Mouse Chd8 Knockout Project (CRISPR/Cas9)
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Human Placenta Exosomes: Biogenesis, Isolation, Composition, and Prospects for Use in Diagnostics
International Journal of Molecular Sciences Review Human Placenta Exosomes: Biogenesis, Isolation, Composition, and Prospects for Use in Diagnostics Evgeniya E. Burkova 1,* , Sergey E. Sedykh 1,2 and Georgy A. Nevinsky 1,2 1 SB RAS Institute of Chemical Biology and Fundamental Medicine, 630090 Novosibirsk, Russia; [email protected] (S.E.S.); [email protected] (G.A.N.) 2 Department of Natural Sciences, Novosibirsk State University, 630090 Novosibirsk, Russia * Correspondence: [email protected]; Tel.: +7-(383)-363-51-27 Abstract: Exosomes are 40–100 nm nanovesicles participating in intercellular communication and transferring various bioactive proteins, mRNAs, miRNAs, and lipids. During pregnancy, the placenta releases exosomes into the maternal circulation. Placental exosomes are detected in the maternal blood even in the first trimester of pregnancy and their numbers increase significantly by the end of pregnancy. Exosomes are necessary for the normal functioning of the placenta and fetal development. Effects of exosomes on target cells depend not only on their concentration but also on their intrinsic components. The biochemical composition of the placental exosomes may cause various complications of pregnancy. Some studies relate the changes in the composition of nanovesicles to placental dysfunction. Isolation of placental exosomes from the blood of pregnant women and the study of protein, lipid, and nucleic composition can lead to the development of methods for early diagnosis of pregnancy pathologies. This review describes the biogenesis of exosomes, methods Citation: Burkova, E.E.; Sedykh, S.E.; of their isolation, analyzes their biochemical composition, and considers the prospects for using Nevinsky, G.A. Human Placenta exosomes to diagnose pregnancy pathologies. -
Supporting Information
Supporting Information Edgar et al. 10.1073/pnas.1601895113 SI Methods (Actimetrics), and recordings were analyzed using LumiCycle Mice. Sample size was determined using the resource equation: Data Analysis software (Actimetrics). E (degrees of freedom in ANOVA) = (total number of exper- – Cell Cycle Analysis of Confluent Cell Monolayers. NIH 3T3, primary imental animals) (number of experimental groups), with −/− sample size adhering to the condition 10 < E < 20. For com- WT, and Bmal1 fibroblasts were sequentially transduced − − parison of MuHV-4 and HSV-1 infection in WT vs. Bmal1 / with lentiviral fluorescent ubiquitin-based cell cycle indicators mice at ZT7 (Fig. 2), the investigator did not know the genotype (FUCCI) mCherry::Cdt1 and amCyan::Geminin reporters (32). of the animals when conducting infections, bioluminescence Dual reporter-positive cells were selected by FACS (Influx Cell imaging, and quantification. For bioluminescence imaging, Sorter; BD Biosciences) and seeded onto 35-mm dishes for mice were injected intraperitoneally with endotoxin-free lucif- subsequent analysis. To confirm that expression of mCherry:: Cdt1 and amCyan::Geminin correspond to G1 (2n DNA con- erin (Promega E6552) using 2 mg total per mouse. Following < ≤ anesthesia with isofluorane, they were scanned with an IVIS tent) and S/G2 (2 n 4 DNA content) cell cycle phases, Lumina (Caliper Life Sciences), 15 min after luciferin admin- respectively, cells were stained with DNA dye DRAQ5 (abcam) and analyzed by flow cytometry (LSR-Fortessa; BD Biosci- istration. Signal intensity was quantified using Living Image ences). To examine dynamics of replicative activity under ex- software (Caliper Life Sciences), obtaining maximum radiance perimental confluent conditions, synchronized FUCCI reporter for designated regions of interest (photons per second per − − − monolayers were observed by time-lapse live cell imaging over square centimeter per Steradian: photons·s 1·cm 2·sr 1), relative 3 d (Nikon Eclipse Ti-E inverted epifluorescent microscope). -
