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https://www.readbyqxmd.com/read/29330802/microinjection-of-antisense-morpholinos-crispr-cas9-rnp-and-rna-dna-into-zebrafish-embryos
#1
Yi Xin, Cunming Duan
In this chapter, we describe a stepwise protocol of microinjection. Using this method, antisense morpholinos, CRISPR-Cas9 ribonucleoprotein complexes, capped mRNA, and DNA can be delivered into fertilized zebrafish eggs to manipulate gene expression during development. This protocol can also be adapted for microinjection in other fish and amphibian species.
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29330354/the-res-complex-is-required-for-efficient-transformation-of-the-precatalytic-b-spliceosome-into-an-activated-bact-complex
#2
Penghui Bao, Cindy L Will, Henning Urlaub, Kum-Loong Boon, Reinhard Lührmann
The precise function of the trimeric retention and splicing (RES) complex in pre-mRNA splicing remains unclear. Here we dissected the role of RES during the assembly and activation of yeast spliceosomes. The efficiency of pre-mRNA splicing was significantly lower in the absence of the RES protein Snu17, and the recruitment of its binding partners, Pml1 (pre-mRNA leakage protein 1) and Bud13 (bud site selection protein 13), to the spliceosome was either abolished or substantially reduced. RES was not required for the assembly of spliceosomal B complexes, but its absence hindered efficient Bact complex formation...
January 12, 2018: Genes & Development
https://www.readbyqxmd.com/read/29326986/measles-virus-background-and-oncolytic-virotherapy
#3
REVIEW
Sankhajit Bhattacharjee, Pramod Kumar Yadava
Measles is a highly transmissible disease caused by measles virus and remains a major cause of child mortality in developing countries. Measles virus nucleoprotein (N) encapsidates the RNA genome of the virus for providing protection from host cell endonucleases and for specific recognition of viral RNA as template for transcription and replication. This protein is over-expressed at the time of viral replication. The C-terminal of N protein is intrinsically disordered, which enables this protein to interact with several host cell proteins...
March 2018: Biochemistry and Biophysics Reports
https://www.readbyqxmd.com/read/29321445/negative-regulation-of-b-cell-responses-and-self-tolerance-to-rna-related-lupus-self-antigen
#4
Takeshi Tsubata
The antibody response to RNA-related antigens such as Sm/RNP requires the endosomal RNA sensor TLR7, and this process is crucial in the development of systemic lupus erythematosus at least in animal models. The inhibitory B cell receptor CD72 is unique because it recognizes Sm/RNP and specifically inhibits the activation of Sm/RNP-reactive B cells by activating SH2-containing protein tyrosine phosphatase 1 (SHP-1). In the normal immune system, Sm/RNP-reactive B cells are tolerized by a unique mechanism that probably involves SHP-1...
2018: Proceedings of the Japan Academy. Series B, Physical and Biological Sciences
https://www.readbyqxmd.com/read/29321324/influenza-c-and-d-viruses-package-eight-organized-ribonucleoprotein-complexes
#5
Sumiho Nakatsu, Shin Murakami, Keiko Shindo, Taisuke Horimoto, Hiroshi Sagara, Takeshi Noda, Yoshihiro Kawaoka
Influenza A and B viruses have eight-segmented, single-stranded, negative-sense RNA genomes, whereas influenza C and D viruses have seven-segmented genomes. Each genomic RNA segment exists in the form of a ribonucleoprotein complex (RNP) in association with nucleoproteins and an RNA-dependent RNA polymerase in virions. Influenza D virus was recently isolated from swine and cattle, but its morphology is not fully studied. Here, we examined the morphological characteristics of D/bovine/Yamagata/10710/2016 (D/Yamagata) and C/Ann Arbor/50 (C/AA), focusing on RNPs packaged within the virions...
January 10, 2018: Journal of Virology
https://www.readbyqxmd.com/read/29309035/dissection-of-affinity-captured-line-1-macromolecular-complexes
#6
Martin S Taylor, Ilya Altukhov, Kelly R Molloy, Paolo Mita, Hua Jiang, Emily M Adney, Aleksandra Wudzinska, Sana Badri, Dmitry Ischenko, George Eng, Kathleen H Burns, David Fenyö, Brian T Chait, Dmitry Alexeev, Michael P Rout, Jef D Boeke, John LaCava
Long Interspersed Nuclear Element-1 (LINE-1, L1) is a mobile genetic element active in human genomes. L1-encoded ORF1 and ORF2 proteins bind L1 RNAs, forming ribonucleoproteins (RNPs). These RNPs interact with diverse host proteins, some repressive and others required for the L1 lifecycle. Using differential affinity purifications, quantitative mass spectrometry, and next generation RNA sequencing, we have characterized the proteins and nucleic acids associated with distinctive, enzymatically active L1 macromolecular complexes...
