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Homologous recombination

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https://www.readbyqxmd.com/read/28092459/toward-high-throughput-and-multiplexed-imaging-of-genome-organization
#1
Eric F Joyce
Dr. Eric Joyce from the Department of Genetics at the University of Pennsylvania was awarded The President's Innovation award at the annual Society of Biomolecular Imaging and Informatics meeting held in Boston, September 2016. Chromosome interactions are a fundamental aspect of nuclear organization that can activate and silence genes or even direct chromosome rearrangements. However, the molecular mechanisms underlying how chromosomal segments find each other and form stable interactions within cells remain unknown...
January 2017: Assay and Drug Development Technologies
https://www.readbyqxmd.com/read/28092034/expression-and-purification-of-sh2-domains-using-baculovirus-expression-system
#2
Mari Ogiue-Ikeda, Kazuya Machida
Recombinant proteins expressed in bacteria are sometimes insoluble, aggregated, and incorrectly folded. For those Src homology 2 (SH2) domains that are insoluble in bacteria, baculovirus-insect cell expression systems can be an alternative to produce soluble and functionally active proteins. We describe a protocol for cloning and purification of GST-tagged SH2 domains using the Bac-to-Bac baculovirus expression system.
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28092033/purification-of-socs-suppressor-of-cytokine-signaling-sh2-domains-for-structural-and-functional-studies
#3
Nicholas P D Liau, Artem Laktyushin, Jeffrey J Babon
Src Homology 2 (SH2) domains are protein domains which have a high binding affinity for specific amino acid sequences containing a phosphorylated tyrosine residue. The Suppressors of Cytokine Signaling (SOCS) proteins use an SH2 domain to bind to components of certain cytokine signaling pathways to downregulate the signaling cascade. The recombinantly produced SH2 domains of various SOCS proteins have been used to undertake structural and functional studies elucidating the method of how such targeting occurs...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28092031/expression-and-production-of-sh2-domain-proteins
#4
Bernard A Liu, Mari Ogiue-Ikeda, Kazuya Machida
The Src Homology 2 (SH2) domain lies at the heart of phosphotyrosine signaling, coordinating signaling events downstream of receptor tyrosine kinases (RTKs), adaptors, and scaffolds. Over a hundred SH2 domains are present in mammals, each having a unique specificity which determines its interactions with multiple binding partners. One of the essential tools necessary for studying and determining the role of SH2 domains in phosphotyrosine signaling is a set of soluble recombinant SH2 proteins. Here we describe methods, based on a broad experience with purification of all SH2 domains, for the production of SH2 domain proteins needed for proteomic and biochemical-based studies such as peptide arrays, mass-spectrometry, protein microarrays, reverse-phase microarrays, and high-throughput fluorescence polarization (HTP-FP)...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28090586/holliday-junction-trap-shows-how-cells-use-recombination-and-a-junction-guardian-role-of-recq-helicase
#5
Jun Xia, Li-Tzu Chen, Qian Mei, Chien-Hui Ma, Jennifer A Halliday, Hsin-Yu Lin, David Magnan, John P Pribis, Devon M Fitzgerald, Holly M Hamilton, Megan Richters, Ralf B Nehring, Xi Shen, Lei Li, David Bates, P J Hastings, Christophe Herman, Makkuni Jayaram, Susan M Rosenberg
DNA repair by homologous recombination (HR) underpins cell survival and fuels genome instability, cancer, and evolution. However, the main kinds and sources of DNA damage repaired by HR in somatic cells and the roles of important HR proteins remain elusive. We present engineered proteins that trap, map, and quantify Holliday junctions (HJs), a central DNA intermediate in HR, based on catalytically deficient mutant RuvC protein of Escherichia coli. We use RuvCDefGFP (RDG) to map genomic footprints of HR at defined DNA breaks in E...
November 2016: Science Advances
https://www.readbyqxmd.com/read/28089683/coupling-of-homologous-recombination-and-the-checkpoint-by-atr
#6
Rémi Buisson, Joshi Niraj, Amélie Rodrigue, Chu Kwen Ho, Johannes Kreuzer, Tzeh Keong Foo, Emilie J-L Hardy, Graham Dellaire, Wilhelm Haas, Bing Xia, Jean-Yves Masson, Lee Zou
ATR is a key regulator of cell-cycle checkpoints and homologous recombination (HR). Paradoxically, ATR inhibits CDKs during checkpoint responses, but CDK activity is required for efficient HR. Here, we show that ATR promotes HR after CDK-driven DNA end resection. ATR stimulates the BRCA1-PALB2 interaction after DNA damage and promotes PALB2 localization to DNA damage sites. ATR enhances BRCA1-PALB2 binding at least in part by inhibiting CDKs. The optimal interaction of BRCA1 and PALB2 requires phosphorylation of PALB2 at S59, an ATR site, and hypo-phosphorylation of S64, a CDK site...
