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https://www.readbyqxmd.com/read/28102837/precise-and-efficient-scarless-genome-editing-in-stem-cells-using-correct
#1
Dylan Kwart, Dominik Paquet, Shaun Teo, Marc Tessier-Lavigne
CRISPR/Cas9 is a promising tool for genome-editing DNA in cells with single-base-pair precision, which allows novel in vitro models of human disease to be generated-e.g., in pluripotent stem cells. However, the accuracy of intended sequence changes can be severely diminished by CRISPR/Cas9's propensity to re-edit previously modified loci, causing unwanted mutations (indels) alongside intended changes. Here we describe a genome-editing framework termed consecutive re-guide or re-Cas steps to erase CRISPR/Cas-blocked targets (CORRECT), which, by exploiting the use of highly efficacious CRISPR/Cas-blocking mutations in two rounds of genome editing, enables accurate, efficient and scarless introduction of specific base changes-for example, in human induced pluripotent (iPS) stem cells...
February 2017: Nature Protocols
https://www.readbyqxmd.com/read/28102835/spinal-cord-regeneration-in-xenopus-laevis
#2
Gabriela Edwards-Faret, Rosana Muñoz, Emilio E Méndez-Olivos, Dasfne Lee-Liu, Victor S Tapia, Juan Larraín
Here we present a protocol for the husbandry of Xenopus laevis tadpoles and froglets, and procedures to study spinal cord regeneration. This includes methods to induce spinal cord injury (SCI); DNA and morpholino electroporation for genetic studies; in vivo imaging for cell analysis; a swimming test to measure functional recovery; and a convenient model for screening for new compounds that promote neural regeneration. These protocols establish X. laevis as a unique model organism for understanding spinal cord regeneration by comparing regenerative and nonregenerative stages...
February 2017: Nature Protocols
https://www.readbyqxmd.com/read/28102819/rethinking-origin-licensing
#3
Stephen P Bell
Human cells that lack a subunit in their origin recognition complex are viable, which suggests the existence of alternative mechanisms to initiate DNA replication.
January 19, 2017: ELife
https://www.readbyqxmd.com/read/28102818/is-cell-size-a-spandrel
#4
Ariel Amir
All organisms control the size of their cells. We focus here on the question of size regulation in bacteria, and suggest that the quantitative laws governing cell size and its dependence on growth rate may arise as byproducts of a regulatory mechanism which evolved to support multiple DNA replication forks. In particular, we show that the increase of bacterial cell size during Lenski's long-term evolution experiments is a natural outcome of this proposal. This suggests that, in the context of evolution, cell size may be a 'spandrel'...
January 19, 2017: ELife
https://www.readbyqxmd.com/read/28102666/a-simulation-approach-for-timing-analysis-of-genetic-logic-circuits
#5
Hasan Baig, Jan Madsen
Constructing genetic logic circuits is an application of synthetic biology, where parts of the DNA of a living cell are engineered to perform a dedicated Boolean function triggered by appropriate concentration levels of certain proteins or by different genetic components. These logic circuits work in a manner similar to electronic logic circuits, but are much more stochastic and hence much harder to characterize. In this paper, we introduce an approach to analyze the threshold value and timing of genetic logic circuits...
January 19, 2017: ACS Synthetic Biology
https://www.readbyqxmd.com/read/28102569/human-%C3%AE-defensin-3-increases-the-tlr9-dependent-response-to-bacterial-dna
#6
Sarah L McGlasson, Fiona Semple, Heather MacPherson, Mohini Gray, Donald J Davidson, Julia R Dorin
Human β-defensin 3 (hBD3) is a cationic antimicrobial peptide with potent bactericidal activity in vitro. HBD3 is produced in response to pathogen challenge and can modulate immune responses. The amplified recognition of self-DNA by human plasmacytoid dendritic cells has been previously reported, but we show here that hBD3 preferentially enhances the response to bacterial DNA in mouse Flt-3 induced dendritic cells (FLDCs) and in human peripheral blood mononuclear cells. We show the effect is mediated through TLR9 and although hBD3 significantly increases the cellular uptake of both E...
