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https://www.readbyqxmd.com/read/29925004/full-functional-knockout-of-placental-growth-factor-by-knockin-with-an-inactive-variant-able-to-heterodimerize-with-vegf-a
#1
Ivana Apicella, Valeria Cicatiello, Dario Acampora, Valeria Tarallo, Sandro De Falco
Placental growth factor (PlGF) is a proangiogenic member of the vascular endothelial growth factor (VEGF) family playing a central role in pathological angiogenesis. PlGF-DE is a PlGF variant unable to bind vascular endothelial growth factor receptor 1 (VEGFR-1) but still able to generate heterodimer with VEGF-A. We have generated PlGF-DE knockin mice that are vital and fertile and show unaltered expression of Plgf, Vegf-a, Vegfr-1, and Vegfr-2 compared with wild-type mice. Interestingly, these mutants showed additional and remarkable angiogenesis impairment in tumor growth, hindlimb ischemia, and choroidal neovascularization compared with both PlGF knockout and wild-type mice...
June 19, 2018: Cell Reports
https://www.readbyqxmd.com/read/29905864/pathogenic-variant-in-ephb4-results-in-central-conducting-lymphatic-anomaly
#2
Dong Li, Tara L Wenger, Christoph Seiler, Michael E March, Alvaro Gutierrez-Uzquiza, Charlly Kao, Elizabeth Bhoj, Lifeng Tian, Misha Rosenbach, Yichuan Liu, Nora Robinson, Mechenzie Behr, Rosetta Chiavacci, Cuiping Hou, Tiancheng Wang, Marina Bakay, Renata Pellegrino da Silva, Jonathan A Perkins, Patrick Sleiman, Michael A Levine, Patricia J Hicks, Maxim Itkin, Yoav Dori, Hakon Hakonarson
Central conducting lymphatic anomaly (CCLA) is one of the complex lymphatic anomalies characterized by dilated lymphatic channels, lymphatic channel dysmotility, and distal obstruction affecting lymphatic drainage. We performed whole exome sequencing (WES) of DNA from a four-generation pedigree and examined the consequences of the variant by transfection of mammalian cells and morpholino and rescue studies in zebrafish. WES revealed a heterozygous mutation in EPHB4 (RefSeq NM_004444.4; c.2334 + 1G>C) and RNA-Seq demonstrated that the EPHB4 mutation destroys the normal donor site, which leads to the use of a cryptic splice donor that results in retention of the intervening 12-bp intron sequence...
June 14, 2018: Human Molecular Genetics
https://www.readbyqxmd.com/read/29898956/congenital-macrothrombocytopenia-with-focal-myelofibrosis-due-to-mutations-in-human-g6b-b-is-rescued-in-humanized-mice
#3
Inga Hofmann, Mitchell J Geer, Timo Vögtle, Andrew Crispin, Dean R Campagna, Alastair Barr, Monica L Calicchio, Silke Heising, Johanna P van Geffen, Marijke J E Kuijpers, Johan W M Heemskerk, Johannes A Eble, Klaus Schmitz-Abe, Esther A Obeng, Michael Douglas, Kathleen Freson, Corinne Pondarré, Rémi Favier, Gavin E Jarvis, Kyriacos Markianos, Ernest Turro, Willem H Ouwehand, Alexandra Mazharian, Mark D Fleming, Yotis A Senis
Unlike primary myelofibrosis (PMF) in adults, myelofibrosis in children is rare. Congenital (inherited) forms of myelofibrosis (cMF) have been described, but the underlying genetic mechanisms remain elusive. Here we describe 4 families with autosomal recessive inherited macrothrombocytopenia with focal myelofibrosis due to germline loss-of-function mutations in the megakaryocyte-specific immunoreceptor tyrosine-based inhibitory motif (ITIM)-containing receptor G6b-B (G6b, C6orf25 or MPIG6B). Patients presented with a mild-to-moderate bleeding diathesis, macrothrombocytopenia, anemia, leukocytosis and atypical megakaryocytes associated with a distinctive, focal, perimegakaryocytic pattern of bone marrow fibrosis...
