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Anti crispr

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https://www.readbyqxmd.com/read/29789882/rapid-development-of-stable-transgene-cho-cell-lines-by-crispr-cas9-mediated-site-specific-integration-into-c12orf35
#1
Menglin Zhao, Jiaxian Wang, Manyu Luo, Han Luo, Meiqi Zhao, Lei Han, Mengxiao Zhang, Hui Yang, Yueqing Xie, Hua Jiang, Lei Feng, Huili Lu, Jianwei Zhu
Chinese hamster ovary (CHO) cells are the most widely used mammalian hosts for recombinant protein production. However, by conventional random integration strategy, development of a high-expressing and stable recombinant CHO cell line has always been a difficult task due to the heterogenic insertion and its caused requirement of multiple rounds of selection. Site-specific integration of transgenes into CHO hot spots is an ideal strategy to overcome these challenges since it can generate isogenic cell lines with consistent productivity and stability...
May 22, 2018: Applied Microbiology and Biotechnology
https://www.readbyqxmd.com/read/29779790/a-hyperthermophilic-phage-decoration-protein-suggests-common-evolutionary-origin-with-herpesvirus-triplex-proteins-and-an-anti-crispr-protein
#2
Nicholas P Stone, Brendan J Hilbert, Daniel Hidalgo, Kevin T Halloran, Jooyoung Lee, Erik J Sontheimer, Brian A Kelch
Virus capsids are protein shells that protect the viral genome from environmental assaults, while maintaining the high internal pressure of the tightly packaged genome. To elucidate how capsids maintain stability under harsh conditions, we investigated the capsid components of the hyperthermophilic phage P74-26. We determined the structure of capsid protein gp87 and show that it has the same fold as decoration proteins in many other phages, despite lacking significant sequence homology. We also find that gp87 is significantly more stable than mesophilic homologs...
April 26, 2018: Structure
https://www.readbyqxmd.com/read/29769725/mxra8-is-a-receptor-for-multiple-arthritogenic-alphaviruses
#3
Rong Zhang, Arthur S Kim, Julie M Fox, Sharmila Nair, Katherine Basore, William B Klimstra, Rebecca Rimkunas, Rachel H Fong, Hueylie Lin, Subhajit Poddar, James E Crowe, Benjamin J Doranz, Daved H Fremont, Michael S Diamond
Arthritogenic alphaviruses comprise a group of enveloped RNA viruses that are transmitted to humans by mosquitoes and cause debilitating acute and chronic musculoskeletal disease 1 . The host factors required for alphavirus entry remain poorly characterized 2 . Here we use a genome-wide CRISPR-Cas9-based screen to identify the cell adhesion molecule Mxra8 as an entry mediator for multiple emerging arthritogenic alphaviruses, including chikungunya, Ross River, Mayaro and O'nyong nyong viruses. Gene editing of mouse Mxra8 or human MXRA8 resulted in reduced levels of viral infection of cells and, reciprocally, ectopic expression of these genes resulted in increased infection...
May 16, 2018: Nature
https://www.readbyqxmd.com/read/29766242/inactivation-of-deubiquitinase-cyld-enhances-therapeutic-antibody-production-in-chinese-hamster-ovary-cells
#4
Yafang Lu, Qin Zhou, Qianqian Han, Pengfei Wu, Lanlan Zhang, Lin Zhu, David T Weaver, Changzhi Xu, Buchang Zhang
Chinese hamster ovary (CHO) cells are promising host engineering cells for industry manufacturing of therapeutic antibodies. However, cell death due to apoptosis remains a huge challenge to augment antibody production, and developing CHO cells with enhanced anti-apoptosis and proliferation ability is fundamental for cell line development and high-yielding bioprocesses. Deubiquitinase cylindromatosis (CYLD) has been proved to be a tumor suppressor by negatively regulating NF-κB and Wnt/β-catenin signaling pathways...