December 28, 2016: Molecular Cell
https://www.readbyqxmd.com/read/28089630/the-aspergillus-nidulans-pbp1-homolog-is-required-for-normal-sexual-development-and-secondary-metabolism
#7
Alexandra A Soukup, Gregory J Fischer, Jerry Luo, Nancy P Keller
P bodies and stress granules are RNA-containing structures governing mRNA degradation and translational arrest, respectively. Saccharomyces cerevisiae Pbp1protein localizes to stress granules and promotes their formation and is involved in proper polyadenylation, suppression of RNA-DNA hybrids, and preventing aberrant rDNA recombination. A genetic screen for Aspergillus nidulans mutants aberrant in secondary metabolism identified the Pbp1 homolog, PbpA. Using Dcp1 (mRNA decapping) as a marker for P-body formation and FabM (Pab1, poly-A binding protein) to track stress granule accumulation, we examine the dynamics of RNA granule formation in A...
January 9, 2017: Fungal Genetics and Biology: FG & B
https://www.readbyqxmd.com/read/28087320/poly-adp-ribose-polymerase-activity-and-inhibition-in-cancer
#8
REVIEW
Caleb Dulaney, Samuel Marcrom, Jennifer Stanley, Eddy S Yang
Genomic instability resultant from defective DNA repair mechanisms is a fundamental hallmark of cancer. The poly(ADP-ribose) polymerase (PARP) proteins 1, 2 and 3 catalyze the polymerization of poly(ADP-ribose) and covalent attachment to proteins in a phylogenetically ancient form of protein modification. PARPs play a role in base excision repair, homologous recombination, and non-homologous end joining. The discovery that loss of PARP activity had cytotoxic effects in cells deficient in homologous recombination has sparked a decade of translational research efforts that culminated in the FDA approval of an oral PARP inhibitor for clinical use in patients with ovarian cancer and defective homologous recombination...
January 10, 2017: Seminars in Cell & Developmental Biology
https://www.readbyqxmd.com/read/28087249/nick-initiated-homologous-recombination-protecting-the-genome-one-strand-at-a-time
#9
REVIEW
Lianne E M Vriend, Przemek M Krawczyk
Homologous recombination (HR) is an essential, widely conserved mechanism that utilizes a template for accurate repair of DNA breaks. Some early HR models, developed over five decades ago, anticipated single-strand breaks (nicks) as initiating lesions. Subsequent studies favored a more double-strand break (DSB)-centered view of HR initiation and at present this pathway is primarily considered to be associated with DSB repair. However, mounting evidence suggests that nicks can indeed initiate HR directly, without first being converted to DSBs...
December 29, 2016: DNA Repair
https://www.readbyqxmd.com/read/28081194/t-cell-epitope-containing-domains-of-ragweed-amb-a-1-and-mugwort-art-v-6-modulate-immunologic-responses-in-humans-and-mice
#10
Ana I Sancho, Michael Wallner, Michael Hauser, Birgit Nagl, Martin Himly, Claudia Asam, Christof Ebner, Beatrice Jahn-Schmid, Barbara Bohle, Fatima Ferreira
BACKGROUND: Ragweed (Ambrosia artemisiifolia) and mugwort (Artemisia vulgaris) are the major cause of pollen allergy in late summer. Allergen-specific lymphocytes are crucial for immune modulation during immunotherapy. We sought to generate and pre-clinically characterise highly immunogenic domains of the homologous pectate lyases in ragweed (Amb a 1) and mugwort pollen (Art v 6) for immunotherapy. METHODS: Domains of Amb a 1 (Amb a 1α) and Art v 6 (Art v 6α) and a hybrid molecule, consisting of both domains, were designed, expressed in E...
2017: PloS One
https://www.readbyqxmd.com/read/28079255/moonlighting-at-replication-forks-a-new-life-for-homologous-recombination-proteins-brca1-brca2-and-rad51
#11
REVIEW
Arun Mouli Kolinjivadi, Vincenzo Sannino, Anna de Antoni, Hervé Técher, Giorgio Baldi, Vincenzo Costanzo
Coordination between DNA replication and DNA repair ensures maintenance of genome integrity, which is lost in cancer cells. Emerging evidence has linked Homologous Recombination (HR) proteins RAD51, BRCA1 and BRCA2 to the stability of nascent DNA. This function appears to be distinct from Double Strand Break (DSB) repair and is in part due to the prevention of MRE11-mediated degradation of nascent DNA at stalled forks. The role of RAD51 in fork protection resembles the activity described for its prokaryotic ortholog RecA, which prevents nuclease-mediated degradation of DNA and promotes replication fork restart in cells challenged by DNA damaging agents...