January 19, 2017: European Journal of Immunology
https://www.readbyqxmd.com/read/28102347/low-level-predation-by-lytic-phage-phiipla-rodi-promotes-biofilm-formation-and-triggers-the-stringent-response-in-staphylococcus-aureus
#7
Lucía Fernández, Silvia González, Ana Belén Campelo, Beatriz Martínez, Ana Rodríguez, Pilar García
An important lesson from the war on pathogenic bacteria has been the need to understand the physiological responses and evolution of natural microbial communities. Bacterial populations in the environment are generally forming biofilms subject to some level of phage predation. These multicellular communities are notoriously resistant to antimicrobials and, consequently, very difficult to eradicate. This has sparked the search for new therapeutic alternatives, including phage therapy. This study demonstrates that S...
January 19, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28102346/human-p53-interacts-with-the-elongating-rnapii-complex-and-is-required-for-the-release-of-actinomycin-d-induced-transcription-blockage
#8
Barbara N Borsos, Ildikó Huliák, Hajnalka Majoros, Zsuzsanna Ujfaludi, Ákos Gyenis, Peter Pukler, Imre M Boros, Tibor Pankotai
The p53 tumour suppressor regulates the transcription initiation of selected genes by binding to specific DNA sequences at their promoters. Here we report a novel role of p53 in transcription elongation in human cells. Our data demonstrate that upon transcription elongation blockage, p53 is associated with genes that have not been reported as its direct targets. p53 could be co-immunoprecipitated with active forms of DNA-directed RNA polymerase II subunit 1 (RPB1), highlighting its association with the elongating RNA polymerase II...
January 19, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28102297/whsc1l1-mediated-egfr-mono-methylation-enhances-the-cytoplasmic-and-nuclear-oncogenic-activity-of-egfr-in-head-and-neck-cancer
#9
Vassiliki Saloura, Theodore Vougiouklakis, Makda Zewde, Xiaolan Deng, Kazuma Kiyotani, Jae-Hyun Park, Yo Matsuo, Mark Lingen, Takehiro Suzuki, Naoshi Dohmae, Ryuji Hamamoto, Yusuke Nakamura
While multiple post-translational modifications have been reported to regulate the function of epidermal growth factor receptor (EGFR), the effect of protein methylation on its function has not been well characterized. In this study, we show that WHSC1L1 mono-methylates lysine 721 in the tyrosine kinase domain of EGFR, and that this methylation leads to enhanced activation of its downstream ERK cascade without EGF stimulation. We also show that EGFR K721 mono-methylation not only affects the function of cytoplasmic EGFR, but also that of nuclear EGFR...
January 19, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28102121/inflammasomes-in-myeloid-cells-warriors-within
#10
Sushmita Jha, W June Brickey, Jenny Pan-Yun Ting
The inflammasome is a large multimeric protein complex comprising an effector protein that demonstrates specificity for a variety of activators or ligands; an adaptor molecule; and procaspase-1, which is converted to caspase-1 upon inflammasome activation. Inflammasomes are expressed primarily by myeloid cells and are located within the cell. The macromolecular inflammasome structure can be visualized by cryo-electron microscopy. This complex has been found to play a role in a variety of disease models in mice, and several have been genetically linked to human diseases...
January 2017: Microbiology Spectrum
https://www.readbyqxmd.com/read/28102109/epigenetic-regulation-of-rgs2-regulator-of-g-protein-signaling-2-in-chemoresistant-ovarian-cancer-cells
#11
Ercan Cacan
Regulator of G-protein signaling 2 (RGS2) is a GTPase-activating protein functioning as an inhibitor of G-protein coupled receptors (GPCRs). RGS2 dysregulation was implicated in solid tumour development and RGS2 downregulation has been reported in prostate and ovarian cancer progression. However, the molecular mechanism by which RGS2 expression is suppressed in ovarian cancer remains unknown. The expression and epigenetic regulation of RGS2 in chemosensitive and chemoresistant ovarian cancer cells were determined by qRT-PCR and chromatin immunoprecipitation assays, respectively...