May 15, 2018: Applied Microbiology and Biotechnology
https://www.readbyqxmd.com/read/29756770/versatile-high-throughput-fluorescence-assay-for-monitoring-cas9-activity
#5
Kyle Seamon, Yooli K Light, Edwin Saada, Joseph S Schoeniger, Brooke Harmon
The RNA-guided DNA nuclease Cas9 is now widely used for the targeted modification of genomes of human cells and various organisms. Despite the extensive use of CRISPR systems for genome engineering and the rapid discovery and engineering of new CRISPR-associated nucleases, there are no high-throughput assays for measuring enzymatic activity. The current laboratory and future therapeutic uses of CRISPR technology have a significant risk of accidental exposure or clinical off-target effects, under-scoring the need for therapeutically-effective inhibitors of Cas9...
May 14, 2018: Analytical Chemistry
https://www.readbyqxmd.com/read/29755980/importance-of-validating-antibodies-and-small-compound-inhibitors-using-genetic-knockout-studies-t-cell-receptor-induced-cyld-phosphorylation-by-ikk%C3%AE%C2%B5-tbk1-as-a-case-study
#6
Marie Lork, Marja Kreike, Jens Staal, Rudi Beyaert
CYLD is a deubiquitinating enzyme that plays a crucial role in immunity and inflammation as a negative regulator of NF-κB transcription factor and JNK kinase signaling. Defects in either of these pathways contribute to the progression of numerous inflammatory and autoimmune disorders. Therefore, we set out to unravel molecular mechanisms that control CYLD activity in the context of T cell receptor (TCR) signaling. More specifically, we focused on CYLD phosphorylation at Ser418, which can be detected upon immunoblotting of cell extracts with phospho(Ser418)-CYLD specific antibodies...
2018: Frontiers in Cell and Developmental Biology
https://www.readbyqxmd.com/read/29748615/specific-targeting-of-point-mutations-in-egfr-l858r-positive-lung-cancer-by-crispr-cas9
#7
Alvin Ho-Kwan Cheung, Chit Chow, Jinglin Zhang, Yuhang Zhou, Tingting Huang, Kayla Ching-Kei Ng, Terry Cho-Tsun Or, Yoyo Yao Yao, Yujuan Dong, Jackie Mei-Wah Fung, Lei Xiong, Aden Ka-Yin Chan, Wai-Ming Raymond Lung, Wei Kang, Ka-Fai To
Cancer cells are defined genetically by the mutations they harbor, commonly single nucleotide substitutions. Therapeutic approaches which specifically target cancer cells by recognizing these defining genetic aberrations are expected to exhibit minimal side-effects. However, current protein-based targeted therapy is greatly limited by the range of genes that can be targeted, as well as by acquired resistance. We hypothesized that a therapeutic oligonucleotide-based strategy may address this need of specific cancer targeting...
May 10, 2018: Laboratory Investigation; a Journal of Technical Methods and Pathology
https://www.readbyqxmd.com/read/29748595/crispr-lbcpf1-prevents-choroidal-neovascularization-in-a-mouse-model-of-age-related-macular-degeneration
#8
Taeyoung Koo, Sung Wook Park, Dong Hyun Jo, Daesik Kim, Jin Hyoung Kim, Hee-Yeon Cho, Jeungeun Kim, Jeong Hun Kim, Jin-Soo Kim
LbCpf1, derived from Lachnospiraceae bacterium ND2006, is a CRISPR RNA-guided endonuclease and holds promise for therapeutic applications. Here we show that LbCpf1 can be used for therapeutic gene editing in a mouse model of age-related macular degeneration (AMD). The intravitreal delivery of LbCpf1, targeted to two angiogenesis-associated genes encoding vascular endothelial growth factor A (Vegfa) and hypoxia inducing factor 1a (Hif1a), using adeno-associated virus, led to efficient gene disruption with no apparent off-target effects in the retina and retinal pigment epithelium (RPE) cells...
