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Inducible crispr

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https://www.readbyqxmd.com/read/28720717/sting-is-an-essential-mediator-of-the-ku70-mediated-production-of-ifn-%C3%AE-1-in-response-to-exogenous-dna
#1
Hongyan Sui, Ming Zhou, Hiromi Imamichi, Xiaoli Jiao, Brad T Sherman, H Clifford Lane, Tomozumi Imamichi
We previously identified Ku70, a subunit of a DNA repair protein complex, as a cytosolic DNA sensor that induces the production of interferon-λ1 (IFN-λ1) by human primary cells and cell lines. IFN-λ1 is a type III IFN and has similar antiviral activity to that of the type I IFNs (IFN-α and IFN-β). We observed that human embryonic kidney (HEK) 293T cells, which are deficient in the innate immune adaptor protein STING (stimulator of IFN genes), did not produce IFN-λ1 in response to DNA unless they were reconstituted with STING...
July 18, 2017: Science Signaling
https://www.readbyqxmd.com/read/28717054/simple-generation-of-hairless-mice-for-in-vivo-imaging
#2
Yoshikazu Hoshino, Seiya Mizuno, Kanako Kato, Saori Mizuno-Iijima, Yoko Tanimoto, Miyuki Ishida, Noriko Kajiwara, Tomoki Sakasai, Yoshihiro Miwa, Satoru Takahashi, Ken-Ichi Yagami, Fumihiro Sugiyama
The in vivo imaging of mice makes it possible to analyze disease progress non-invasively through reporter gene expression. As the removal of hair improves the accuracy of in vivo imaging, gene-modified mice with a reporter gene are often crossed with Hos:HR-1 mutant mice homozygous for the spontaneous Hr(hr) mutation that exhibit a hair loss phenotype. However, it is time consuming to produce mice carrying both the reporter gene and mutant Hr(hr) gene by mating. In addition, there is a risk that genetic background of the gene-modified mice would be altered by mating...
July 18, 2017: Experimental Animals
https://www.readbyqxmd.com/read/28716076/klf8-regulates-left-right-asymmetric-patterning-through-modulation-of-kupffer-s-vesicle-morphogenesis-and-spaw-expression
#3
Che-Yi Lin, Ming-Yuan Tsai, Yu-Hsiu Liu, Yu-Fen Lu, Yi-Chung Chen, Yun-Ren Lai, Hsin-Chi Liao, Huang-Wei Lien, Chung-Hsiang Yang, Chang-Jen Huang, Sheng-Ping L Hwang
BACKGROUND: Although vertebrates are bilaterally symmetric organisms, their internal organs are distributed asymmetrically along a left-right axis. Disruption of left-right axis asymmetric patterning often occurs in human genetic disorders. In zebrafish embryos, Kupffer's vesicle, like the mouse node, breaks symmetry by inducing asymmetric expression of the Nodal-related gene, spaw, in the left lateral plate mesoderm (LPM). Spaw then stimulates transcription of itself and downstream genes, including lft1, lft2, and pitx2, specifically in the left side of the diencephalon, heart and LPM...
July 17, 2017: Journal of Biomedical Science
https://www.readbyqxmd.com/read/28712502/gene-editing-with-talen-and-crispr-cas-in-rice
#4
Honghao Bi, Bing Yang
Engineered, site-specific nucleases induce genomic double-strand DNA breaks and break repair processes enable genome editing in a plethora of eukaryotic genomes. TALENs (transcription activator-like effector nucleases) and CRISPR/Cas (clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins) are potent biotechnological tools used for genome editing. In rice, species-tailored editing tools have proven to be efficient and easy to use. Both tools are capable of generating DNA double-strand breaks (DSBs) in vivo and such breaks can be repaired either by error-prone NHEJ (nonhomologous end joining) that leads to nucleotide insertions or deletions or by HDR (homology-directed repair) if an appropriate exogenous DNA template is provided...