January 8, 2018: ELife
https://www.readbyqxmd.com/read/29302061/importance-of-the-1-7-configuration-of-ribonucleoprotein-complexes-for-influenza-a-virus-genome-packaging
#7
Takeshi Noda, Shin Murakami, Sumiho Nakatsu, Hirotaka Imai, Yukiko Muramoto, Keiko Shindo, Hiroshi Sagara, Yoshihiro Kawaoka
The influenza A virus genome is composed of eight single-stranded negative-sense RNAs. Eight distinct viral RNA segments (vRNAs) are selectively packaged into progeny virions, with eight vRNAs in ribonucleoprotein complexes (RNPs) arranged in a specific "1+7" pattern, that is, one central RNP surrounded by seven RNPs. Here we report the genome packaging of an artificially generated seven-segment virus that lacks the hemagglutinin (HA) vRNA. Electron microscopy shows that, even in the presence of only seven vRNAs, the virions efficiently package eight RNPs arranged in the same "1+7" pattern as wild-type virions...
January 4, 2018: Nature Communications
https://www.readbyqxmd.com/read/29290190/determination-of-specificity-and-pattern-of-antinuclear-antibodies-ana-in-systemic-rheumatic-disease-patients-positive-for-ana-testing
#8
Hamid Nawaz Tipu, Muhammad Mukarram Bashir
OBJECTIVE: To determine probability of finding antinuclear antibodies (ANA) and anti extractable nuclear antigens (ENA) positive samples and associating ANA patterns with anti-ENA reactivities among a consecutive cohort of samples of systemic rheumatic disease patients referred for ANA testing. STUDY DESIGN: Prospective cohort study. PLACE AND DURATION OF STUDY: Immunology Department, Armed Forces Institute of Pathology, Rawalpindi, Pakistan, from January to June 2016...
January 2018: Journal of the College of Physicians and Surgeons—Pakistan: JCPSP
https://www.readbyqxmd.com/read/29274563/iga-anti-%C3%AE-2-glycoprotein-i-antibodies-experience-from-a-large-center
#9
Alexandru Vlagea, Dora Pascual-Salcedo, Rita Álvarez Doforno, Paz Lavilla, Jesús Diez, Beatriz Padilla Merlano, María V Cuesta, Antonio Gil
OBJECTIVE: IgG and IgM antibodies directed at β2-glycoprotein I are included in the classification criteria for the antiphospholipid syndrome (APS) while the IgA antibodies against β2-glycoprotein I (IgA aβ2GPI) are not. Conflicting data about the significance of IgA aβ2GPI and APS manifestation can be found and more studies are necessary in order to define the diagnostic value of IgA aβ2GPI. In the present article, we investigated the possible role of IgA aβ2GPI as marker of APS...
December 12, 2017: Thrombosis Research
https://www.readbyqxmd.com/read/29273498/engineering-protein-secreting-plasma-cells-by-homology-directed-repair-in-primary-human-b-cells
#10
King L Hung, Iana Meitlis, Malika Hale, Chun-Yu Chen, Swati Singh, Shaun W Jackson, Carol H Miao, Iram F Khan, David J Rawlings, Richard G James
The ability to engineer primary human B cells to differentiate into long-lived plasma cells and secrete a de novo protein may allow the creation of novel plasma cell therapies for protein deficiency diseases and other clinical applications. We initially developed methods for efficient genome editing of primary B cells isolated from peripheral blood. By delivering CRISPR/CRISPR-associated protein 9 (Cas9) ribonucleoprotein (RNP) complexes under conditions of rapid B cell expansion, we achieved site-specific gene disruption at multiple loci in primary human B cells (with editing rates of up to 94%)...