June 13, 2018: Blood
https://www.readbyqxmd.com/read/29898399/autophagy-differentially-regulates-insulin-production-and-insulin-sensitivity
#4
Soh Yamamoto, Kenta Kuramoto, Nan Wang, Xiaolei Situ, Medha Priyadarshini, Weiran Zhang, Jose Cordoba-Chacon, Brian T Layden, Congcong He
Autophagy, a stress-induced lysosomal degradative pathway, has been assumed to exert similar metabolic effects in different organs. Here, we establish a model where autophagy plays different roles in insulin-producing β cells versus insulin-responsive cells, utilizing knockin (Becn1F121A ) mice manifesting constitutively active autophagy. With a high-fat-diet challenge, the autophagy-hyperactive mice unexpectedly show impaired glucose tolerance, but improved insulin sensitivity, compared to mice with normal autophagy...
June 12, 2018: Cell Reports
https://www.readbyqxmd.com/read/29898394/variable-satb1-levels-regulate-hematopoietic-stem-cell-heterogeneity-with-distinct-lineage-fate
#5
Yukiko Doi, Takafumi Yokota, Yusuke Satoh, Daisuke Okuzaki, Masahiro Tokunaga, Tomohiko Ishibashi, Takao Sudo, Tomoaki Ueda, Yasuhiro Shingai, Michiko Ichii, Akira Tanimura, Sachiko Ezoe, Hirohiko Shibayama, Terumi Kohwi-Shigematsu, Junji Takeda, Kenji Oritani, Yuzuru Kanakura
Hematopoietic stem cells (HSCs) comprise a heterogeneous population exhibiting self-renewal and differentiation capabilities; however, the mechanisms involved in maintaining this heterogeneity remain unclear. Here, we show that SATB1 is involved in regulating HSC heterogeneity. Results in conditional Satb1-knockout mice revealed that SATB1 was important for the self-renewal and lymphopoiesis of adult HSCs. Additionally, HSCs from Satb1/Tomato-knockin reporter mice were classified based on SATB1/Tomato intensity, with transplantation experiments revealing stronger differentiation toward the lymphocytic lineage along with high SATB1 levels, whereas SATB1- HSCs followed the myeloid lineage in agreement with genome-wide transcription and cell culture studies...
June 12, 2018: Cell Reports
https://www.readbyqxmd.com/read/29888643/rapid-response-and-slow-recovery-of-the-h3k4me3-epigenomic-marker-in-the-liver-after-light-mediated-phase-advances-of-the-circadian-clock
#6
Dmytro Grygoryev, Michael R Rountree, Furaha Rwatambuga, Anna Ohlrich, Ayaka Kukino, Matthew P Butler, Charles N Allen, Mitchell S Turker
Mammalian tissues display circadian rhythms in transcription, translation, and histone modifications. Here we asked how an advance of the light-dark cycle alters daily rhythms in the liver epigenome at the H3K4me3 (trimethylation of lysine 4 on histone 3) modification, which is found at active and poised gene promoters. H3K4me3 levels were first measured at 4 time points (zeitgeber time [ZT] 3, 8, 15, and 20) during a normal 12L:12D light-dark cycle. Peak levels were observed during the early dark phase at ZT15 and dropped to low levels around lights-on (ZT0) between ZT20 and ZT3...
June 1, 2018: Journal of Biological Rhythms
https://www.readbyqxmd.com/read/29887318/paneth-cell-multipotency-induced-by-notch-activation-following-injury
#7
Shiyan Yu, Kevin Tong, Yanlin Zhao, Iyshwarya Balasubramanian, George S Yap, Ronaldo P Ferraris, Edward M Bonder, Michael P Verzi, Nan Gao
Paneth cells are post-mitotic intestinal epithelial cells supporting the stem cell niche and mucosal immunity. Paneth cell pathologies are observed in various gastrointestinal diseases, but their plasticity and response to genomic and environmental challenges remain unclear. Using a knockin allele engineered at the mouse Lyz1 locus, we performed detailed Paneth cell-lineage tracing. Irradiation induced a subset of Paneth cells to proliferate and differentiate into villus epithelial cells. RNA sequencing (RNA-seq) revealed that Paneth cells sorted from irradiated mice acquired a stem cell-like transcriptome; when cultured in vitro, these individual Paneth cells formed organoids...