2017: Progress in Molecular Biology and Translational Science
https://www.readbyqxmd.com/read/28710851/generation-of-apoptosis-resistant-hek293-cells-with-crispr-cas-mediated-quadruple-gene-knockout-for-improved-protein-and-virus-production
#5
Weifeng Zhang, Dan Xiao, Linlin Shan, Junli Zhao, Qinwen Mao, Haibin Xia
Apoptosis has important functions during pathophysiologic processes. However, from a biopharmaceutical point of view, active apoptosis of host cells is undesirable during viral packaging or protein expression, because it decreases the efficiency of viral or protein production. Here we used the CRISPR/Cas technique to knock out four pro-apoptotic genes, Caspase3, Caspase6, Caspase7 and AIF1, in HEK293 cells, and successfully produced an apoptosis-resistant cell line. Furthermore, this cell line showed higher expression levels of pro-apoptotic proteins and higher packaging efficiency for the virus carrying these proteins than control HEK293 cells...
July 15, 2017: Biotechnology and Bioengineering
https://www.readbyqxmd.com/read/28710427/the-essential-role-of-tap73-in-bortezomib-induced-apoptosis-in-p53-deficient-colorectal-cancer-cells
#6
Yasamin Dabiri, Sara Kalman, Clara-Marie Gürth, Jee Young Kim, Viola Mayer, Xinlai Cheng
Mutations in the tumor suppressor p53 are among the most highly occurring events in colorectal cancer (CRC). Such mutations have been shown to influence the sensitivity of cancer cells to chemotherapeutic agents. However their impact on the efficacy of the proteasomal inhibitor bortezomib remains controversial. We thus re-evaluated the toxicity of bortezomib in the CRC cell lines HCT116 wt (wild-type) and its p53-/- clone. Transient resistance to bortezomib treatment was observed in p53-null cells that was later accompanied by an increase in levels and nuclear translocation of TAp73, an isoform of the p53-homologue p73, as well as induction of apoptosis...
July 14, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28707405/generation-of-biallelic-f0-mutants-in-medaka-using-the-crispr-cas9-system
#7
Rie Sawamura, Natsumi Osafune, Takahiro Murakami, Fumiya Furukawa, Takeshi Kitano
Several animal models generated by genome editing methods develop somatic mosaic mutations including wild-type genome sequence in F0 generation because it is difficult to use editing tools at the one-cell stage. Producing complete knockout animals quickly is a great advantage in determining the function of target genes. This study investigated the generation of F0 knockout medaka using the CRISPR/Cas9 system. To determine whether this editing system induced mutations in the medaka genome at the one-cell stage, recombinant Cas9 protein, tracrRNA and crRNA for dead end (dnd), which is essential for germ cell development, were injected into one-cell stage embryos of olvas-DsRedExpress transgenic medaka...
July 14, 2017: Genes to Cells: Devoted to Molecular & Cellular Mechanisms
https://www.readbyqxmd.com/read/28706518/immunization-with-toxoplasma-gondii-gra17-deletion-mutant-induces-partial-protection-and-survival-in-challenged-mice
#8
Jin-Lei Wang, Hany M Elsheikha, Wei-Ning Zhu, Kai Chen, Ting-Ting Li, Dong-Mei Yue, Xiao-Xuan Zhang, Si-Yang Huang, Xing-Quan Zhu
Toxoplasmosis remains a world-threatening disease largely because of the lack of a fully effective vaccine. Here, we created a ΔGRA17 mutant by disrupting the virulence factor GRA17 using CRISPR-Cas9 method. Then, we tested whether ΔGRA17 tachyzoites can be used as a live-attenuated vaccine against acute, chronic, and congenital Toxoplasma gondii infection in mice. Immune response evoked by ΔGRA17 immunization suggested a sequential Th1 and Th2 T cell response, indicated by high levels of Th1 and a mixed Th1/Th2 cytokines at 28 and 70 days after immunization, respectively...