November 22, 2017: Molecular Therapy: the Journal of the American Society of Gene Therapy
https://www.readbyqxmd.com/read/29236766/influences-of-micro-geomorphology-on-the-stoichiometry-of-c-n-and-p-in-chenier-island-soils-and-plants-in-the-yellow-river-delta-china
#11
Fanzhu Qu, Ling Meng, Junbao Yu, Jingtao Liu, Jingkuan Sun, Hongjun Yang, Linshui Dong
Studies have indicated that consistent or well-constrained (relatively low variability) carbon:nitrogen:phosphorus (C:N:P) ratios exist in large-scale ecosystems, including both marine and terrestrial ecosystems. Little is known about the C, N and P stoichiometric ratios that exist in the soils and plants of Chenier Island in the Yellow River Delta (YRD). We examined the distribution patterns and relationships of C, N and P stoichiometry in the soils and plants of Chenier Island, as well as the potential influences of the island's micro-geomorphology...
2017: PloS One
https://www.readbyqxmd.com/read/29226180/cytosolic-and-nuclear-delivery-of-crispr-cas9-ribonucleoprotein-for-gene-editing-using-arginine-functionalized-gold-nanoparticles
#12
Rubul Mout, Vincent M Rotello
In this protocol, engineered Cas9-ribonucleoprotein (Cas9 protein and sgRNA, together called Cas9-RNP) and gold nanoparticles are used to make nanoassemblies that are employed to deliver Cas9-RNP into cell cytoplasm and nucleus. Cas9 protein is engineered with an N-terminus glutamic acid tag (E-tag or En, where n = the number of glutamic acid in an E-tag and usually n = 15 or 20), C-terminus nuclear localizing signal (NLS), and a C-terminus 6xHis-tag. [Cas9En hereafter] To use this protocol, the first step is to generate the required materials (gold nanoparticles, recombinant Cas9En, and sgRNA)...
October 20, 2017: Bio-protocol
https://www.readbyqxmd.com/read/29214036/strong-viral-associations-with-sle-among-filipinos
#13
Evan S Vista, Michael H Weisman, Mariko L Ishimori, Hua Chen, Rebecka L Bourn, Ben F Bruner, Laniyati Hamijoyo, Robelle D Tanangunan, Noga J Gal, Julie M Robertson, John B Harley, Joel M Guthridge, Sandra V Navarra, Judith A James
Objectives: Epstein-Barr virus (EBV) is considered an important environmental factor in SLE aetiology, but the relationship between SLE and EBV in the Filipino population is unknown. We tested associations between SLE, lupus-associated autoantibodies and seropositivity for EBV and other herpes viruses in the Filipino population. Methods: Sera from Filipino patients with SLE (n=233), unaffected first-degree relatives (FDRs, n=543) and unrelated controls (n=221) were tested for antibodies against EBV, cytomegalovirus (CMV) and herpes simplex viruses (HSV-1 and HSV-2) by standardised ELISAs...
2017: Lupus Science & Medicine
https://www.readbyqxmd.com/read/29197185/structure-and-interaction-of-corynebacterium-pseudotuberculosis-cold-shock-protein-a-with-y-box-single-stranded-dna-fragment
#14
Ícaro P Caruso, Vineet Panwalkar, Monika A Coronado, Andrew J Dingley, Marinônio L Cornélio, Dieter Willbold, Raghuvir K Arni, Raphael J Eberle
Cold shock proteins (Csps) function to preserve cell viability at low temperatures by binding to nucleic acids and consequently control gene expression. The mesophilic bacterium Corynebacterium pseudotuberculosis is the causative agent of caseous lymphadenitis in animals, and infection to livestock is a considerable economic burden worldwide. In this report, the structure of cold shock protein A from C. pseudotuberculosis (Cp-CspA) and biochemical analysis of its temperature-dependent interaction with a Y-box single-stranded DNA (ssDNA) motif is presented...
December 2, 2017: FEBS Journal
https://www.readbyqxmd.com/read/29191215/domestication-of-self-splicing-introns-during-eukaryogenesis-the-rise-of-the-complex-spliceosomal-machinery
#15
REVIEW
Julian Vosseberg, Berend Snel
ᅟ: The spliceosome is a eukaryote-specific complex that is essential for the removal of introns from pre-mRNA. It consists of five small nuclear RNAs (snRNAs) and over a hundred proteins, making it one of the most complex molecular machineries. Most of this complexity has emerged during eukaryogenesis, a period that is characterised by a drastic increase in cellular and genomic complexity. Although not fully resolved, recent findings have started to shed some light on how and why the spliceosome originated...