May 23, 2018: Cell Stem Cell
https://www.readbyqxmd.com/read/29887311/no-blokes-is-essential-for-male-viability-and-x-chromosome-gene-expression-in-the-australian-sheep-blowfly
#8
Rebecca J Davis, Esther J Belikoff, Elizabeth H Scholl, Fang Li, Maxwell J Scott
It has been hypothesized that the Drosophila 4th  chromosome is derived from an ancient X chromosome [1]. In the Australian sheep blowfly, Lucilia cuprina, the heterochromatic X chromosome contains few active genes and orthologs of Drosophila X-linked genes are autosomal. Of 8 X-linked genes identified previously in L. cuprina, 6 were orthologs of Drosophila 4th -chromosome genes [2]. The X-linked genes were expressed equally in males and females. Here we identify an additional 51 X-linked genes and show that most are dosage compensated...
May 29, 2018: Current Biology: CB
https://www.readbyqxmd.com/read/29862665/targeted-nucleotide-editing-technologies-for-microbial-metabolic-engineering
#9
REVIEW
Takayuki Arazoe, Akihiko Kondo, Keiji Nishida
Since the emergence of programmable RNA-guided nucleases based on Clustered regularly interspaced short palindromic repeats (CRISPR)/ CRISPR-associated protein (Cas) systems, genome editing technologies have become a simplified and versatile tool for genome editing in various organisms and cell types. Although genome editing enables efficient genome manipulations, such as gene disruptions, gene knockins, and chromosomal translocations via DNA double-strand break (DSB) repair in eukaryotes, DSBs induced by the CRISPR/Cas system are lethal or severely toxic to many microorganisms...
June 3, 2018: Biotechnology Journal
https://www.readbyqxmd.com/read/29861284/the-temporal-dynamics-of-arc-expression-regulate-cognitive-flexibility
#10
Mark J Wall, Dawn R Collins, Samantha L Chery, Zachary D Allen, Elissa D Pastuzyn, Arlene J George, Viktoriya D Nikolova, Sheryl S Moy, Benjamin D Philpot, Jason D Shepherd, Jürgen Müller, Michael D Ehlers, Angela M Mabb, Sonia A L Corrêa
Neuronal activity regulates the transcription and translation of the immediate-early gene Arc/Arg3.1, a key mediator of synaptic plasticity. Proteasome-dependent degradation of Arc tightly limits its temporal expression, yet the significance of this regulation remains unknown. We disrupted the temporal control of Arc degradation by creating an Arc knockin mouse (ArcKR) where the predominant Arc ubiquitination sites were mutated. ArcKR mice had intact spatial learning but showed specific deficits in selecting an optimal strategy during reversal learning...
May 24, 2018: Neuron
https://www.readbyqxmd.com/read/29858049/generation-of-hutat2-fc-knockin-primary-human-monocytes-using-crispr-cas9
#11
Bowen Wang, Jiahui Zuo, Wenzhen Kang, Qianqi Wei, Jianhui Li, Chunfu Wang, Zhihui Liu, Yuanan Lu, Yan Zhuang, Bianli Dang, Qing Liu, Wen Kang, Yongtao Sun
The ability of monocytes to travel through the bloodstream, traverse tissue barriers, and aggregate at disease sites endows these cells with the attractive potential to carry therapeutic genes into the nervous system. However, gene editing in primary human monocytes has long been a challenge. Here, we applied the CRISPR/Cas9 system to deliver the large functional Hutat2:Fc DNA fragment into the genome of primary monocytes to neutralize HIV-1 transactivator of transcription (Tat), an essential neurotoxic factor that causes HIV-associated neurocognitive disorder (HAND) in the nervous system...