2017: Frontiers in Immunology
https://www.readbyqxmd.com/read/28705828/crispr-induced-null-alleles-show-that-frost-protects-drosophila-melanogaster-reproduction-after-cold-exposure
#9
Claire E Newman, Jantina Toxopeus, Hiroko Udaka, Soohyun Ahn, David M Martynowicz, Steffen P Graether, Brent J Sinclair, Anthony Percival-Smith
The ability to survive and reproduce after cold exposure is important in all kingdoms of life. However, even in a sophisticated genetic model system like Drosophila melanogaster, few genes have been identified as functioning in cold tolerance. The accumulation of the Frost (Fst) gene transcript increases after cold exposure, making it a good candidate for a gene that has a role in cold tolerance. However, despite extensive RNAi knockdown analysis, no role in cold tolerance has been assigned to Fst CRISPR is an effective technique for completely knocking down genes, and less likely to produce off-target effects than GAL4-UAS RNAi systems...
July 13, 2017: Journal of Experimental Biology
https://www.readbyqxmd.com/read/28702937/genetic-engineering-strategies-for-biotic-and-abiotic-stress-tolerance-and-quality-enhancement-in-horticultural-crops-a-comprehensive-review
#10
REVIEW
Nehanjali Parmar, Kunwar Harendra Singh, Deepika Sharma, Lal Singh, Pankaj Kumar, J Nanjundan, Yasin Jeshima Khan, Devendra Kumar Chauhan, Ajay Kumar Thakur
Genetic engineering technique offers myriads of applications in improvement of horticultural crops for biotic and abiotic stress tolerance, and produce quality enhancement. During last two decades, a large number of transgenic horticultural crops has been developed and more are underway. A number of genes including natural and synthetic Cry genes, protease inhibitors, trypsin inhibitors and cystatin genes have been used to incorporate insect and nematode resistance. For providing protection against fungal and bacterial diseases, various genes like chitinase, glucanase, osmotin, defensin and pathogenesis-related genes are being transferred to many horticultural crops world over...
August 2017: 3 Biotech
https://www.readbyqxmd.com/read/28701484/sirt1-regulates-macrophage-self-renewal
#11
Francesco Imperatore, Julien Maurizio, Stephanie Vargas Aguilar, Clara J Busch, Jérémy Favret, Elisabeth Kowenz-Leutz, Wilfried Cathou, Rebecca Gentek, Pierre Perrin, Achim Leutz, Carole Berruyer, Michael H Sieweke
Mature differentiated macrophages can self-maintain by local proliferation in tissues and can be extensively expanded in culture under specific conditions, but the mechanisms of this phenomenon remain only partially defined. Here, we show that SIRT1, an evolutionary conserved regulator of life span, positively affects macrophage self-renewal ability in vitro and in vivo Overexpression of SIRT1 during bone marrow-derived macrophage differentiation increased their proliferative capacity. Conversely, decrease of SIRT1 expression by shRNA inactivation, CRISPR/Cas9 mediated deletion and pharmacological inhibition restricted macrophage self-renewal in culture...
July 12, 2017: EMBO Journal
https://www.readbyqxmd.com/read/28701475/hif-activation-causes-synthetic-lethality-between-the-vhl-tumor-suppressor-and-the-ezh1-histone-methyltransferase
#12
Abhishek A Chakraborty, Eijiro Nakamura, Jun Qi, Amanda Creech, Jacob D Jaffe, Joshiawa Paulk, Jesse S Novak, Kshithija Nagulapalli, Samuel K McBrayer, Glenn S Cowley, Javier Pineda, Jiaxi Song, Yaoyu E Wang, Steven A Carr, David E Root, Sabina Signoretti, James E Bradner, William G Kaelin
Inactivation of the von Hippel-Lindau tumor suppressor protein (pVHL) is the signature lesion in the most common form of kidney cancer, clear cell renal cell carcinoma (ccRCC). pVHL loss causes the transcriptional activation of hypoxia-inducible factor (HIF) target genes, including many genes that encode histone lysine demethylases. Moreover, chromatin regulators are frequently mutated in this disease. We found that ccRCC displays increased H3K27 acetylation and a shift toward mono- or unmethylated H3K27 caused by an HIF-dependent increase in H3K27 demethylase activity...