May 10, 2018: Nature Communications
https://www.readbyqxmd.com/read/29740340/tissue-distribution-of-kir7-1-inwardly-rectifying-k-channel-probed-in-a-knock-in-mouse-expressing-a-haemagglutinin-tagged-protein
#9
Isabel Cornejo, Sandra Villanueva, Johanna Burgos, Karen I López-Cayuqueo, Régine Chambrey, Francisca Julio-Kalajzić, Neudo Buelvas, María I Niemeyer, Dulce Figueiras-Fierro, Peter D Brown, Francisco V Sepúlveda, L P Cid
Kir7.1 encoded by the Kcnj13 gene in the mouse is an inwardly rectifying K+ channel present in epithelia where it shares membrane localization with the Na+ /K+ -pump. Further investigations of the localisation and function of Kir7.1 would benefit from the availability of a knockout mouse, but perinatal mortality attributed to cleft palate in the neonate has thwarted this research. To facilitate localisation studies we now use CRISPR/Cas9 technology to generate a knock-in mouse, the Kir7.1-HA that expresses the channel tagged with a haemagglutinin (HA) epitope...
2018: Frontiers in Physiology
https://www.readbyqxmd.com/read/29735912/towards-resolving-the-pro-and-anti-tumor-effects-of-the-aryl-hydrocarbon-receptor
#10
Supraja Narasimhan, Elizabeth Stanford Zulick, Olga Novikov, Ashley J Parks, Jennifer J Schlezinger, Zhongyan Wang, Fabrice Laroche, Hui Feng, Francesca Mulas, Stefano Monti, David H Sherr
We have postulated that the aryl hydrocarbon receptor (AHR) drives the later, more lethal stages of some cancers when chronically activated by endogenous ligands. However, other studies have suggested that, under some circumstances, the AHR can oppose tumor aggression. Resolving this apparent contradiction is critical to the design of AHR-targeted cancer therapeutics. Molecular (siRNA, shRNA, AHR repressor, CRISPR-Cas9) and pharmacological (AHR inhibitors) approaches were used to confirm the hypothesis that AHR inhibition reduces human cancer cell invasion (irregular colony growth in 3D Matrigel cultures and Boyden chambers), migration (scratch wound assay) and metastasis (human cancer cell xenografts in zebrafish)...
May 7, 2018: International Journal of Molecular Sciences
https://www.readbyqxmd.com/read/29723654/ctcf-knockout-reveals-an-essential-role-for-this-protein-during-the-zebrafish-development
#11
Francisco Carmona-Aldana, Cecilia Zampedri, Fernando Suaste-Olmos, Adrián Murillo-de-Ozores, Georgina Guerrero, Rodrigo Arzate-Mejía, Ernesto Maldonado, Rosa Navarro, Jesús Chimal-Monroy, Félix Recillas-Targa
Chromatin regulation and organization are essential processes that regulate gene activity. The CCCTC-binding factor (CTCF) is a protein with different and important molecular functions related with chromatin dynamics. It is conserved since invertebrates to vertebrates, posing it as a factor with an important role in the physiology. In this work, we aimed to understand the distribution and functional relevance of CTCF during the embryonic development of the zebrafish (Danio rerio). We generated a zebrafish specific anti-Ctcf antibody, and found this protein to be ubiquitous, through different stages and tissues...
May 1, 2018: Mechanisms of Development
https://www.readbyqxmd.com/read/29721381/foxo1-promotes-resistance-of-non-hodgkin-lymphomas-to-anti-cd20-based-therapy
#12
Beata Pyrzynska, Michal Dwojak, Abdessamad Zerrouqi, Giulia Morlino, Piotr Zapala, Nina Miazek, Agnieszka Zagozdzon, Kamil Bojarczuk, Malgorzata Bobrowicz, Marta Siernicka, Marcin M Machnicki, Stefania Gobessi, Joanna Barankiewicz, Ewa Lech-Maranda, Dimitar G Efremov, Przemyslaw Juszczynski, Dinis Calado, Jakub Golab, Magdalena Winiarska
Diminished overall survival rate of non-Hodgkin lymphoma (NHL) patients treated with a combination regimen of rituximab, cyclophosphamide, doxorubicin, vincristine and prednisone (R-CHOP) has been recently linked to recurrent somatic mutations activating FOXO1. Despite of the clinical relevance of this finding, the molecular mechanism driving resistance to R-CHOP therapy remains largely unknown. Herein, we investigated the potential role of FOXO1 in the therapeutic efficacy of rituximab, the only targeted therapy included in the R-CHOP regimen...