June 1, 2018: Molecular Therapy. Nucleic Acids
https://www.readbyqxmd.com/read/29849099/the-c-ebp%C3%AE-lip-isoform-rescues-loss-of-c-ebp%C3%AE-function-in-the-mouse
#12
Valérie Bégay, Christian Baumeier, Karin Zimmermann, Arnd Heuser, Achim Leutz
The transcription factor C/EBPβ regulates hematopoiesis, bone, liver, fat, and skin homeostasis, and female reproduction. C/EBPβ protein expression from its single transcript occurs by alternative in-frame translation initiation at consecutive start sites to generate three isoforms, two long (LAP*, LAP) and one truncated (LIP), with the same C-terminal bZip dimerization domain. The long C/EBPβ isoforms are considered gene activators, whereas the LIP isoform reportedly acts as a dominant-negative repressor...
May 30, 2018: Scientific Reports
https://www.readbyqxmd.com/read/29805302/dna-damage-checkpoint-pathway-modulates-the-regulation-of-skeletal-growth-and-osteoblastic-bone-formation-by-parathyroid-hormone-related-peptide
#13
Ying Zhang, Guangpei Chen, Zhen Gu, Haijian Sun, Andrew Karaplis, David Goltzman, Dengshun Miao
We previously demonstrated that parathyroid hormone-related peptide (PTHrP) 1-84 knockin ( Pthrp KI) mice, which lacked a PTHrP nuclear localization sequence (NLS) and C-terminus, displayed early senescence, defective osteoblastic bone formation, and skeletal growth retardation. However, the mechanism of action of the PTHrP NLS and C-terminus in regulating development of skeleton is still unclear. In this study, we examined alterations of oxidative stress and DNA damage response-related molecules in Pthrp KI skeletal tissue...
2018: International Journal of Biological Sciences
https://www.readbyqxmd.com/read/29787711/tild-crispr-allows-for-efficient-and-precise-gene-knockin-in-mouse-and-human-cells
#14
Xuan Yao, Meiling Zhang, Xing Wang, Wenqin Ying, Xinde Hu, Pengfei Dai, Feilong Meng, Linyu Shi, Yun Sun, Ning Yao, Wanxia Zhong, Yun Li, Keliang Wu, Weiping Li, Zi-Jiang Chen, Hui Yang
The targeting efficiency of knockin sequences via homologous recombination (HR) is generally low. Here we describe a method we call Tild-CRISPR (targeted integration with linearized dsDNA-CRISPR), a targeting strategy in which a PCR-amplified or precisely enzyme-cut transgene donor with 800-bp homology arms is injected with Cas9 mRNA and single guide RNA into mouse zygotes. Compared with existing targeting strategies, this method achieved much higher knockin efficiency in mouse embryos, as well as brain tissue...
May 21, 2018: Developmental Cell
https://www.readbyqxmd.com/read/29786791/protocols-for-studies-on-tmprss2-erg-in-prostate-cancer
#15
Hubert Pakula, Douglas E Linn, Daniel R Schmidt, Marit Van Gorsel, Matthew G Vander Heiden, Zhe Li
TMPRSS2/ERG is the most common type of gene fusions found in human prostate cancer. There are two important features of TMPRSS2/ERG fusions. One is that these gene fusions lead to ectopic expression of ERG, an ETS family transcription factor, in prostate epithelial cells from the 5' control region of an androgen/estrogen dual-responsive gene, TMPRSS2; the other is that ~60% of these fusions are generated via intrachromosomal deletion of the interstitial region between TMPRSS2 and ERG. To recapitulate these important aspects of TMPRSS2/ERG fusions, we generated several TMPRSS2/ERG knockin mouse models based on the endogenous Tmprss2 locus...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29780003/a-disease-associated-aifm1-variant-induces-severe-myopathy-in-knockin-mice
#16
Lena Wischhof, Anna Gioran, Dagmar Sonntag-Bensch, Antonia Piazzesi, Miriam Stork, Pierluigi Nicotera, Daniele Bano
OBJECTIVE: Mutations in the AIFM1 gene have been identified in recessive X-linked mitochondrial diseases. Functional and molecular consequences of these pathogenic AIFM1 mutations have been poorly studied in vivo. METHODS/RESULTS: Here we provide evidence that the disease-associated apoptosis-inducing factor (AIF) deletion arginine 201 (R200 in rodents) causes pathology in knockin mice. Within a few months, posttranslational loss of the mutant AIF protein induces severe myopathy associated with a lower number of cytochrome c oxidase-positive muscle fibers...