A Chemical Proteomic Approach to Investigate Rab Prenylation in Living Systems
A chemical proteomic approach to investigate Rab prenylation in living systems By Alexandra Fay Helen Berry A thesis submitted to Imperial College London in candidature for the degree of Doctor of Philosophy of Imperial College. Department of Chemistry Imperial College London Exhibition Road London SW7 2AZ August 2012 Declaration of Originality I, Alexandra Fay Helen Berry, hereby declare that this thesis, and all the work presented in it, is my own and that it has been generated by me as the result of my own original research, unless otherwise stated. 2 Abstract Protein prenylation is an important post-translational modification that occurs in all eukaryotes; defects in the prenylation machinery can lead to toxicity or pathogenesis. Prenylation is the modification of a protein with a farnesyl or geranylgeranyl isoprenoid, and it facilitates protein- membrane and protein-protein interactions. Proteins of the Ras superfamily of small GTPases are almost all prenylated and of these the Rab family of proteins forms the largest group. Rab proteins are geranylgeranylated with up to two geranylgeranyl groups by the enzyme Rab geranylgeranyltransferase (RGGT). Prenylation of Rabs allows them to locate to the correct intracellular membranes and carry out their roles in vesicle trafficking. Traditional methods for probing prenylation involve the use of tritiated geranylgeranyl pyrophosphate which is hazardous, has lengthy detection times, and is insufficiently sensitive. The work described in this thesis developed systems for labelling Rabs and other geranylgeranylated proteins using a technique known as tagging-by-substrate, enabling rapid analysis of defective Rab prenylation in cells and tissues. An azide analogue of the geranylgeranyl pyrophosphate substrate of RGGT (AzGGpp) was applied for in vitro prenylation of Rabs by recombinant enzyme. -
Literature Mining Sustains and Enhances Knowledge Discovery from Omic Studies
LITERATURE MINING SUSTAINS AND ENHANCES KNOWLEDGE DISCOVERY FROM OMIC STUDIES by Rick Matthew Jordan B.S. Biology, University of Pittsburgh, 1996 M.S. Molecular Biology/Biotechnology, East Carolina University, 2001 M.S. Biomedical Informatics, University of Pittsburgh, 2005 Submitted to the Graduate Faculty of School of Medicine in partial fulfillment of the requirements for the degree of Doctor of Philosophy University of Pittsburgh 2016 UNIVERSITY OF PITTSBURGH SCHOOL OF MEDICINE This dissertation was presented by Rick Matthew Jordan It was defended on December 2, 2015 and approved by Shyam Visweswaran, M.D., Ph.D., Associate Professor Rebecca Jacobson, M.D., M.S., Professor Songjian Lu, Ph.D., Assistant Professor Dissertation Advisor: Vanathi Gopalakrishnan, Ph.D., Associate Professor ii Copyright © by Rick Matthew Jordan 2016 iii LITERATURE MINING SUSTAINS AND ENHANCES KNOWLEDGE DISCOVERY FROM OMIC STUDIES Rick Matthew Jordan, M.S. University of Pittsburgh, 2016 Genomic, proteomic and other experimentally generated data from studies of biological systems aiming to discover disease biomarkers are currently analyzed without sufficient supporting evidence from the literature due to complexities associated with automated processing. Extracting prior knowledge about markers associated with biological sample types and disease states from the literature is tedious, and little research has been performed to understand how to use this knowledge to inform the generation of classification models from ‘omic’ data. Using pathway analysis methods to better understand the underlying biology of complex diseases such as breast and lung cancers is state-of-the-art. However, the problem of how to combine literature- mining evidence with pathway analysis evidence is an open problem in biomedical informatics research. -