January 19, 2017: Journal of Chemotherapy
https://www.readbyqxmd.com/read/28102058/neonatal-immune-state-is-influenced-by-maternal-allergic-rhinitis-and-associated-with-regulatory-t-cells
#12
Lu Tan, Jing Ou, Zezhang Tao, Yonggang Kong, Yu Xu
PURPOSE: Maternal influences contribute to the origin of allergic diseases, but the mechanisms are not clear. The current literature prompted the role of epigenetics in the development of allergic diseases. We sought to investigate the roles of regulatory T (Treg) cells and Forkhead box p3 (Foxp3) DNA methylation in the process of maternal transmission of allergic rhinitis (AR) susceptibility. METHODS: BALB/c female mice (AR mother) were sensitized by intraperitoneal injection of Dermatophagoides pteronyssinus (Der p) 1 on day 1 and 7...
March 2017: Allergy, Asthma & Immunology Research
https://www.readbyqxmd.com/read/28102036/combining-one-step-sanger-sequencing-with-phasing-probe-hybridization-for-hla-class-i-typing-yields-rapid-g-group-resolution-predicting-99-of-unique-full-length-protein-sequences
#13
Bin Tu, Carly Masaberg, Lihua Hou, Daniel Behm, Peter Brescia, Nuri Cha, Kanthi Kariyawasam, Jar How Lee, Thoa Nong, John Sells, Paul Tausch, Ruyan Yang, Jennifer Ng, Carolyn Katovich Hurley
BACKGROUND: Sanger-based DNA sequencing of exons 2+3 of HLA class I alleles from a heterozygote frequently results in two or more alternative genotypes. This study was undertaken to reduce the time and effort required to produce a single high resolution HLA genotype. MATERIALS AND METHODS: Samples were typed in parallel by Sanger sequencing and oligonucleotide probe hybridization. This workflow, together with optimization of analysis software, was tested and refined during the typing of over 42,000 volunteers for an unrelated hematopoietic progenitor cell donor registry...
February 2017: HLA
https://www.readbyqxmd.com/read/28102032/nonpromoter-methylation-of-the-cdkn2a-gene-with-active-transcription-is-associated-with-improved-locoregional-control-in-laryngeal-squamous-cell-carcinoma
#14
Miriam M Ben-Dayan, Thomas J Ow, Thomas J Belbin, Joshua Wetzler, Richard V Smith, Geoffrey Childs, Brenda Diergaarde, D Neil Hayes, Jennifer R Grandis, Michael B Prystowsky, Nicolas F Schlecht
We previously reported a novel association between CDKN2A nonpromoter methylation and transcription (ARF/INK4a) in human papillomavirus associated oropharyngeal tumors. In this study we assessed whether nonpromoter CDKN2A methylation in laryngeal squamous cell carcinomas (LXSCC) conferred a similar association with transcription that predicted patient outcome. We compared DNA methylation and ARF/INK4a RNA expression levels for the CDKN2A locus using the Illumina HumanMethylation27 beadchip and RT-PCR in 43 LXSCC tumor samples collected from a prospective study of head and neck cancer patients treated at Montefiore Medical Center (MMC)...
January 19, 2017: Cancer Medicine
https://www.readbyqxmd.com/read/28102011/local-myogenic-pulp-derived-cell-injection-enhances-craniofacial-muscle-regeneration-in-vivo
#15
J E Jung, M J Song, S Shin, Y J Choi, K H Kim, C J Chung
OBJECTIVES: To enhance myogenic differentiation in pulp cells isolated from extracted premolars by epigenetic modification using a DNA demethylation agent, 5-aza-2'-deoxycytidine (5-Aza), and to evaluate the potent stimulatory effect of 5-Aza-treated pulp cell injection for craniofacial muscle regeneration in vivo. SETTING AND SAMPLE POPULATION: Pulp cells were isolated from premolars extracted for orthodontic purposes from four adults (age range, 18-22.1 years)...