May 8, 2018: Molecular Metabolism
https://www.readbyqxmd.com/read/29779223/gene-editing-vectors-for-studying-nicotinic-acetylcholine-receptors-in-cholinergic-transmission
#17
Can Peng, Yijin Yan, Veronica J Kim, Staci E Engle, Jennifer N Berry, J Michael McIntosh, Rachael L Neve, Ryan M Drenan
Nicotinic acetylcholine receptors (nAChRs), prototype members of the cys-loop ligand gated ion channel family, are key mediators of cholinergic transmission in the central nervous system. Despite their importance, technical gaps exist in our ability to dissect the function of individual subunits in the brain. To overcome these barriers, we designed CRISPR/Cas9 small guide RNA sequences (sgRNAs) for production of loss-of-function alleles in mouse nAChR genes. These sgRNAs were validated in vitro via deep sequencing...
May 19, 2018: European Journal of Neuroscience
https://www.readbyqxmd.com/read/29754960/high-level-precise-knockin-of-ipscs-by-simultaneous-reprogramming-and-genome-editing-of-human-peripheral-blood-mononuclear-cells
#18
Wei Wen, Xinxin Cheng, Yawen Fu, Feiying Meng, Jian-Ping Zhang, Lu Zhang, Xiao-Lan Li, Zhixue Yang, Jing Xu, Feng Zhang, Gary D Botimer, Weiping Yuan, Changkai Sun, Tao Cheng, Xiao-Bing Zhang
We have developed an improved episomal vector system for efficient generation of integration-free induced pluripotent stem cells (iPSCs) from peripheral blood mononuclear cells. More recently, we reported that the use of an optimized CRISPR-Cas9 system together with a double-cut donor increases homology-directed repair-mediated precise gene knockin efficiency by 5- to 10-fold. Here, we report the integration of blood cell reprogramming and genome editing in a single step. We found that expression of Cas9 and KLF4 using a single vector significantly increases genome editing efficiency, and addition of SV40LT further enhances knockin efficiency...
May 4, 2018: Stem Cell Reports
https://www.readbyqxmd.com/read/29733298/snasp-inhibits-tlr-signaling-to-regulate-immune-response-in-sepsis
#19
Feng-Ming Yang, Yong Zuo, Wei Zhou, Chuan Xia, Bumsuk Hahm, Mark Sullivan, Jinke Cheng, Hui-Ming Chang, Edward Th Yeh
Many Toll-like receptors (TLRs) signal through TNF receptor-associated factor 6 (TRAF6) to activate innate immune responses. Here, we show that somatic nuclear autoantigenic sperm protein (sNASP) binds to TRAF6 to prevent TRAF6 autoubiquitination in unstimulated macrophages. Following LPS stimulation, a complex consisting of sNASP, TRAF6, IRAK4, and casein kinase 2 (CK2) is formed. CK2 phosphorylates sNASP at serine 158, allowing sNASP to dissociate from TRAF6. Free TRAF6 is then autoubiquitinated, followed by activation of downstream signaling pathways...
May 7, 2018: Journal of Clinical Investigation
https://www.readbyqxmd.com/read/29729423/interaction-between-a-mapt-variant-causing-frontotemporal-dementia-and-mutant-app-affects-axonal-transport
#20
Robert Adalbert, Stefan Milde, Claire Durrant, Kunie Ando, Virginie Stygelbout, Zehra Yilmaz, Stacey Gould, Jean-Pierre Brion, Michael P Coleman
In Alzheimer's disease, many indicators point to a central role for poor axonal transport, but the potential for stimulating axonal transport to alleviate the disease remains largely untested. Previously, we reported enhanced anterograde axonal transport of mitochondria in 8- to 11-month-old MAPTP301L knockin mice, a genetic model of frontotemporal dementia with parkinsonism-17T. In this study, we further characterized the axonal transport of mitochondria in younger MAPTP301L mice crossed with the familial Alzheimer's disease model, TgCRND8, aiming to test whether boosting axonal transport in young TgCRND8 mice can alleviate axonal swelling...
April 5, 2018: Neurobiology of Aging
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