Human Brain Organoids Reveal Accelerated Development of Cortical Neuron Classes As a Shared Feature of Autism Risk Genes
bioRxiv preprint doi: https://doi.org/10.1101/2020.11.10.376509; this version posted November 12, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Human brain organoids reveal accelerated development of cortical neuron classes as a shared feature of autism risk genes Bruna Paulsen1,2,†, Silvia Velasco1,2,†,#, Amanda J. Kedaigle1,2,3,†, Martina Pigoni1,2,†, Giorgia Quadrato4,5 Anthony Deo2,6,7,8, Xian Adiconis2,3, Ana Uzquiano1,2, Kwanho Kim1,2,3, Sean K. Simmons2,3, Kalliopi Tsafou2, Alex Albanese9, Rafaela Sartore1,2, Catherine Abbate1,2, Ashley Tucewicz1,2, Samantha Smith1,2, Kwanghun Chung9,10,11,12, Kasper Lage2,13, Aviv Regev3,14, Joshua Z. Levin2,3, Paola Arlotta1,2,# † These authors contributed equally to the work # Correspondence should be addressed to [email protected] and [email protected] 1 Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138, USA 2 Stanley Center for Psychiatric Research, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA 3 Klarman Cell Observatory, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA 4 Department of Stem Cell Biology and Regenerative Medicine, Keck School of Medicine, University of Southern California, Los Angeles, CA 90033, USA; 5 Eli and Edythe Broad CIRM Center for Regenerative Medicine and Stem Cell Research at the University of Southern California, Los Angeles, CA 90033, USA. 6 Department of Psychiatry, -
Chd8 Mutation Leads to Autistic-Like Behaviors and Impaired Striatal Circuits
Chd8 Mutation Leads to Autistic-like Behaviors and Impaired Striatal Circuits The MIT Faculty has made this article openly available. Please share how this access benefits you. Your story matters. Citation Platt, Randall J.; Zhou, Yang; Slaymaker, Ian M.; Shetty, Ashwin S.; Weisbach, Niels R.; Kim, Jin-Ah; Sharma, Jitendra et al. “Chd8 Mutation Leads to Autistic-Like Behaviors and Impaired Striatal Circuits.” Cell Reports 19, no. 2 (April 2017): 335–350 © 2017 The Authors As Published http://dx.doi.org/10.1016/j.celrep.2017.03.052 Publisher Elsevier Version Final published version Citable link http://hdl.handle.net/1721.1/109830 Terms of Use Creative Commons Attribution-NonCommercial-NoDerivs License Detailed Terms http://creativecommons.org/licenses/by-nc-nd/4.0/ Article Chd8 Mutation Leads to Autistic-like Behaviors and Impaired Striatal Circuits Graphical Abstract Authors Randall J. Platt, Yang Zhou, Ian M. Slaymaker, ..., Gerald R. Crabtree, Guoping Feng, Feng Zhang Correspondence [email protected] (R.J.P.), [email protected] (F.Z.) Macrocephaly Gene dysregulation In Brief Platt et al. demonstrate that loss-of- +/– Chd8 function mutation of the high-confidence ASD-associated gene Chd8 results in behavioral and synaptic defects in mice. ASD-like behavior Synaptic dysfunction Highlights À d Chd8+/ mice display macrocephaly and ASD-like behaviors d Major insult to genome regulation and cellular processes in Chd8+/– mice d LOF Chd8 mutations cause synaptic dysfunction in the nucleus accumbens d NAc-specific perturbation of Chd8 in adult Cas9 mice recapitulates behavior Platt et al., 2017, Cell Reports 19, 335–350 April 11, 2017 ª 2017 The Authors. -