2018: Oncoimmunology
https://www.readbyqxmd.com/read/29720387/microrna-focused-crispr-cas9-library-screen-reveals-fitness-associated-mirnas
#13
Jessica S Kurata, Ren-Jang Lin
MicroRNAs (miRNAs) are posttranscriptional gene regulators that play important roles in the control of cell fitness, differentiation, and development. The CRISPR-Cas9 gene-editing system is composed of the Cas9 nuclease in complex with a single guide RNA (sgRNA) and directs DNA cleavage at a predetermined site. Several CRISPR-Cas9 libraries have been constructed for genome-scale knockout screens of protein function; however few libraries have included miRNA genes. Here we constructed a miRNA-focused CRISPR-Cas9 library that targets 1,594 (85%) annotated human miRNA stem-loops...
May 2, 2018: RNA
https://www.readbyqxmd.com/read/29716628/nanog-interaction-with-the-androgen-receptor-signaling-axis-induce-ovarian-cancer-stem-cell-regulation-studies-based-on-the-crispr-cas9-system
#14
Kaijian Ling, Lupin Jiang, Shi Liang, Joseph Kwong, Leiyan Yang, Yudi Li, PingYin, Qingchun Deng, Zhiqing Liang
BACKGROUND: Ovarian cancer stem cells (OCSCs) contribute to the poor prognosis of ovarian cancer. Involvement of the androgen receptor (AR) in the malignant behaviors of other tumors has been reported. However, whether AR associates with Nanog (a stem cell marker) and participates in OCSC functions remain unclear. In this study, we investigated the interaction of Nanog with AR and examined whether this interaction induced stem-like properties in ovarian cancer cells. METHODS: AR and Nanog expression in ovarian tumors was evaluated...
May 2, 2018: Journal of Ovarian Research
https://www.readbyqxmd.com/read/29709474/keratinocyte-growth-factor-protects-endometrial-cells-from-oxygen-glucose-deprivation-re-oxygenation-via-activating-nrf2-signaling
#15
Xuting Shi, Hia-Yan Liu, Shu-Ping Li, Hong-Bin Xu
Oxygen and glucose deprivation (OGD)-re-oxygenation (OGDR) exposure to endometrial cells mimics ischemia-reperfusion injury. The present study tests the potential effect of keratinocyte growth factor (KGF) on the process. We show that KGF receptor KGFR is expressed in human endometrial T-HESC cells and primary murine endometrial cells. KGF pre-treatment protected endometrial cells from OGDR, inhibiting cell viability reduction and cell death. KGF attenuated OGDR-induced programmed necrosis in endometrial cells...
April 27, 2018: Biochemical and Biophysical Research Communications
https://www.readbyqxmd.com/read/29706942/oral-administration-of-recombinant-saccharomyces-boulardii-expressing-ovalbumin-cpe-fusion-protein-induces-antibody-response-in-mice
#16
Ghasem Bagherpour, Hosnie Ghasemi, Bahare Zand, Najmeh Zarei, Farzin Roohvand, Esmat M Ardakani, Mohammad Azizi, Vahid Khalaj
Saccharomyces boulardii , a subspecies of Saccharomyces cerevisiae , is a well-known eukaryotic probiotic with many benefits for human health. In the present study, a recombinant strain of S. boulardii was prepared to use as a potential oral vaccine delivery vehicle. In this sense, a ura3 auxotroph strain of S. boulardii CNCM I-745 (known as S. cerevisiae HANSEN CBS 5926, Yomogi® ) was generated using CRISPR/Cas9 methodology. Then a gene construct encoding a highly immunogenic protein, ovalbumin (OVA), was prepared and transformed into the ura3- S...