February 2017: Orthodontics & Craniofacial Research
https://www.readbyqxmd.com/read/28101907/increased-arf-p53-activity-in-stem-cells-aging-and-cancer
#16
REVIEW
Estefania Carrasco-Garcia, Manuel Moreno, Leire Moreno-Cugnon, Ander Matheu
Arf/p53 pathway protects the cells against DNA damage induced by acute stress. This characteristic is the responsible for its tumor suppressor activity. Moreover, it regulates the chronic type of stress associated with aging. This is the basis of its anti-aging activity. Indeed, increased gene dosage of Arf/p53 displays elongated longevity and delayed aging. At a cellular level, it has been recently shown that increased dosage of Arf/p53 delays age-associated stem cell exhaustion and the subsequent decline in tissue homeostasis and regeneration...
January 19, 2017: Aging Cell
https://www.readbyqxmd.com/read/28101820/insulin-insulin-like-growth-factor-1-signalling-iis-based-regulation-of-lifespan-across-species
#17
REVIEW
Rebecca Mathew, Manika Pal Bhadra, Utpal Bhadra
An organism's well-being is facilitated by numerous molecular and biochemical pathways that ensure homeostasis within cells and tissues. Aging causes a gradual let-down in the maintenance of homeostasis due to various endogenous and environmental challenges, leading to amassing of damages, functional deterioration of different tissues and vulnerability to ailments. Nutrient sensing pathways that maintain glucose homeostasis in body are involved in regulation of aging. Insulin/insulin-like growth factor-1 (IGF-1) signalling (IIS) pathway was the first nutrient sensing pathway discovered to affect the aging process...
January 18, 2017: Biogerontology
https://www.readbyqxmd.com/read/28101801/identification-of-changes-in-the-human-papilloma-virus-16-hpv16-genome-during-early-dissemination-of-cervical-cancer-cells-may-complement-histological-diagnosis-of-lymph-node-metastasis
#18
Anirban Roychowdhury, Sudip Samadder, Md Saimul Islam, Kalyansree Chaudhury, Anup Roy, Dipanwita Banerjee, Ranajit Mandal, Partha S Basu, Susanta Roychoudhury, Chinmay Kumar Panda
Cancer of the uterine cervix (CACX) is one of the most common carcinoma affecting women worldwide. During treatment, histologically lymph node (LN) metastasis and presence of HPV DNA in blood plasma act as a major prognostic factor. Despite the lack of apparent LN involvement, some early-invasive CACX patients have shown recurrences and poor survival. This is suggestive of undetected early dissemination of cancer cells characterized by presence of HPV DNA in histologically non-metastatic LNs which finally progresses into histologically visible metastasis...
January 19, 2017: Pathology Oncology Research: POR
https://www.readbyqxmd.com/read/28101670/recombination-correlates-with-synaptonemal-complex-length-and-chromatin-loop-size-in-bovids-insights-into-mammalian-meiotic-chromosomal-organization
#19
Aurora Ruiz-Herrera, Miluse Vozdova, Jonathan Fernández, Hana Sebestova, Laia Capilla, Jan Frohlich, Covadonga Vara, Adrià Hernández-Marsal, Jaroslav Sipek, Terence J Robinson, Jiri Rubes
Homologous chromosomes exchange genetic information through recombination during meiosis, a process that increases genetic diversity, and is fundamental to sexual reproduction. In an attempt to shed light on the dynamics of mammalian recombination and its implications for genome organization, we have studied the recombination characteristics of 112 individuals belonging to 28 different species in the family Bovidae. In particular, we analyzed the distribution of RAD51 and MLH1 foci during the meiotic prophase I that serve, respectively, as proxies for double-strand breaks (DSBs) which form in early stages of meiosis and for crossovers...
January 18, 2017: Chromosoma
https://www.readbyqxmd.com/read/28101569/resveratrol-inhibits-adventitial-fibroblast-proliferation-and-induces-cell-apoptosis-through-the-sirt1-pathway
#20
Lin Ling, Shaohua Gu, Yan Cheng
Atherosclerosis is one of the most important causes of cardiovascular disease and studies have showed that adventitial fibroblasts, which are considered to be the most common cell type of the vascular adventitia, are involved in the development of early atherosclerotic plaques. Resveratrol is a plant polyphenolic compound confirmed to have anti‑atherosclerotic and cardioprotective effects. The aim of the present study was to investigate the effects of resveratrol on adventitial fibroblasts in vitro and to clarify the underlying mechanism...
February 2017: Molecular Medicine Reports
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