Mutations and Modeling of the Chromatin Remodeler CHD8 Define an Emerging Autism Etiology
REVIEW published: 17 December 2015 doi: 10.3389/fnins.2015.00477 Mutations and Modeling of the Chromatin Remodeler CHD8 Define an Emerging Autism Etiology Rebecca A. Barnard 1, Matthew B. Pomaville 1, 2 and Brian J. O’Roak 1* 1 Department of Molecular & Medical Genetics, Oregon Health & Science University, Portland, OR, USA, 2 Department of Biology, California State University, Fresno, CA, USA Autism Spectrum Disorder (ASD) is a common neurodevelopmental disorder with a strong but complex genetic component. Recent family based exome-sequencing strategies have identified recurrent de novo mutations at specific genes, providing strong evidence for ASD risk, but also highlighting the extreme genetic heterogeneity of the disorder. However, disruptions in these genes converge on key molecular pathways early in development. In particular, functional enrichment analyses have found that there is a bias toward genes involved in transcriptional regulation, such as chromatin modifiers. Here we review recent genetic, animal model, co-expression network, and functional genomics studies relating to the high confidence ASD risk gene, CHD8. CHD8, a chromatin remodeling factor, may serve as a “master regulator” of a common ASD etiology. Individuals with a CHD8 mutation show an ASD subtype that includes similar Edited by: physical characteristics, such as macrocephaly and prolonged GI problems including Gul Dolen, recurrent constipation. Similarly, animal models of CHD8 disruption exhibit enlarged Johns Hopkins University, USA head circumference and reduced gut motility phenotypes. Systems biology approaches Reviewed by: Jeffrey Varner, suggest CHD8 and other candidate ASD risk genes are enriched during mid-fetal Cornell University, USA development, which may represent a critical time window in ASD etiology. -
Brain-Specific Functional Relationship Networks Inform Autism Spectrum
Duda et al. Translational Psychiatry (2018) 8:56 DOI 10.1038/s41398-018-0098-6 Translational Psychiatry ARTICLE Open Access Brain-specific functional relationship networks inform autism spectrum disorder gene prediction Marlena Duda 1, Hongjiu Zhang1, Hong-Dong Li1,2,DennisP.Wall 3,4,MargitBurmeister1,5,6,7 andYuanfangGuan1,8,9 Abstract Autism spectrum disorder (ASD) is a neuropsychiatric disorder with strong evidence of genetic contribution, and increased research efforts have resulted in an ever-growing list of ASD candidate genes. However, only a fraction of the hundreds of nominated ASD-related genes have identified de novo or transmitted loss of function (LOF) mutations that can be directly attributed to the disorder. For this reason, a means of prioritizing candidate genes for ASD would help filter out false-positive results and allow researchers to focus on genes that are more likely to be causative. Here we constructed a machine learning model by leveraging a brain-specific functional relationship network (FRN) of genes to produce a genome-wide ranking of ASD risk genes. We rigorously validated our gene ranking using results from two independent sequencing experiments, together representing over 5000 simplex and multiplex ASD families. Finally, through functional enrichment analysis on our highly prioritized candidate gene network, we identified a small number of pathways that are key in early neural development, providing further support for their potential role in ASD. 1234567890():,; 1234567890():,; Introduction genomic variants for ASD. These studies, which primarily The term autism spectrum disorder (ASD) describes a focus on de novo loss-of-function (LOF) mutations, have range of complex neurodevelopmental phenotypes found significant associations with autism in anywhere marked by impaired social interaction and communica- from 65 to 150 genes; however, statistical genetic models tion skills along with restricted interests and repetitive estimate the total number of genes that contribute to ASD behaviors1. -