July 12, 2017: Science Translational Medicine
https://www.readbyqxmd.com/read/28701426/dynamics-of-in-vivo-asc-speck-formation
#13
Paola Kuri, Nicole L Schieber, Thomas Thumberger, Joachim Wittbrodt, Yannick Schwab, Maria Leptin
Activated danger or pathogen sensors trigger assembly of the inflammasome adaptor ASC into specks, large signaling platforms considered hallmarks of inflammasome activation. Because a lack of in vivo tools has prevented the study of endogenous ASC dynamics, we generated a live ASC reporter through CRISPR/Cas9 tagging of the endogenous gene in zebrafish. We see strong ASC expression in the skin and other epithelia that act as barriers to insult. A toxic stimulus triggered speck formation and rapid pyroptosis in keratinocytes in vivo...
July 12, 2017: Journal of Cell Biology
https://www.readbyqxmd.com/read/28700213/enabling-graded-and-large-scale-multiplex-of-desired-genes-using-a-dual-mode-dcas9-activator-in-saccharomyces-cerevisiae
#14
Matthew Deaner, Julio Mejia, Hal S Alper
Standard approaches for dCas9-based modification of gene expression are limited in the ability to multiplex targets, establish streamlined cassettes, and utilize commonly studied Pol II promoters. In this work, we re-purpose the dCas9-VPR activator to act as a dual-mode activator/repressor that can be programmed based solely on target position at gene loci. Furthermore, we implement this approach using a streamlined Pol II-ribozyme system that allows expression of many sgRNAs from a single transcript. By "stepping" dCas9-VPR within the promoter region and ORF we create graded activation and repression (respectively) of target genes, allowing precise control over multiplexed gene modulation...
July 12, 2017: ACS Synthetic Biology
https://www.readbyqxmd.com/read/28699686/ablation-of-ceramide-synthase-2-exacerbates-dextran-sodium-sulphate-induced-colitis-in-mice-due-to-increased-intestinal-permeability
#15
Ye-Ryung Kim, Giora Volpert, Kyong-Oh Shin, So-Yeon Kim, Sun-Hye Shin, Younghay Lee, Sun Hee Sung, Yong-Moon Lee, Jung-Hyuck Ahn, Yael Pewzner-Jung, Woo-Jae Park, Anthony H Futerman, Joo-Won Park
Ceramides mediate crucial cellular processes including cell death and inflammation and have recently been implicated in inflammatory bowel disease. Ceramides consist of a sphingoid long-chain base to which fatty acids of various length can be attached. We now investigate the effect of alerting the ceramide acyl chain length on a mouse model of colitis. Ceramide synthase (CerS) 2 null mice, which lack very-long acyl chain ceramides with concomitant increase of long chain bases and C16-ceramides, were more susceptible to dextran sodium sulphate-induced colitis, and their survival rate was markedly decreased compared with that of wild-type littermates...
July 12, 2017: Journal of Cellular and Molecular Medicine
https://www.readbyqxmd.com/read/28696528/endogenous-sequence-patterns-predispose-the-repair-modes-of-crispr-cas9-induced-dna-double-strand-breaks-in-arabidopsis-thaliana
#16
Giang T H Vu, Hieu X Cao, Friedrich Fauser, Bernd Reiss, Holger Puchta, Ingo Schubert
The possibility to predict the outcome of targeted DNA double strand break (DSB) repair would be desirable for genome editing. Furthermore the consequences of mis-repair of potentially cell-lethal DSBs and the underlying pathways are not yet fully understood. Here we study the CRISPR/Cas9-induced mutation spectra at three selected endogenous loci in Arabidopsis thaliana by deep sequencing of long amplicon libraries. Notably, we found sequence-dependent genomic features impacting the DNA repair outcome. Deletions of 1-<1000 bp size and/or very short insertions, deletions >1 kpb (all due to NHEJ) and deletions combined with insertions between 5->100 bp (caused by an SDSA-like mechanism) occurred most frequently at all three loci...