January 12, 2017: FEBS Letters
https://www.readbyqxmd.com/read/28077781/effects-of-an-unusual-poison-identify-a-lifespan-role-for-topoisomerase-2-in-saccharomyces-cerevisiae
#12
Gregory Tombline, Jonathan I Millen, Bogdan Polevoda, Matan Rapaport, Bonnie Baxter, Michael Van Meter, Matthew Gilbertson, Joe Madrey, Gary A Piazza, Lynn Rasmussen, Krister Wennerberg, E Lucile White, John L Nitiss, David S Goldfarb
A progressive loss of genome maintenance has been implicated as both a cause and consequence of aging. Here we present evidence supporting the hypothesis that an age-associated decay in genome maintenance promotes aging in Saccharomyces cerevisiae (yeast) due to an inability to sense or repair DNA damage by topoisomerase 2 (yTop2). We describe the characterization of LS1, identified in a high throughput screen for small molecules that shorten the replicative lifespan of yeast. LS1 accelerates aging without affecting proliferative growth or viability...
January 5, 2017: Aging
https://www.readbyqxmd.com/read/28077631/vaccine-efficacy-of-an-inactivated-chimeric-ha-h9-h5n2-avian-influenza-virus-and-its-suitability-for-the-marker-vaccine-strategy
#13
Se Mi Kim, Young-Il Kim, Su-Jin Park, Eun-Ha Kim, Hyeok-Il Kwon, Young-Jae Si, In-Won Lee, Min-Suk Song, Young Ki Choi
: In order to produce a dual effective vaccine against H9 and H5 avian influenza viruses that aligns with the DIVA (differentiating infected from vaccinated animals) strategy, we generated a chimeric H9/H5N2 recombinant vaccine which expressed the whole HA1 region of A/CK/Korea/04163/04 H9N2 and HA2 region of recent HPAI A/MD/Korea/W452/14 H5N8 viruses. The Chimeric H9/H5N2 virus showed similar in vitro and in vivo growth properties and virulence to the LPAI H9 influenza virus. An inactivated vaccine based on this chimeric virus induced serum neutralizing (SN) antibodies against both H9 and H5 viruses, but only induced cross-reactive hemmaglutination inhibition (HI) antibody against H9 viruses...
January 11, 2017: Journal of Virology
https://www.readbyqxmd.com/read/28076794/brca1-directs-the-repair-pathway-to-homologous-recombination-by-promoting-53bp1-dephosphorylation
#14
Mayu Isono, Atsuko Niimi, Takahiro Oike, Yoshihiko Hagiwara, Hiro Sato, Ryota Sekine, Yukari Yoshida, Shin-Ya Isobe, Chikashi Obuse, Ryotaro Nishi, Elena Petricci, Shinichiro Nakada, Takashi Nakano, Atsushi Shibata
BRCA1 promotes homologous recombination (HR) by activating DNA-end resection. By contrast, 53BP1 forms a barrier that inhibits DNA-end resection. Here, we show that BRCA1 promotes DNA-end resection by relieving the 53BP1-dependent barrier. We show that 53BP1 is phosphorylated by ATM in S/G2 phase, promoting RIF1 recruitment, which inhibits resection. 53BP1 is promptly dephosphorylated and RIF1 released, despite remaining unrepaired DNA double-strand breaks (DSBs). When resection is impaired by CtIP/MRE11 endonuclease inhibition, 53BP1 phosphorylation and RIF1 are sustained due to ongoing ATM signaling...
January 10, 2017: Cell Reports
https://www.readbyqxmd.com/read/28073912/auxiliary-metabolic-genes-distinct-features-of-cyanophage-encoded-t-type-phycobiliprotein-lyase-%C3%AE-cpet
#15
Raphael Gasper, Julia Schwach, Jana Hartmann, Andrea Holtkamp, Jessica Wiethaus, Natascha Riedel, Eckhard Hofmann, Nicole Frankenberg-Dinkel
Auxiliary metabolic genes (AMG) are commonly found in the genomes of phages that infect cyanobacteria and increase cyanophage's fitness. AMGs are often homologs of host genes, and also typically related to photosynthesis. For example, the θcpeT gene in the cyanophage P-HM1 encodes a putative phycobiliprotein lyase related to cyanobacterial T-type lyases, which facilitate attachment of linear tetrapyrrole chromophores to Cys-155 of phycobiliprotein β-subunits, suggesting that θCpeT may also help assemble light-harvesting phycobiliproteins during infection...