RAB2B (Human) Recombinant Protein (P01)
Produktinformation Diagnostik & molekulare Diagnostik Laborgeräte & Service Zellkultur & Verbrauchsmaterial Forschungsprodukte & Biochemikalien Weitere Information auf den folgenden Seiten! See the following pages for more information! Lieferung & Zahlungsart Lieferung: frei Haus Bestellung auf Rechnung SZABO-SCANDIC Lieferung: € 10,- HandelsgmbH & Co KG Erstbestellung Vorauskassa Quellenstraße 110, A-1100 Wien T. +43(0)1 489 3961-0 Zuschläge F. +43(0)1 489 3961-7 [email protected] • Mindermengenzuschlag www.szabo-scandic.com • Trockeneiszuschlag • Gefahrgutzuschlag linkedin.com/company/szaboscandic • Expressversand facebook.com/szaboscandic RAB2B (Human) Recombinant Ras superfamily that contain 4 highly conserved regions Protein (P01) involved in GTP binding and hydrolysis. Rab proteins are prenylated, membrane-bound proteins involved in Catalog Number: H00084932-P01 vesicular fusion and trafficking; see MIM 179508.[supplied by OMIM] Regulation Status: For research use only (RUO) Product Description: Human RAB2B full-length ORF ( NP_116235.2, 1 a.a. - 216 a.a.) recombinant protein with GST-tag at N-terminal. Sequence: MTYAYLFKYIIIGDTGVGKSCLLLQFTDKRFQPVHDLTI GVEFGARMVNIDGKQIKLQIWDTAGQESFRSITRSYYR GAAGALLVYDITRRETFNHLTSWLEDARQHSSSNMVI MLIGNKSDLESRRDVKREEGEAFAREHGLIFMETSAKT ACNVEEAFINTAKEIYRKIQQGLFDVHNEANGIKIGPQQ SISTSVGPSASQRNSRDIGSNSGCC Host: Wheat Germ (in vitro) Theoretical MW (kDa): 50.6 Applications: AP, Array, ELISA, WB-Re (See our web site product page for detailed applications information) Protocols: See our web site at http://www.abnova.com/support/protocols.asp -
Isolation and Identification of Proteins from Swine Sperm Chromatin and Nuclear Matrix
DOI: 10.21451/1984-3143-AR816 Anim. Reprod., v.14, n.2, p.418-428, Apr./Jun. 2017 Isolation and identification of proteins from swine sperm chromatin and nuclear matrix Guilherme Arantes Mendonça1,3, Romualdo Morandi Filho2, Elisson Terêncio Souza2, Thais Schwarz Gaggini1, Marina Cruvinel Assunção Silva-Mendonça1, Robson Carlos Antunes1, Marcelo Emílio Beletti1,2 1Post-graduation Program in Veterinary Science, Federal University of Uberlandia, Uberlandia, MG, Brazil. 2Post-graduation Program in Cellular and Molecular Biology, Federal University of Uberlandia, Uberlandia, MG, Brazil. Abstract (Yamauchi et al., 2011). According to the same authors, these active sperm chromatin sites in protamine toroids The aim of this study was to perform a may contain important epigenetic information for the proteomic analysis to isolate and identify proteins from developing embryo. the swine sperm nuclear matrix to contribute to a The isolated use of genomic and transcriptomic database of swine sperm nuclear proteins. We used pre- information may be insufficient to fully understand a chilled diluted semen from seven boars (19 to 24 week- complex organism because proteomics and old) from the commercial line Landrace x Large White transcriptomics can be discordant and DNA-RNA x Pietran. The semen was processed to separate the relationships cannot be fully correlated. Thus, sperm heads and extract the chromatin and nuclear measurements of other metabolic levels should also be matrix for protein quantification and analysis by mass obtained, such as the study of proteins (Wright et al., spectrometry, by LTQ Orbitrap ELITE mass 2012). According to these same authors, large-scale spectrometer (Thermo-Finnigan) coupled to a nanoflow protein research in organisms (i.e., the proteome-protein chromatography system (LC-MS/MS). -
