IRF8, which we have termed "HDP-on...
December 15, 2017: Molecular Therapy. Methods & Clinical Development
https://www.readbyqxmd.com/read/29031725/intestinal-sirt3-overexpression-in-mice-improves-whole-body-glucose-homeostasis-independent-of-body-weight
#18
Deepti Ramachandran, Rosmarie Clara, Shahana Fedele, Junmin Hu, Endre Lackzo, Jing-Yi Huang, Eric Verdin, Wolfgang Langhans, Abdelhak Mansouri
OBJECTIVE: Intestinal metabolism might play a greater role in regulating whole body metabolism than previously believed. We aimed to enhance enterocyte metabolism in mice and investigate if it plays a role in diet-induced obesity (DIO) and its comorbidities. METHODS: Using the cre-loxP system, we overexpressed the mitochondrial NAD(+) dependent protein deacetylase SIRT3 in enterocytes of mice (iSIRT3 mice). We chronically fed iSIRT3 mice and floxed-SIRT3 control (S3fl) mice a low-fat, control diet (CD) or a high-fat diet (HFD) and then phenotyped the mice...
October 2017: Molecular Metabolism
https://www.readbyqxmd.com/read/29030008/assessment-of-leukocyte-activity-in-mice-devoid-of-the-glucocorticoid-receptor-in-the-noradrenergic-system-gr-dbhcre
#19
Adam Roman, Justyna Kuśmierczyk, Grzegorz Kreiner, Irena Nalepa
Disturbances in brain monoamines, overactivity of the hypothalamo-pituitary adrenal (HPA) axis and pro-inflammatory tendency in the immune system are the key features of depressive disorders. Recently, several murine lines with mutations in glucocorticoid receptors (GRs) have been generated and these animals may be utilized for study depressive-like disorders. In the present study, we have investigated whether selective ablation of GRs in noradrenergic neurons affects functional properties of leukocytes and redirects them towards pro-inflammatory activity...
October 8, 2017: Immunobiology
https://www.readbyqxmd.com/read/29029620/tgf%C3%AE-superfamily-signaling-and-uterine-decidualization
#20
REVIEW
Nan Ni, Qinglei Li
Decidualization is an intricate biological process where extensive morphological, functional, and genetic changes take place in endometrial stromal cells to support the development of an implanting blastocyst. Deficiencies in decidualization are associated with pregnancy complications and reproductive diseases. Decidualization is coordinately regulated by steroid hormones, growth factors, and molecular and epigenetic mechanisms. Transforming growth factor β (TGFβ) superfamily signaling regulates multifaceted reproductive processes...
October 13, 2017: Reproductive Biology and Endocrinology: RB&E
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