2018: Frontiers in Microbiology
https://www.readbyqxmd.com/read/29704372/-systematic-discovery-of-novel-prokaryotic-defense-systems-progress-and-prospects
#17
Ming Li, Fei Yue Cheng, Lu Yao Gong, Hua Xiang
Microbial defense systems provide fundamental elements for biotechnology development. For example, molecular cloning techniques greatly depend on the discovery of the bacterial restriction-modification system, and similarly, deciphering the CRISPR adaptive immunity gives rise to the revolutionary genome editing technique CRISPR-Cas9. Therefore, exploring novel microbial defense systems has attracted world-wide attention, and new systems like prokaryotic Argonautes (pAgos) and the defense island system associated with restriction-modification (DISARM) were successively discovered and investigated...
April 20, 2018: Yi Chuan, Hereditas
https://www.readbyqxmd.com/read/29694447/pharmacological-versus-genetic-inhibition-of-heme-oxygenase-1-the-comparison-of-metalloporphyrins-shrna-and-crispr-cas9-system
#18
Olga Mucha, Paulina Podkalicka, Maria Czarnek, Anna Biela, Mateusz Mieczkowski, Neli Kachamakova-Trojanowska, Jacek Stepniewski, Alicja Jozkowicz, Jozef Dulak, Agnieszka Loboda
Inhibition of heme oxygenase-1 (HO-1, encoded by HMOX1), a cytoprotective, anti-apoptotic and anti-inflammatory enzyme, may serve as a valuable therapy in various pathophysiological processes, including tumorigenesis. We compared the effect of chemical inhibitors - metalloporphyrins, with genetic tools - shRNA and CRISPR/Cas9 systems, to knock-down (KD)/knock-out (KO) HO-1 expression/activity. 293T cells were incubated with metalloporphyrins, tin and zinc protoporphyrins (SnPPIX and ZnPPIX, respectively) or were either transduced with lentiviral vectors encoding different shRNA sequences against HO-1 or were modified by CRISPR/Cas9 system targeting HMOX1...
April 25, 2018: Acta Biochimica Polonica
https://www.readbyqxmd.com/read/29693372/biosynthesis-of-the-15-membered-ring-depsipeptide-neoantimycin
#19
Will Skyrud, Joyce Liu, Divya Thankachan, Maria Cabrera, Ryan F Seipke, Wenjun Zhang
Antimycins are a family of natural products possessing outstanding biological activities and unique structures, which have intrigued chemists for over a half century. Of particular interest are the ring-expanded antimycins that show promising anti-cancer potential and whose biosynthesis remains uncharacterized. Specifically, neoantimycin and its analogs have been shown to be effective regulators of the oncogenic proteins GRP78/BiP and K-Ras. The neoantimycin structural skeleton is built on a 15-membered tetralactone ring containing one methyl, one hydroxy, one benzyl and three alkylmoieties, as well as an amide linkage to a conserved 3-formamidosalicylic acid moiety...
April 25, 2018: ACS Chemical Biology
https://www.readbyqxmd.com/read/29691200/crispr-cas9-genome-editing-fueling-the-revolution-in-cancer-immunotherapy
#20
REVIEW
Xiaojun Liu, Yangbing Zhao
The development of genomic editing technologies expands the landscape of T cell engineering for adoptive cell therapy. Among the multiple tools that can be used, CRISPR/Cas9 has been shown to be relatively easy to use, simple to design and cost effective with highly efficient multiplex genome engineering capabilities. Allogeneic universal chimeric antigen receptor (CAR) T cells can be produced by disrupting T cell receptor (TCR) and beta-2-microglobulin (B2M) in CAR T cells or by directly knocking in a CAR at the disrupted TRAC locus...
April 21, 2018: Current Research in Translational Medicine
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