Agricultural University of Athens
ΓΕΩΠΟΝΙΚΟ ΠΑΝΕΠΙΣΤΗΜΙΟ ΑΘΗΝΩΝ ΣΧΟΛΗ ΕΠΙΣΤΗΜΩΝ ΤΩΝ ΖΩΩΝ ΤΜΗΜΑ ΕΠΙΣΤΗΜΗΣ ΖΩΙΚΗΣ ΠΑΡΑΓΩΓΗΣ ΕΡΓΑΣΤΗΡΙΟ ΓΕΝΙΚΗΣ ΚΑΙ ΕΙΔΙΚΗΣ ΖΩΟΤΕΧΝΙΑΣ ΔΙΔΑΚΤΟΡΙΚΗ ΔΙΑΤΡΙΒΗ Εντοπισμός γονιδιωματικών περιοχών και δικτύων γονιδίων που επηρεάζουν παραγωγικές και αναπαραγωγικές ιδιότητες σε πληθυσμούς κρεοπαραγωγικών ορνιθίων ΕΙΡΗΝΗ Κ. ΤΑΡΣΑΝΗ ΕΠΙΒΛΕΠΩΝ ΚΑΘΗΓΗΤΗΣ: ΑΝΤΩΝΙΟΣ ΚΟΜΙΝΑΚΗΣ ΑΘΗΝΑ 2020 ΔΙΔΑΚΤΟΡΙΚΗ ΔΙΑΤΡΙΒΗ Εντοπισμός γονιδιωματικών περιοχών και δικτύων γονιδίων που επηρεάζουν παραγωγικές και αναπαραγωγικές ιδιότητες σε πληθυσμούς κρεοπαραγωγικών ορνιθίων Genome-wide association analysis and gene network analysis for (re)production traits in commercial broilers ΕΙΡΗΝΗ Κ. ΤΑΡΣΑΝΗ ΕΠΙΒΛΕΠΩΝ ΚΑΘΗΓΗΤΗΣ: ΑΝΤΩΝΙΟΣ ΚΟΜΙΝΑΚΗΣ Τριμελής Επιτροπή: Aντώνιος Κομινάκης (Αν. Καθ. ΓΠΑ) Ανδρέας Κράνης (Eρευν. B, Παν. Εδιμβούργου) Αριάδνη Χάγερ (Επ. Καθ. ΓΠΑ) Επταμελής εξεταστική επιτροπή: Aντώνιος Κομινάκης (Αν. Καθ. ΓΠΑ) Ανδρέας Κράνης (Eρευν. B, Παν. Εδιμβούργου) Αριάδνη Χάγερ (Επ. Καθ. ΓΠΑ) Πηνελόπη Μπεμπέλη (Καθ. ΓΠΑ) Δημήτριος Βλαχάκης (Επ. Καθ. ΓΠΑ) Ευάγγελος Ζωίδης (Επ.Καθ. ΓΠΑ) Γεώργιος Θεοδώρου (Επ.Καθ. ΓΠΑ) 2 Εντοπισμός γονιδιωματικών περιοχών και δικτύων γονιδίων που επηρεάζουν παραγωγικές και αναπαραγωγικές ιδιότητες σε πληθυσμούς κρεοπαραγωγικών ορνιθίων Περίληψη Σκοπός της παρούσας διδακτορικής διατριβής ήταν ο εντοπισμός γενετικών δεικτών και υποψηφίων γονιδίων που εμπλέκονται στο γενετικό έλεγχο δύο τυπικών πολυγονιδιακών ιδιοτήτων σε κρεοπαραγωγικά ορνίθια. Μία ιδιότητα σχετίζεται με την ανάπτυξη (σωματικό βάρος στις 35 ημέρες, ΣΒ) και η άλλη με την αναπαραγωγική -
Comprehensive Knockout Analysis of the Rab Family Gtpases in Epithelial Cells
Comprehensive knockout analysis of the Rab family GTPases in epithelial cells 著者 Yuta Homma, Riko Kinoshita, Yoshihiko Kuchitsu, Paulina S Wawro, Soujiro Marubashi, Mai E Oguchi, Mori´e Ishida, Naonobu Fujita, Mitsunori Fukuda journal or The journal of cell biology publication title volume 218 number 6 page range 2035-2050 year 2019-05-09 URL http://hdl.handle.net/10097/00127022 doi: 10.1083/jcb.201810134 Creative Commons : 表示 - 非営利 http://creativecommons.org/licenses/by-nc/3.0/deed.ja Published Online: 9 May, 2019 | Supp Info: http://doi.org/10.1083/jcb.201810134 Downloaded from jcb.rupress.org on November 6, 2019 TOOLS Comprehensive knockout analysis of the Rab family GTPases in epithelial cells Yuta Homma, Riko Kinoshita, Yoshihiko Kuchitsu, Paulina S. Wawro, Soujiro Marubashi, Mai E. Oguchi, Morie´ Ishida, Naonobu Fujita, and Mitsunori Fukuda The Rab family of small GTPases comprises the largest number of proteins (∼60 in mammals) among the regulators of intracellular membrane trafficking, but the precise function of many Rabs and the functional redundancy and diversity of Rabs remain largely unknown. Here, we generated a comprehensive collection of knockout (KO) MDCK cells for the entire Rab family. We knocked out closely related paralogs simultaneously (Rab subfamily knockout) to circumvent functional compensation and found that Rab1A/B and Rab5A/B/C are critical for cell survival and/or growth. In addition, we demonstrated that Rab6-KO cells lack the basement membrane, likely because of the inability to secrete extracellular matrix components. Further analysis revealed the general requirement of Rab6 for secretion of soluble cargos. Transport of transmembrane cargos to the plasma membrane was also significantly delayed in Rab6-KO cells, but the phenotype was relatively mild.