January 10, 2017: Journal of Biological Chemistry
https://www.readbyqxmd.com/read/28073364/lack-of-mre11-rad50-nbs1-mrn-complex-detection-occurs-frequently-in-low-grade-epithelial-ovarian-cancer
#16
Simone Brandt, Eleftherios P Samartzis, Anne-Katrin Zimmermann, Daniel Fink, Holger Moch, Aurelia Noske, Konstantin J Dedes
BACKGROUND: BRCA1/2-deficient ovarian carcinomas are recognized as target for Poly (ADP-ribose) polymerase (PARP) inhibitors. BRCA1 and BRCA2 proteins are involved in homologous recombination repair of double-strand DNA breaks. The relevance of other homologous recombination repair proteins, e.g. MRE11, RAD50, NBS1 (MRN complex) in ovarian carcinomas is unclear. The objective of this study was to investigate the prevalence of lack of MRE11, RAD50, NBS1 protein detection in epithelial ovarian cancer (EOC)...
January 10, 2017: BMC Cancer
https://www.readbyqxmd.com/read/28073246/a-new-era-of-genome-integration-simply-cut-and-paste
#17
Zihe Liu, Youyun Liang, Ee-Lui Ang, Huimin Zhao
Genome integration is a powerful tool in both basic and applied biological research. However, traditional genome integration, which is typically mediated by homologous recombination, has been constrained by low efficiencies and limited host range. In recent years, the emergence of homing endonucleases and programmable nucleases has greatly enhanced integration efficiencies and allowed alternative integration mechanisms such as non-homologous end joining and microhomology-mediated end joining, enabling integration in hosts deficient in homologous recombination...
January 10, 2017: ACS Synthetic Biology
https://www.readbyqxmd.com/read/28070534/design-and-construction-of-a-cloning-vector-containing-the-hspx-gene-of-mycobacterium-tuberculosis
#18
Yaghoubi Atieh, Aryan Ehsan, Zare Hosna, Alami Shadi, Teimourpour Roghayeh, Meshkat Zahra
BACKGROUND: Tuberculosis (TB) is a major cause of death worldwide. Finding an effective vaccine against TB is the best way to control it. Several vaccines against this disease have been developed but none are completely protective. The aim of this study was to design and construct a cloning vector containing the Mycobacterium tuberculosis (M. tuberculosis) heat shock protein X (hspX). METHODS: First, an hspX fragment was amplified by PCR and cloned into plasmid pcDNA3...
October 2016: Reports of Biochemistry & Molecular Biology
https://www.readbyqxmd.com/read/28069956/shuttling-along-dna-and-directed-processing-of-d-loops-by-recq-helicase-support-quality-control-of-homologous-recombination
#19
Gábor M Harami, Yeonee Seol, Junghoon In, Veronika Ferencziová, Máté Martina, Máté Gyimesi, Kata Sarlós, Zoltán J Kovács, Nikolett T Nagy, Yuze Sun, Tibor Vellai, Keir C Neuman, Mihály Kovács
Cells must continuously repair inevitable DNA damage while avoiding the deleterious consequences of imprecise repair. Distinction between legitimate and illegitimate repair processes is thought to be achieved in part through differential recognition and processing of specific noncanonical DNA structures, although the mechanistic basis of discrimination remains poorly defined. Here, we show that Escherichia coli RecQ, a central DNA recombination and repair enzyme, exhibits differential processing of DNA substrates based on their geometry and structure...
January 9, 2017: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/28069867/the-carboxyl-terminal-region-of-dok-7-plays-a-key-but-not-essential-role-in-activation-of-muscle-specific-receptor-kinase-musk-and-neuromuscular-synapse-formation
#20
Ryo Ueta, Tohru Tezuka, Yosuke Izawa, Sadanori Miyoshi, Satoru Nagatoishi, Kouhei Tsumoto, Yuji Yamanashi
As the synapse between a motor neuron and skeletal muscle, the neuromuscular junction (NMJ) is required for muscle contraction. The formation and maintenance of NMJs are controlled by the muscle-specific receptor kinase MuSK. Dok-7 is the essential cytoplasmic activator of MuSK, and indeed mice lacking Dok-7 form no NMJs. Moreover, DOK7 gene mutations underlie DOK7 myasthenia, an NMJ synaptopathy. Previously, we failed to detect MuSK activation in myotubes by Dok-7 mutated in the N-terminal pleckstrin homology (PH) or phosphotyrosine binding (PTB) domain or that lacked the C-terminal region (Dok-7-ΔC)...
January 7, 2017: Journal of Biochemistry
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