December 1, 2017: Biology Direct
https://www.readbyqxmd.com/read/29190635/neonatal-lupus-case-series-of-a-tertiary-hospital
#16
Ana Raquel Teixeira, Mariana Rodrigues, Hercília Guimarães, Cláudia Moura, Iva Brito
Neonatal lupus (NL) is a very rare condition with an estimated incidence of 1 in 20.000 pregnancies. It is caused by the transplacental passage of autoantibodies anti-Ro/SSA, antiSa/SSB antibodies and/or anti-U1 RNP antibodies into the fetal circulation. The mother may be completely asymptomatic or have a known inflammatory rheumatic disease, such as Sjögren syndrome (SS) or Systemic Lupus Erythematosus (SLE). Clinical manifestations are diverse, being the most common cutaneous and cardiac. The authors present a case series of eight cases diagnosed with NL between January 2008 and December 2016 in a tertiary hospital and a brief revision of the literature...
November 29, 2017: Acta Reumatológica Portuguesa
https://www.readbyqxmd.com/read/29186851/gnss-electronic-compass-road-segment-information-fusion-for-vehicle-to-vehicle-collision-avoidance-application
#17
Rui Sun, Qi Cheng, Dabin Xue, Guanyu Wang, Washington Yotto Ochieng
The increasing number of vehicles in modern cities brings the problem of increasing crashes. One of the applications or services of Intelligent Transportation Systems (ITS) conceived to improve safety and reduce congestion is collision avoidance. This safety critical application requires sub-meter level vehicle state estimation accuracy with very high integrity, continuity and availability, to detect an impending collision and issue a warning or intervene in the case that the warning is not heeded. Because of the challenging city environment, to date there is no approved method capable of delivering this high level of performance in vehicle state estimation...
November 25, 2017: Sensors
https://www.readbyqxmd.com/read/29177505/efficient-rna-pseudouridylation-by-eukaryotic-h-aca-ribonucleoproteins-requires-high-affinity-binding-and-correct-positioning-of-guide-rna
#18
Evan A Caton, Erin K Kelly, Rajashekhar Kamalampeta, Ute Kothe
H/ACA ribonucleoproteins (H/ACA RNPs) are responsible for introducing many pseudouridines into RNAs, but are also involved in other cellular functions. Utilizing a purified and reconstituted yeast H/ACA RNP system that is active in pseudouridine formation under physiological conditions, we describe here the quantitative characterization of H/ACA RNP formation and function. This analysis reveals a surprisingly tight interaction of H/ACA guide RNA with the Cbf5p-Nop10p-Gar1p trimeric protein complex whereas Nhp2p binds comparably weakly to H/ACA guide RNA...
November 21, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/29176610/reconstitution-of-the-complete-pathway-of-its2-processing-at-the-pre-ribosome
#19
Lisa Fromm, Sebastian Falk, Dirk Flemming, Jan Michael Schuller, Matthias Thoms, Elena Conti, Ed Hurt
Removal of internal transcribed spacer 2 (ITS2) from pre-ribosomal RNA is essential to make functional ribosomes. This complicated processing reaction begins with a single endonucleolytic cleavage followed by exonucleolytic trimming at both new cleavage sites to generate mature 5.8S and 25S rRNA. We reconstituted the 7S→5.8S processing branch within ITS2 using purified exosome and its nuclear cofactors. We find that both Rrp44's ribonuclease activities are required for initial RNA shortening followed by hand over to the exonuclease Rrp6...
November 27, 2017: Nature Communications
https://www.readbyqxmd.com/read/29155789/efficient-generation-and-editing-of-feeder-free-ipscs-from-human-pancreatic-cells-using-the-crispr-cas9-system
#20
Anjali Nandal, Barbara Mallon, Bhanu P Telugu
Embryonic and induced pluripotent stem cells can self-renew and differentiate into multiple cell types of the body. The pluripotent cells are thus coveted for research in regenerative medicine and are currently in clinical trials for eye diseases, diabetes, heart diseases, and other disorders. The potential to differentiate into specialized cell types coupled with the recent advances in genome editing technologies including the CRISPR/Cas system have provided additional opportunities for tailoring the genome of iPSC for varied applications including disease modeling, gene therapy, and biasing pathways of differentiation, to name a few...
November 8, 2017: Journal of Visualized Experiments: JoVE
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