July 11, 2017: Plant Journal: for Cell and Molecular Biology
https://www.readbyqxmd.com/read/28695896/r-spondin-chromosome-rearrangements-drive-wnt-dependent-tumour-initiation-and-maintenance-in-the-intestine
#17
Teng Han, Emma M Schatoff, Charles Murphy, Maria Paz Zafra, John E Wilkinson, Olivier Elemento, Lukas E Dow
Defining the genetic drivers of cancer progression is a key in understanding disease biology and developing effective targeted therapies. Chromosome rearrangements are a common feature of human malignancies, but whether they represent bona fide cancer drivers and therapeutically actionable targets, requires functional testing. Here, we describe the generation of transgenic, inducible CRISPR-based mouse systems to engineer and study recurrent colon cancer-associated EIF3E-RSPO2 and PTPRK-RSPO3 chromosome rearrangements in vivo...
July 11, 2017: Nature Communications
https://www.readbyqxmd.com/read/28692648/zebrafish-slc30a10-deficiency-revealed-a-novel-compensatory-mechanism-of-atp2c1-in-maintaining-manganese-homeostasis
#18
Zhidan Xia, Jiayu Wei, Yingniang Li, Jia Wang, Wenwen Li, Kai Wang, Xiaoli Hong, Lu Zhao, Caiyong Chen, Junxia Min, Fudi Wang
Recent studies found that mutations in the human SLC30A10 gene, which encodes a manganese (Mn) efflux transporter, are associated with hypermanganesemia with dystonia, polycythemia, and cirrhosis (HMDPC). However, the relationship between Mn metabolism and HMDPC is poorly understood, and no specific treatments are available for this disorder. Here, we generated two zebrafish slc30a10 mutant lines using the CRISPR/Cas9 system. Compared to wild-type animals, mutant adult animals developed significantly higher systemic Mn levels, and Mn accumulated in the brain and liver of mutant embryos in response to exogenous Mn...
July 10, 2017: PLoS Genetics
https://www.readbyqxmd.com/read/28688920/single-cell-rna-sequencing-reveals-an-altered-gene-expression-pattern-as-a-result-of-crispr-cas9-mediated-deletion-of-gene-33-mig6-and-chronic-exposure-to-hexavalent-chromium-in-human-lung-epithelial-cells
#19
Soyoung Park, Xiaowen Zhang, Cen Li, Changhong Yin, Jiangwei Li, John T Fallon, Weihua Huang, Dazhong Xu
Gene 33 (Mig6, ERRFI1) is an adaptor protein with multiple cellular functions. We recently reported that depletion of this protein promotes lung epithelial cell transformation induced by hexavalent chromium [Cr(VI)]. However, the early molecular events that mediate this process are not clear. In the present study, we used single-cell RNA sequencing to compare gene expression profiles between BEAS-2B lung epithelial cells chronically exposed to a sublethal dose of Cr(VI) with or without CRISPR/cas9-mediated deletion of Gene 33...
July 5, 2017: Toxicology and Applied Pharmacology
https://www.readbyqxmd.com/read/28684898/autophagy-regulates-the-stemness-of-cervical-cancer-stem-cells
#20
Yi Yang, Li Yu, Jin Li, Ya Hong Yuan, Xiao Li Wang, Shi Rong Yan, Dong Sheng Li, Yan Ding
Cancer stem cells (CSCs) are a rare population of multipotent cells with the capacity to self-renew. It has been reported that there are CSCs in cervical cancer cells. Pluripotency-associated (PA) transcription factors such as Oct4, Sox2, Nanog and CD44 have been used to isolate CSCs subpopulations. In this study, we showed that autophagy plays an important role in the biological behavior of cervical cancer cells. The expression of the autophagy protein Beclin 1 and LC3B was higher in tumorspheres established from human cervical cancers cell lines (and CaSki) than in the parental adherent cells...
2017: Biologics: Targets & Therapy
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